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This tribute is dedicated to Sidney (Sid) Altman, one of the veritable architects of modern RNA research. Sid is widely known as the co-recipient of the 1989 Nobel Prize in Chemistry, an honor shared with Tom Cech for the discovery of catalytic RNAs. However, in addition to being a distinguished scientist, the collection of essays in this Gedenkschrift highlight Sid's expertise in research and administration, breadth of advising, and humanism. A biography and an interview transcript bookend the writings by some of his trainees, colleagues at Yale, and friends at large. This collective ink celebrates Sid's outsized influence on all the lives he touched. We hope this tribute will inspire all readers.
RNA. 2022 Nov;28(11):1393–1429.
SIDNEY ALTMAN: AN EXEMPLARY SCIENTIST, TEACHER, AND FRIEND (1939–2022)
(Left) Press conference at the Kline Biology Tower, Yale University, on October 12, 1989, following the announcement by the Royal Swedish Academy of Sciences. (Left to right) Donna Wesolowski, Heidi Gold, Cecilia Guerrier-Takada, Sidney Altman, Ania Knapp, Karen Peck-Miller, and Nadya Lumelsky. (Right) Some alumni members of the Altman Laboratory gathered in New Haven to commemorate Sid's 60th birthday (August 1999). The flags of countries of the participants were placed at the podium. The international makeup of his team was a great source of pride for Sid.
(Photos courtesy of Dr. John Arnez.)
Note:Quotations attributed to Sidney Altman himself (in italics) are drawn from his articles, emails, and unpublished memoir.
The human stories that undergird major scientific advances are dissimilar tapestries, each showcasing a different interweaving of individual values, research mentors, environments, and serendipity. The life of Sidney (Sid) Altman is no exception. From modest beginnings, he embarked on a challenging and exciting odyssey that now is an important chapter in the history of modern biology. His discovery of a catalytic role for RNA, a finding also made independently by Tom Cech, upended the longstanding view that the ability to accelerate chemical reactions is the sole realm of proteins. Through their landmark findings, Altman and Cech inspired the identification of unexpected and novel roles for RNAs (beyond their role as information carriers) and the pursuit of RNAs as tools, drug targets, and drugs/vaccines.
Early life: Sid was born in Montreal, Canada, in May 1939, to Jewish émigré parents. Almost 20 years before Sid's birth, both his parents had migrated to Canada: his mother Ray Arlin from Białystok, Poland, and his father Victor Altman from Chornyi Ostriv, Ukraine. Ray, one of eleven children born to a Talmudic scholar and a homemaker, worked initially in a textile mill in Montreal. Victor, a penniless immigrant, spent long days as a laborer on a collective farm in Ontario, a requirement imposed by the Jewish agency that sponsored his immigration to Canada. In 1930, Victor soon had his own grocery store in Montreal's west end district of Notre-Dame-de-Grâce and had married Ray. Sid's father worked very hard to support his family and to save for his children's education. His parents’ efforts left Sid with an enduring conviction in the importance of hard work, a belief that he unfailingly emphasized to those who sought his counsel. “It was from [my parents] that I learned that hard work in stable surroundings could yield rewards, even if only in infinitesimally small increments.”
Even though neither of them had completed high school, Victor and Ray placed a high value on the education of their two sons (Sid was predeceased by an older brother, Marvin). At home, they drew from three cultures and languages (Yiddish, Russian, and English) as sources of parental advice. For his formal K-12 education, they sent Sid to Montreal public schools. To ensure his fluency and pronunciation in French, his parents arranged for a special tutor (Madame Gendron, for whom Sid retained a lifelong affection) to supplement the French classes at school. Sid greatly enjoyed these private lessons since the discussions of France and French culture were quite unlike those in his school textbooks. He also attended Hebrew school on Sundays, winning book prizes that he retained for his entire life. Sid could sight-read music and play the piano, having been trained by a sequence of tutors, many of whom were post-World War II refugees that his parents wanted to support. He maintained a lifelong interest in classical music, with Mozart's clarinet concerto (K.622) being a strong favorite.
Higher education and research career: In May 1960, Sid received a BS from MIT, with a major in physics. He had worked with Lee Grodzins, a new faculty member, to obtain experimental support for the nonconservation of parity. Sid's undergraduate research thesis, which was dedicated to his parents, showed that electrons decaying from 90Sr are polarized and that there is asymmetry in the number of electrons emitted, relative to the direction of the spin (more electrons were observed opposite the direction of the spin). Sid attributed this rewarding bench experience to Grodzins’ approach: He would propose a problem and wait for Sid's solution before stepping in with advice. Sid cherished this training philosophy and always acknowledged his debt to Grodzins, who guided his first foray into research.
Sid joined the doctoral program in physics at Columbia but left when he was unable to find an advisor. By his own account, he quit before he would have been expelled! He then enrolled as a summer student at the University of Colorado in Boulder, where a chance encounter with George Gamow, a famous physicist and Russian émigré, led Sid to consider the graduate program in Biophysics at the University of Colorado Medical School in Denver.
Sid was soon engaged in his doctoral research in Denver with Leonard Lerman (who was, incidentally, born in the United States to Jewish immigrants from Ukraine). Lerman had recently returned from the Medical Research Council Laboratory of Molecular Biology (MRC LMB) in Cambridge, England, having completed a sabbatical with Sydney Brenner and Francis Crick. Lerman had generated mutations in DNA using acridine dyes, whose planar structures enable their intercalation with DNA. For his doctoral dissertation, Sid examined the effects of 9-aminoacridine on replication of the bacteriophage T4 and identified distinct effects on both the replication and the packaging of DNA. During his years in the Lerman laboratory, Sid taught himself molecular biology and sharpened his independent research skills. A tribute to Lerman by Sid and Tom Maniatis (another of Lerman's doctoral students), includes the following paragraph: “[Lerman] demanded rigorous and independent thinking, and a thorough grasp of scientific principles. In some ways, his method of teaching was Socratic, as the questions we asked were usually answered by another question, which probed our depth of understanding and thinking. A discussion of how we might attack the problem at hand would follow, usually leading to his comment that ‘You decide!’ While this approach could sometimes lead to despair, on many occasions it resulted in the conception of an independent study, a new direction …”
Lerman moved to Vanderbilt and Sid accompanied him to complete his research project. Then, at a conference in Tennessee, Sid met Matt Meselson and landed his first postdoctoral position in Meselson's laboratory at Harvard. Meselson told Sid that he would need to find his own funding and, with a strong recommendation from Lerman, Sid was awarded a Damon Runyon Fellowship. At Harvard, Sid demonstrated that expression of a previously unidentified endonuclease was induced in T4-infected Escherichia coli, with resultant single-strand breaks in the bacteriophage DNA. Sid's heart was, however, somewhere else entirely—not in Cambridge, Massachusetts but in Cambridge, England. Since the early 1960s, when he had first read Sydney Brenner's pioneering work on RNA, Sid had aspired to work with him, a sentiment amplified severalfold after listening to a spectacular lecture by Brenner. Toward the end of his postdoctoral appointment at Harvard, Sid met Brenner in Boston and was excited to be offered an opportunity, at the LMB in Cambridge, to use NMR to study the structure of tRNA. Sid was delighted because, previously, Brenner had told him that there was no available bench space at the LMB. Sid would later remark that his sojourn at the LMB was a fabulous time when he got to observe at close quarters the penetrating intelligence of an amazing group of scientists (including Fred Sanger).
By the time Sid arrived at the LMB as an Anna Fuller Fund fellow, in October 1969, advances in tRNA crystallization had rendered moot any NMR-based approach to the determination of the structure of tRNA. So, when Brenner and Francis Crick asked Sid to come up with a different research plan, he proposed the isolation of acridine-induced mutants of tRNATyr. Sid's treatment of E. coli with a potent acridine half-mustard compound led to the identification of a mutant tRNA that was much longer than any mature tRNA. He purified the radiolabeled tRNATyr precursor and determined its nucleotide sequence.
While we now appreciate that many mature RNAs are first produced as longer precursors, Sid's early findings spawned numerous studies of bacterial RNA processing. Because the mutant tRNA contained more nucleotide residues than the mature tRNA, Sid reasoned that it was probably a precursor that could not be processed by the ribonucleases normally involved in 5′ and 3′ trimming. With the help of Hugh Robertson, who was also a postdoctoral fellow at the LMB and who, along with William (Bill) McClain, would become a close friend, the enzymatic activity responsible for releasing the 5′ leader was isolated from E. coli. The enzyme was named RNase P (with P referring to precursor). In their 1971 JBC paper, Sid and Hugh noted that there was a large amount of UV-absorbing material in the peak fractions. A forerunner of what was to come!
Sid's work at the LMB led to his appointment as an Assistant Professor at Yale University, where his biochemical characterization of bacterial RNase P began in earnest. To the intense frustration of those involved in these efforts, a pesky, copurifying RNA appeared in all the fractions with RNase P activity. Sid tasked Ben Stark, a meticulous graduate student, to get rid of the RNA and focus on the protein responsible for the processing of the pre-tRNA. Ben's painstaking experiments convinced his skeptical advisor of the inescapable requirement for RNA in the cleavage activity of RNase P (a ribonucleoprotein). When Sid was granted tenure in 1977, he wrote to Sydney Brenner thanking him for his support and acknowledging his memorable stay at the LMB. He ended the letter with “You may be interested to know that RNase P has an RNA moiety, i.e., at least one or possibly two discrete RNA species are essential for the function of the enzyme.”
The 1978 PNAS paper, in which Sid and his coworkers posited that the RNA in RNase P might be playing a supporting role in the catalytic activity of RNase P, was very different from the 1983 Cell paper, in which the RNA moiety was demonstrated to be the catalytic subunit of RNase P! Within five short years, there was a sea change. The first signs in this difficult journey came from the painstaking biochemical studies performed by Ryszard Kole, a Polish postdoctoral fellow in Sid's laboratory. By separating the RNA from the protein component of E. coli RNase P, Ryszard showed that the protein (contrary to dogma) lacked pre-tRNA cleavage activity, but the combination of the RNA and the protein resulted in activity. This finding led Kole and Altman to state in their 1981 Biochemistry paper, “The possibility that the [RNase P] RNA molecule participates in the formation of an active site of an enzyme appears novel.” Subsequently, in the fall of 1983, during reconstitution of the respective RNA and protein subunits of E. coli and Bacillus subtilis RNase P (in collaboration with Norman Pace's laboratory), Cecilia Guerrier-Takada, an exceptional postdoctoral scientist in Sid's laboratory, found that each RNA subunit by itself had catalytic activity in the presence of 60 mM Mg2+, a condition that had not previously been tested! Soon afterwards (Guerrier-Takada and Altman 1984), Cecilia showed that the E. coli RNase P RNA, synthesized by in vitro transcription, could catalyze the cleavage reaction, firmly ruling out any possibility that contaminating protein(s) might have been responsible for cleavage activity. These experiments were only possible because the gene for the RNA had been cloned and sequenced, a notable accomplishment in the early 1980s by Robin Reed, a graduate student in the Altman laboratory.
Sid was willing to challenge accepted dogma, albeit very cautiously. However, he had tremendous difficulty in convincing others of his discovery. Tom Cech's independent discovery of the self-splicing group I intron of Tetrahymena helped decisively to turn the tide in their collective favor. Sid always felt that the scientific community had shown its best and worst sides during this trying period. He gained much from the support of his family, colleagues, and mentors. Sid recounted a conversation with Matt Meselson in which he shared his uphill battles with the doubters. Meselson's response was, “If you have done your experiments correctly and you believe in your results, then you have no choice but to trust what nature is telling you. Sooner or later, the truth will be apparent.” These remarks boosted Sid during some of his darkest hours. While time tends to soften the rough edges of all human engagements, Sid felt, even during his later years, that he had not been treated fairly in the late 1970s and early 1980s.
During his career, Sid made more than a few serendipitous choices that endowed him with expertise that paid handsome dividends later. For example, the chance meeting with Gamow that led to Lerman and the abiding interest in acridines, which indirectly paved the way to his work on RNase P. However, there was nothing fortuitous about the discovery of RNA as the catalytic component of RNase P. Sid and his coworkers were tenacious and single-minded experimentalists who drilled deep to uncover one of Mother Nature's fascinating secrets. The independent advances made by Tom Cech and his team were equally impressive. Sid and Tom Cech, joined soon by many other investigators, began to unravel the biochemistry of RNA catalysis. RNA enzymes were found to be not so different from proteinaceous enzymes insofar as they are modular, they exploit acid-base chemistry and metal ions, and both exploit a number of weak interactions to promote formation of the enzyme-substrate complex. This was the dawn of a new era in biocatalysis!
Advising: “I am forever grateful to my mentors during my career. Perhaps, I was able to repay that enduring gift with my postdocs and students.” It is true that the apple does not fall far from the tree and this adage applies to scientific progeny as much as to parents and children. I have many scientific siblings from Sid's laboratory at Yale University (51 were postdoctoral fellows, 15 were doctoral students, and 22 were undergraduate trainees). Some of my “siblings” have contributed their rich and varied perspectives to this volume. Adding to their contributions, here are some of Sid's attributes that, I believe, were universally appreciated: a strong work ethic, integrity, intellectual curiosity, and inclusiveness. He was a positive role model and his norms were embraced by his trainees and colleagues.
Sid believed that scholarship was fruitful when accompanied by perseverance and hard work. “It could be a letter carrier, it could be a sales manager in a store or a university professor, all the people who do well work very hard. Nobody who has a record of achievement has been lazy about it.” Moreover, he felt that difficult problems were solved only by obsessive effort. “It is the intense mental preoccupation with a problem that is a major factor: seeing it from every conceivable point of view; examining options for solutions and choosing among them. This scrutiny must encompass every detail, winnowing out those that can lead to complex, fruitless pursuits.” These traits, when complemented by sincerity of purpose, ensured rigor and quality in all his pursuits.
For Sid, integrity meant incorruptibility of the scientific enterprise. He chose to be a scientist because he believed that research endeavors were completely objective (see article by Julius Lucks and Venkat Gopalan in this tribute). Sid was cautious by nature but also open-minded. One might argue that Sid was initially resistant to his students’ ideas (e.g., a catalytic role for RNA, see article by Ben Stark), but he always looked for a chain of reasonable inferences that was based on strong experimental evidence. He was averse to any overstatement. While his conservative approach did, at times, dilute the importance of some claims, Sid despised showboating above all. In fact, as I know only too well, he swiftly consigned to the garbage can any expansive and speculative sentences in the Discussion sections of a manuscript draft.
Sid was a true scholar who read broadly and was an excellent writer. His interests spanned art, music, and history. He would surprise me with emails that included specific inquiries (e.g., a question about the Brahmo Samaj movement in India in the early 1800s) or descriptions of the inadequacy of a museum exhibit on British India that failed to convey the mores and lives of the locals. He had a keen eye for detail and could converse on varied topics in many fields. It was stimulating to be in his company. As Dean of Yale College in the mid-1980s, Sid provided stewardship during the revision of the undergraduate curriculum, ensuring that science majors would gain an appreciation of the arts and humanities and, reciprocally, that non-scientists would recognize the value of science.
Every research team grows naturally, but hidden beacons often guide the leader in the development of her/his group. Sid showed, by example, how to implement an inclusive culture that thrived on intellectual ferment. In this regard, two factors might have influenced him. First and foremost, Sid's time at the LMB with Sydney Brenner showed him that dedication to research was a paramount requirement. Gender, age, nationality, and any other classification were unimportant. Reflecting his experiences at the LMB, he valued the opinions and initiatives of all those who worked with him. Over a 40-year period, his laboratory had many female scientists and technicians and Sid had the highest respect for all of them and their contributions (in particular, Dr. Cecilia Guerrier-Takada; see photo, left). Second, he never forgot that his parents had been poor immigrants to Canada, a country that offered them opportunities for growth and enrichment in a stable and safe environment. Thus, he had a particularly soft spot for immigrants (see photo, right). Foreigners typically outnumbered US citizens in his group. On the occasion of his 70th birthday, some international alumni (myself included) dedicated some review articles on RNase P to Sid. He responded: “I am personally grateful to you for the friendship… You represent six different countries. For me, there is no greater satisfaction from my professional life than to have done a little bit in helping you progress from your homes around the globe to the respected positions that you have now. From one immigrant to the others, thank you.”
Individuality: The superficial appraisal of any public (or private) figure is invariably misleading because the paradoxes in all our personalities are either ignored or glossed over. Sid was viewed by some as overbearing and impatient. Such characterizations are incorrect.
Unwelcome labels prompted by Sid's bluntness and independence overlook the primacy that he placed on honesty and adherence to principles. Nuances were few and far between. However, he never indulged in self-importance and did not like others to fuss over him. He acknowledged his own failings with great humility (see Sid's letter in Ben Stark's article in this tribute). In fact, Sid's unmistakable wit was most apparent when he was self-deprecatory. For example, motivated by his desire to be an author in his late teens, he boldly approached the Arts and Book review editor of the Montreal Star expressing his interest in writing for the newspaper. When Sid returned with a sample book review that the editor had requested, the editor advised, “If you have a talent for science, stick to it while you are young.” Sid concluded that the editor “was very perceptive, or very wise, or very diplomatic, or all three”! Another example of Sid's self-deprecatory wit is obvious from his description of how Sydney Brenner and Francis Crick responded to his idea of studying acridine-induced mutants of tRNAs. Upon sensing their ennui after hearing his idea, Sid wrote that he “distinctly felt that if they spoke to me again, it would be an accident.”
Sid was impatient with anyone who he felt impeded the research that he held dear, but his vast reserves of patience were amply evident when he waited (even for years!) for the completion of difficult research objectives. Recognizing that “work on RNase P was usually a three- to four-year period of boredom and fright,” he was quite tolerant of research excursions from which nothing of any apparent significance emerged.
Sid was a private person, and he was often brusque. It was easy to conclude, from his façade, that Sid was distant. However, such a conclusion is at odds with his devoted friendship to those who knew him well. Whether it was sending a beautiful bouquet on my wedding day, a thoughtful greeting on my first Father's Day, or a poignant condolence note on my loss of a close family member, he consistently showed uncommon grace and caring. Such warmth and goodwill endure.
Sid was never aloof. To the contrary, he acted swiftly when he identified a good reason for doing so. His strong convictions shaped his public stance on many issues. For example, Sid was one of the signatories supporting the Precision Agriculture (GMO) Campaign, largely because he believed that biotechnology could enhance food security and help eliminate poverty. Also, he was a cosignatory, with many other leading scientists, in condemning the international boycott of Israeli academics. He was a vocal proponent of academic and scientific freedom.
Envoi: Numerous trainees and a legion of close friends and colleagues had the privilege of working with Sid. While some of their voices in this tribute are merely a whisper from this shared past, they provide a compelling and collective narrative for those who did not know Sid, and they reveal a portrait of the man as an exemplary scientist, conscientious teacher, and loyal friend. Varied facets of Sid's personality emerge under these different lenses, each tempered by deep reflection over time. An inescapable conclusion from these wide-ranging personal remembrances is that Sid had a profound influence on all the lives he touched. By evoking memories that celebrate the richness of Sid's life, the following essays—including some by those who knew him personally or professionally for nearly half a century—show why his legacy will live forever.
Not marble nor the gilded monuments
Of princes shall outlive this powerful rhyme,
But you shall shine more bright in these contents
Than unswept stone besmeared with sluttish time.
When wasteful war shall statues overturn,
And broils root out the work of masonry,
Nor Mars his sword nor war's quick fire shall burn
The living record of your memory.
‘Gainst death and all-oblivious enmity
Shall you pace forth; your praise shall still find room
Even in the eyes of all posterity
That wear this world out to the ending doom.
(Shakespeare, Sonnet 55)
Footnotes
Venkat Gopalan is extremely grateful to Ann Altman for editing and feedback.
REFERENCES
Guerrier-Takada C, Gardiner K, Marsh T, Pace N, Altman S. 1983. The RNA moiety of ribonuclease P is the catalytic subunit of the enzyme. Cell
35: 849–857. 10.1016/0092-8674(83)90117-4 [DOI] [PubMed] [Google Scholar]
Guerrier-Takada C, Altman S. 1984. Catalytic activity of an RNA molecule prepared by transcription in vitro. Science
223: 285–286. 10.1126/science.6199841 [DOI] [PubMed] [Google Scholar]
Stark BC, Kole R, Bowman EJ, Altman S. 1978. Ribonuclease P: an enzyme with an essential RNA component. Proc Natl Acad Sci
75: 3717–3721. 10.1073/Pnas.75.8.3717 [DOI] [PMC free article] [PubMed] [Google Scholar]
In 1971, I was a graduate student in Dieter Söll's laboratory, at Yale University, when a parade of postdoctoral fellows from the Medical Research Council Laboratories in Cambridge, England, came to Yale to give job seminars. I remember Peter Lengyel introducing a radiant Joan Steitz with his inimitable Hungarian charm, as well as cutting-edge seminars by gifted young men, such as Hugh Robertson, Sam Ward, and Sidney Altman himself.
In the fall of that same year, Sidney, a successful candidate, set up his laboratory on the eighth floor of Kline Biology Tower, six floors above the Söll laboratory. One thing led to another and, in 1972, Sidney and I were married, as he continued to lead his group and I continued the research that would lead to my doctorate in molecular biophysics and biochemistry.
Inevitably, I got to know Sidney's graduate students, in particular, Ben Stark and Al Bothwell. I also got to know the postdoctoral fellows, often meeting them just after their arrival and helping them to get settled. In particular, I remember Ryszard Kole arriving with his wife Jola and just two suitcases; I remember taking Sergei Kazakov to the local inexpensive department store and being somewhat ashamed that there was more to buy in this one store in Connecticut than in all of Novosibirsk.
Over the years, Sidney welcomed young postdoctoral fellows from all over the world, for example, from China, Russia, India, South Korea, Spain, Italy, and Sweden. In my view, the subsequent success of all these young people is Sidney's greatest achievement. And this tribute is a reflection of that success. Sidney's scientific family tree (https://academictree.org/chemistry/tree.php?pid=52166), which is the tree to which all his students and postdoctoral fellows belong, includes Niels Bohr, Erwin Schrödinger, Ernest Rutherford, Linus Pauling, and Sydney Brenner. The additions to this scientific family tree that reflect Sidney's mentorship are his most fitting memorial.
RNA. 2022 Nov;28(11):1393–1429.
A SHORT HISTORY OF THE DISCOVERY OF THE ESSENTIAL RNA COMPONENT OF RNase P1
(Left) The Altman lab in 1974. Seated left to right: Cris Mickiewicz, Lois Atkins, Sheldon Feinstein, Rick Garber, Ray Koski, Ginger Warner, and Paolo Arcari. Standing left to right: Al Bothwell, Frank Costantini, Sid, me, and Bob Geggle. (Right) A letter sent to me by Sid from Bristol, England in 1978.
(Photos courtesy of Benjamin Stark.)
The story starts in 1973. In the fall of 1972 (my second year as a graduate student in the Biology Department at Yale), I had joined Sid Altman's laboratory and in my first semester I had worked on a small project with tRNA precursors that was modestly successful. In 1973, I began my main thesis project to purify to homogeneity RNase P from Escherichia coli (the organism in which Sid and his early collaborators had first found it) and characterize it. At that time, this undertaking was a fairly typical and reasonable project in molecular biology, although in this instance made challenging by the cumbersome nature of the RNase P assay. This assay required a polyacrylamide gel and a particular tRNA precursor that I had to make about each month from an in vivo labeling with large amounts of P-32. Nevertheless, this part of the project moved along pretty well. More demanding was the purification itself. In retrospect, of course, it is easy to see why it was so difficult. I was using standard procedures to purify a protein enzyme, when in fact I was purifying an enzyme that is, by mass, ∼90% RNA and ∼10% protein.
By scaling up the purification procedure, I was able to prepare large quantities of partially purified enzyme. However, additional ion-exchange chromatographic steps did not purify the enzyme further, because RNase P and the contaminants had roughly the same (negative) charge. So, I explored alternative purification steps and decided on preparative electrophoresis using “native” gels, despite the poor yield from this extremely tedious approach. Consequently, when the RNase P-containing fractions from the native gel elution were assessed by SDS-PAGE and Coomassie blue staining (remember, I was still looking for a protein), I often noticed very faint bands or nothing at all.
At this time Paolo Arcari, a postdoctoral fellow in Sid's laboratory (see photo of Sid's group in 1974), was using methylene blue to stain RNA in PAGE gels. Reasoning that negatively charged proteins in my peak fractions may not be binding Coomassie blue, I began in January 1975 to take SDS-polyacrylamide gels that had been stained with Coomassie and “overstain” them with methylene blue. When these overstained gels were destained, there were several new bands including two prominent bands in the gels which contained the peak activity fractions from the native gel purification step. Two of these were called M1 and M2 (I termed them as such in May-June 1975). Likewise, the Coomassie-stained bands in the same fraction were denoted C1–C6. In the cleanest RNase P preparations, there were only C5 and M1 and M2 in the peak fraction (see Fig. 1B,C of the 1978 paper in PNAS [Stark et al. 1978]). As it turned out, these components are the components of native RNase P. C5 is the protein component and M1 and M2 are the RNA components; M1 has been extensively studied and since shown to be the catalytic part of the enzyme while the function of M2 is still unknown.
Initially, I thought M1 and M2 really were proteins. It fit well with the ideas I had at that time because M1 and M2 (if they had been proteins) were measured on my gels with molecular weights of about 60,000 and 70,000 Da, respectively (their actual sizes are larger because RNA and protein molecules do not run the same in this gel system). A tetramer of two copies of M1 and two copies of M2 would have had a molecular weight of 260,000 Da, almost exactly what I had measured for native RNase P by gel filtration in other experiments. It would also parallel the “alpha 2-beta 2 structure” known to exist for many tRNA synthetases, enzymes which akin to RNase P interact with tRNAs.
Of course, it was a formal possibility that M1 and M2 were RNAs, but for a long time the simplicity of the idea that they were proteins and the model described above overruled my common sense. It would have been very easy to determine whether M1 and M2 were RNAs or proteins, but I waited a long time to test this (see below). In fact, I was so sure of the idea that methylene blue could stain certain (acidic) proteins that I even tested this with T1 ribonuclease (a very acidic protein). Methylene blue staining of T1 ribonuclease on SDS gels, however, was poor. I should have seen the weakness in my argument right away, but I did not.
A few months later (fall 1975), I was talking with Skip Binder (another graduate student in our department) about whether M1 and M2 might be RNAs. He suggested that I just go ahead and test my idea directly. It was actually a very easy experiment. It entailed treating native gel-purified RNase P fractions with ribonuclease before running the samples on SDS gels to determine if M1 and M2 disappeared. The first such experiment, according to my notebook, was in November 1975. The experiment did not give clear results, but I continued to refine it. The first experiments that showed clearly that M1 and M2 are RNAs and not proteins were done in late March 1976. Eventually, as I repeated the experiment more and refined it more, it was clear that this result was fully reproducible. When I examined a large pile of photos of SDS gels from RNase P purifications, M1 and M2 were always there. Furthermore, they were most intense in the peak RNase P activity fractions, purified along with the RNase P activity, and their intensity in various fractions nearly paralleled that of RNase P activity in these fractions. I became convinced that M1 and M2 were part of RNase P.
Obviously in 1975–1976, the idea that an enzyme was not protein was absolute heresy. I had never heard of anything like this myself, and, of course, the dogma maintained that this notion was crazy. Unknown to me (I found these papers sometime later), there had been at least three previous reports of ribonucleoprotein enzymes in the literature. When I proposed this idea to Sid, he was against it. At the time, I think he saw me as a very green, though hard-working, graduate student who was not very successful in his PhD research. I did not know what to do. I was convinced I was on to something, but Sid did not want me to pursue it thinking it was a crazy idea by someone who was a bit too naïve. I am afraid I got to whining about this situation quite a bit, largely to Sheldon Feinstein, my lab partner, fellow graduate student, and close friend (see his contribution in this tribute). In October 1976, Sheldon had had enough of my complaining and he told me that, if I felt that strongly about the RNA nature of RNase P, I should have a meeting of my committee and try to convince them that I should devise a way to test this idea. Actually, I did have an idea of how to go about this (see below), but it was not a very good one.
It must have been around October 6, 1976 that my meeting was held. My committee consisted of Sid, Don Crothers (a distinguished member of the Chemistry Department), and Peter Rae (a member of the Biology Department, who was very popular with graduate students). I pitched my idea of the RNA nature of RNase P to the committee. My recollection is that Sid and Don Crothers were negative about this proposal and asked me how I would go about proving my idea. I suggested pretreating my RNase P preparations with a ribonuclease and assessing whether the RNase P activity disappeared. The problem, of course, was that in order to assay my ribonuclease-treated RNase P fraction I would have to use an RNA substrate, and any ribonuclease I had used to degrade the RNase P would also degrade the substrate and render the assay impossible. I proposed to use ribonuclease T1; because of its small size, it might be possible to separate it from RNase P by gel filtration (remember I knew RNase P had a molecular weight of ∼260,000). The technical problems with this approach, however, were daunting, as even modest carryover and contamination with highly active ribonuclease T1 (post-gel filtration) would ruin the assay.
It is here that Peter Rae stepped in. He noted that micrococcal nuclease, an enzyme used, I think, routinely in his lab, would degrade RNA in a calcium-dependent manner. Since RNase P did not require calcium, I could perform the RNase P treatment in the presence of micrococcal nuclease and calcium, then dialyze out the calcium from the digestion mix (relatively easily), add back magnesium to support RNase P activity, and then assay for RNase P activity. The micrococcal nuclease would still contaminate the RNase P but it would not matter, because, in the absence of calcium, the micrococcal nuclease would be inactive and would not interfere with the RNase P assay by degrading the RNA substrate.
According to my notebook, I got some micrococcal nuclease from Tom Barnett, a grad student in Peter Rae's laboratory, on October 7, 1976. Over the next few days, I worked out the conditions for my experiments and these results are shown in Figures 3A and 3B of the 1978 PNAS paper. Sid was still skeptical. But within a few days, controls and repeats showed that the phenomenon was real: RNase P really does have a required RNA component, that is, enzymes do not have to be proteins only. Six months later I defended my thesis. In an initial thesis draft, I had suggested that perhaps the RNA in RNase P was catalytic, but Sid made me exclude that part in favor of the more conservative idea that the RNA moiety of the enzyme may position the tRNA precursor in the active site by complementary base pairing with portions of the substrate.
After my defense, I stayed on in Sid's laboratory for a month or two and continued my work. It was then that I did the first experiments on reconstitution of RNase P from protein and RNA components (which were not convincingly successful). The rest is postscript. Sid took up my cause and made it his cause as well. It seemed as difficult for him to convince the scientific establishment as it had been for me to convince him (see photo of letter). Nature summarily rejected our first manuscript on the topic, later calling it a “quirk” in a News and Views report following the publication of the PNAS paper. And there were other cases dismissing the importance of the discovery as well.
The tide slowly began to turn though. In 1979 came the original work on snRNAs from Joan Steitz's laboratory, and in 1980 a paper from Norm Pace's laboratory showed a required RNA component in RNase P from Bacillus subtilis. Other RNA-containing enzymes were discovered soon after. By 1981–1983 came the discovery of self-splicing of Tetrahymena rRNA precursors by the Cech laboratory and the key discovery that M1 RNA is the catalytic subunit of RNase P by Sid's laboratory; these findings formed the basis for their 1989 Nobel Prize.
When I had my own laboratory, I worked trying to find RNase P in higher plants for a short while. However, circumstances led me in other research directions, which were interesting and fruitful for me. But my RNase P experience in graduate school certainly remains a singular part of my life.
Footnotes
1
Parts of this story have been previously published by Satoshi Ikuta (ISBN 4-534-03513-6) and in reviews by me and Bill McClain, Lien Lai, and Venkat Gopalan. Thanks to Rick Garber, Venkat Gopalan, and Bill McClain for helpful comments.
REFERENCE
Stark BC, Kole R, Bowman EJ, Altman S. 1978. Ribonuclease P: an enzyme with an essential RNA component. Proc Natl Acad Sci
75: 3717–3721. 10.1073/Pnas.75.8.3717 [DOI] [PMC free article] [PubMed] [Google Scholar]
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RECOLLECTIONS OF A GRADUATE STUDENT IN SIDNEY ALTMAN'S LABORATORY
I joined Sidney Altman's laboratory in 1972. Sid (as he wanted us to call him) had recently been recruited by Yale after a postdoctoral fellowship at the MRC Laboratory of Molecular Biology, Cambridge. A nucleus of five new graduate students joined Sid's lab in a short span of two years: Alfred Bothwell joined first, I was next, followed by Benjamin Stark, Richard Garber, and then Raymond Koski. At this beginning stage, the laboratory was mostly male PhD students, although there were female technicians. Subsequently, some female PhD students (Cris Mickiewicz and Janet Emanuel) as well as postdoctoral fellows joined the lab. Sid and the students would occasionally take a break from work and go outside to throw a baseball around for a few minutes. Sid was also a talented hockey player, having grown up in Montreal and had played varsity ice hockey. I remember that on several occasions he arranged for the lab members to attend Yale hockey games at the Ingalls rink. Sid was always interested in sports and this passion showed in these breaks and outings.
When I joined Sid's lab, I was really a novice in biological research. Sid gave me a choice of two projects, one studying ribonuclease P (RNase P) and the other involving nonsense suppressors. I chose the latter, finding it more interesting as it involved deciphering the effect of codon context on translational efficiency. Of course, it turned out that I chose the project that did not lead to the Nobel Prize! In hindsight, I now realize that my research, which sought to tease apart the nexus between codon context and translation, would have been accomplished much more easily and better with recombinant DNA methods, but that was still a few years into the future. However, despite the difficulty of my project, Sid was able to direct my research and, indeed, the research of the early group of five students who successfully completed their dissertations. In retrospect, it is remarkable that Sid, who had not previously directed any PhD student, did so quite well at this early stage in his career.
The RNase P project, which was assigned to Benjamin Stark, turned out to be the one that was most interesting. Ben's job was to purify the enzyme, earning him the nickname “Benzyme.” Ben worked diligently on the purification, the difficulty of which was substantial because assaying the enzyme activity required repeated preparation of a short-lived, radioactive substrate. As Ben continued the purification, the purified enzyme began to take on the absorption spectrum of RNA. At that time it was well-established that enzymes were proteins, so it seemed unlikely that the RNA could actually be part of the enzyme. Instead, the RNA was thought to be an impurity and Sid was questioning Ben's competence! (As an aside, there is an interesting parallel to this in a story that I heard from a postdoc who worked in the laboratory of Arthur Kornberg, another Nobel laureate, on purification of DNA polymerase. The postdoc's task was to remove the “nasty” DNAse activity from the polymerase. He could not accomplish this goal and was nearly thrown out of the Kornberg lab! In the end, it turned out that the DNAse activity was an integral part of the polymerase and provided a proofreading function that lowered the mutagenic rate.) In the case of RNase P, it was difficult to test the function of the RNA in the enzyme, because if an enzyme was used to break down the RNA, that enzyme would interfere with the activity assay by degrading the RNase P substrate. However, Ben did perform some key experiments based on an ingenious suggestion from one of his thesis committee members, Peter Rae (see article by Ben Stark in this tribute). The final step to the Nobel Prize was accomplished by the direct demonstration that the RNA in RNase P, when prepared in vitro, had catalytic activity without assistance from any protein. By the time of this remarkable finding, Ben and I (as well as the initial core of PhD students) had graduated and moved on to the next phases of our careers.
Looking back, I recall that Sid was very analytical and had a great deal of self-confidence, which somehow rubbed off on me, perhaps even before I had any reason to have confidence. His mentoring was key in setting me up for a long career in science. He taught me and other mentees many important lessons, chief among them that unexpected observations can lead to important discoveries, that just because one's research results are met with skepticism does not make them wrong, and that even the greatest scientists are not infallible.
Thanks to Dr. Benjamin C. Stark for reading a draft of this essay and correcting several errors and to Dr. Venkat Gopalan for editing it.
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MEMORIES FROM THE EARLY YEARS OF THE SIDNEY ALTMAN LABORATORY
Two decades after the discovery of the double helix and before the dawn of DNA sequencing, molecular biology was in an exciting age of exploration. We arrived at Yale with the goal of immersing ourselves in molecular biology research. As some of Sid's first graduate students, we were introduced to new methods built on a foundation established by classical bacterial genetics. Sid taught us that rigorous science required passion, concentration, perseverance, and critical review. He enabled us to contribute to the emerging field of RNA–protein interactions and ribonucleoprotein complexes. When one of us (Stark) did the first experiments demonstrating that the RNase P RNA component was essential for catalytic activity, we all realized that RNase P differed from the conventional definitions of enzymes in all textbooks then available (see preceding article by Ben Stark). At monthly lab meetings, during which each of us would present a research article, Sid taught us to critically evaluate its scientific rigor, especially with respect to methods used and the conclusions drawn. These learned skills were later applied by each of us in our respective fields and made us better scientists, clinicians, and educators.
Sid encouraged us to work with eukaryotic systems to further our knowledge of RNA–protein interactions, including those involving mRNAs. This venture into an uncharted area took him out of his comfort zone and involved much intellectual and funding support, which he gladly gave. We all shared the excitement of these early RNA research investigations and witnessed how RNA biology has blossomed to the point of becoming pivotal in the fight against COVID and in other clinical applications. For Sid's stewardship and mentoring, we are therefore eternally grateful.
Sid also displayed his love of sport, particularly ice hockey. There was a custom to give a gag gift to the advisor at the annual departmental holiday party, and one year we gave Sid the “Sidney Cup” in honor of his time on the MIT hockey team! Our recollection was that it was a foil-covered two-liter plastic beaker with a cardboard hockey stick inside! Sid would gladly participate in softball games at our departmental picnics (see photo). During these times he was able to don his more playful side, which we all enjoyed. We cherish these memories.
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THE SPINACH CHLOROPLAST RNase P: THE FIRST PROTEIN-ONLY RNase P!
My first meeting with Sid Altman was in spring 1972 of my senior year at Yale College. He had arrived at Yale the fall semester before. I approached him to inquire about undergraduate research for one semester. I did not work on RNase P, however.
The exposure to RNA biology in Sid's lab, albeit brief, led me to a long and engaging RNA research career, first as a graduate student with David Apirion at the Washington University School of Medicine in St. Louis, then as a postdoctoral fellow in the laboratory of John Abelson at the University of California, San Diego, and finally as a faculty member in the Department of Molecular Biosciences at the University of Kansas. What I did not anticipate was that I would encounter the same difficulties that Sid did, although with an interesting twist!
At the University of Kansas, my graduate students and I set forth to find the chloroplast RNase P protein/RNA complex. It had always been assumed that chloroplast RNase P, like the nuclear cousin, would consist of a protein and an RNA, and that the catalytic subunit would be in the RNA. But we discovered that tRNA 5′ maturation in plant chloroplasts is accomplished by a protein, rather than by a catalytic RNA as in almost all other organisms and domains of life! My team reported this discovery in a 1988 paper in The EMBO Journal. Remarkably, the chloroplast RNase P activity is found at a buoyant density of 1.28 g/mL in CsCl density gradients, coincident with what is expected for protein; for comparison, Escherichia coli RNase P (which Sid studied and where catalysis is due to an RNA) bands as an RNA-rich ribonucleoprotein (RNP) at 1.73 g/mL. The corresponding densities in Cs2SO4 gradients are 1.23 g/mL for protein and 1.55 g/mL for RNA. Furthermore, spinach chloroplast RNase P is resistant to micrococcal nuclease even up to a 50-fold greater amount than those used to fully eliminate activity of E. coli RNase P (see Ben Stark's article in this tribute). My group also showed in a paper published in 1997 that, in E. coli with a GMPαS everywhere instead of GMP (i.e., the RP-thio based substrate), the activity was 28,400-fold worse! In further analysis, the enzyme could act only on the base containing the cleavage site; every other site it is indifferent to. In Mg2+, the enzyme could process only at the position either one base before the correct site (that is, between −2 and −1) or after the site (that is, between +1 and +2). In Mn+2, on the other hand, it processed correctly, between −1 and +1!
In a follow-up paper published in RNA in 2000, my group presented conclusive evidence that chloroplast RNase P differed mechanistically from the RNP form of RNase P. The partially purified chloroplast RNase P can cleave accurately a pre-tRNA with an RP-phosphorothioate substitution at the cleavage site; the rate decreased only by two- to sixfold compared to the unmodified substrate. Clearly, the catalytic mechanism of the organellar enzyme was very different from the bacterial counterpart even though the site of cleavage was the same!
In the fall of 2000, shortly after we published the results presented above, Sid and his colleagues presented a perspective that the organellar enzyme might warrant a different Enzyme Commission nomenclature given the mechanistic differences and likely independent evolutionary origins for the two variants. I contended in my “Divergent View” article, which was published in tandem with Sid's piece, that such a distinction was not necessary and that the same trivial name might be acceptable provided prefixes such as “nuclear,” “organellar,” etc. were used. I also suggested an alternative that the RNP form be referred to as “ribozyme RNase P.” (On a side note, the field has now settled on the PRotein-Only Rnase P [PRORP] nomenclature to distinguish this variant from the RNP form discovered by Sid.)
Back to the organellar variant: While my own laboratory could not clone and identify the make-up of the spinach chloroplast enzyme, Walter Rossmanith and coworkers at the University of Vienna succeeded in characterizing human mitochondrial RNase P and showing persuasively that this activity did not require any RNA but only three proteins; the third protein, MRPP3, was the catalytic subunit and was the homolog of the chloroplast enzyme! Although my work did not lead to the discovery of the spinach chloroplast RNase P protein, I was comforted by the thought that I had certainly been on the right track!
As Sid himself wrote in his “Divergent View” article in RNA, one orthodoxy (only proteins can be enzymes) was replaced by another (RNAs can also catalyze reactions). However, the field soon after adhered to a new dogma that an RNA was necessary in all RNase P variants! For reversing the latter idea with our independent discoveries, both Rossmanith and I were chosen in 2012 to deliver the two inaugural Sidney Altman Endowment Lectures at the XXIV International tRNA Conference in Chile. An unexpected turn of events!
When I met Sid in 2009 at a lab reunion to celebrate his 70th birthday, we discussed many matters including the education of evolution in Kansas. The state had become a hotbed given the hearing on how science should be taught, and I was then on the board of Kansas Citizens for Science! Sid gave me a start on my journey in the land of RNA processing and he was always kind and nice to me; I am forever thankful!
Postscript: In the summer of 2014, I had a stroke due to having arterial tortuosity syndrome, a rare genetic disorder. The stroke has left me in a wheelchair, unable to stand, and with a mild anomic aphasia (a communication disorder). This article was penned together with the help of a colleague.
Sid at the doorsteps of the museum in Athens in June 2008.
(Photo courtesy of Leif A. Kirsebom.)
I first met Sid in late summer 1984 when he, together with his family, visited the Department of Molecular Biology at Uppsala University, where I was doing my PhD. He gave an inspiring presentation during which he discussed his recent and exciting findings on how the RNA subunit of RNase P cleaves precursor tRNA even without the protein cofactor. Visitors typically met with faculty researchers and PhD students in the department. During my discussion about my PhD project with Sid, he mentioned that he had heard that I was looking for a post doctoral position and the discussion naturally evolved into the possibility of my joining his laboratory. Following this exchange and an invitation from Sid, I visited his laboratory at Yale University in April 1985 after I had attended the ribosome meeting in Corpus Christi, Texas. We continued our discussions and he suggested that I explore funding possibilities immediately. After receiving financial support from the Swedish Research Council and defending my thesis in December 1985, I joined Sid's laboratory in the end of January 1986.
Given my PhD research with translation and suppressor tRNAs in Escherichia coli, I started to work on the efficiencies of nonsense tRNA suppressors, including the classical tRNATyrSu3 nonsense suppressor. My objective was to examine the suppression efficiencies of these tRNAs in different RNase P mutant backgrounds to get insight into the action of this processing enzyme in vivo. Before the RNase P mutants were introduced into strains carrying different nonsense suppressors, we (I worked together with Maddy Baer) cloned and sequenced the mutants to map the changes in the RNase P RNA and protein (C5) genes. Sid was very supportive and encouraged me to examine various tRNATyrSu3 mutants in the laboratory freezer, in particular tRNATyrSu3A15, which he, as a post doc at the LMB in Cambridge (England), used to demonstrate that the E. coli tRNATyr gene is transcribed as a precursor. I presented to Sid the first draft of the manuscript, which included plenty of data. After reading the draft, he said that I had provided a good summary of the data. Howcver, Sid counseled that each section of a manuscript should include one question, how it was addressed experimentally, the results, and then finally the conclusions. I have tried to follow this advice and pass it on to my trainees as well. The manuscript was revised and published with the title “Differential effects of mutations in the protein and RNA moieties of RNase P on the efficiency of suppression by various tRNA suppressors.” In a follow-up study, we reported how some of these precursor tRNATyrSu3 mutants were cleaved in vitro with and without the C5 protein cofactor, thus giving me experience of also studying RNase P in vitro. In fact, this foray into in vitro work fortuitously shaped my independent career where I focused on dissecting the substrate-recognition properties of bacterial RNase P.
I stayed in Sid's laboratory for almost three years and left in December 1988 to start my own research group at Uppsala University. Again, Sid was supportive as I sought to set up my laboratory. In October 1989, it was announced that Sidney Altman and Tom Cech were awarded the Nobel Prize in Chemistry for their discoveries of catalytic RNAs. I tried to call and congratulate Sid but it was impossible to reach him by his phone in his office or his laboratory. Then, I called the neighboring lab (fortunately, I still had the Yale telephone book!) and asked if I could talk to Sid. This approach turned out to be successful and I got to congratulate him. Later in December 1989, I met Sid, his family and friends during the prize ceremonies and dinner, as well as when he and Tom Cech delivered their Nobel lectures in Uppsala.
Over the years, we stayed in contact and I frequently visited Sid and his laboratory. I updated him about our work on RNase P—in fact, we exchanged emails even until after Christmas last year. In 1995, I invited Sid to act as the faculty opponent during the thesis defense of Staffan Svärd, my first PhD student. Sid kindly accepted, examined Staffan's thesis, questioned him, and led the discussions constructively with insight and keen interest, thus making the public defense instructive for all attendees. Sid and I also met at different conferences. I invited him to a meeting that focused on noncoding RNAs and was held on an island in the Stockholm archipelago at the rim of the Baltic sea. Sid was one of the keynote speakers and he presented his recent work on RNase P and cleavage of new targets. His presence was much appreciated given the fruitful discussions that he engendered. When we were together at the FEBS meeting in Athens, Greece (June 2008), given our mutual interest in art and culture, we walked around Athens, visited a museum (see photo), and greatly enjoyed ourselves.
Sid was fond of Asian food. During my visits to New Haven, we had several pleasant dinners at Asian restaurants. On one occasion, he invited me to join him and Ann to have dinner at a Thai restaurant. Given my customs at home, I asked Sid if I had to wear a tie. I was serious but Sid took it that I was joking given that he always chose rather unpretentious restaurants with good food! Though I do not remember if I wore a tie, the food was tasty and we had a good time.
My fond recollections of Sid revolve around events that happened shortly after I had arrived and started my work in Sid's laboratory. Olof Palme, the Swedish prime minister, was shot and killed in Stockholm (February 1986). The day after the assassination, Sid and I discussed this sad event. I was surprised to learn from Sid that he was familiar with the work of Palme and his achievements. In April 1986, the devastating Chernobyl accident resulted in a radioactive fallout in Sweden. Of course, we discussed this event too. During these exchanges, Sid showed genuine concern and empathy. I was allowed to use the phone in the laboratory (no cell phones available at the time, of course!) to connect with friends and family in Sweden to ensure they were doing well. I appreciated this gesture as well as his meaningful support during these two tragic events. It meant a lot to me, as I had just left home and my thoughts were about Sweden.
Sid has left us and my thoughts go to his family and children who have lost a father and grandfather. Although Sid is gone, I will cherish many wonderful memories. In my world, he will be remembered as a mentor, friend, and a scientist with great integrity who trusted and pursued his ideas. Importantly, he was a scientist who “made sense and use of the unexpected.”
After receiving the sad news of the death of Sidney Altman, I took a long afternoon walk recalling my years with Sid when I was his graduate student and part of his research group. Then I had a look at the pictures in my photo albums to recall our times together, in the laboratory, at group meetings, parties, and seminars. It is with fondness and gratitude that I remember these times.
I am very grateful to Sid for having accepted me into his research group and then shepherding me through the ups and downs of research. He was a terrific person, scientist, teacher, and mentor. I came to his lab at the suggestion of Tom Steitz, whose group I had joined with the intention of crystallizing RNase P, as I had become interested in RNA structure and function during my first year of graduate studies. Sid welcomed me and immediately got me started, pairing me with a postdoctoral researcher who taught me the ins and outs of RNA and protein purification. Other members of the laboratory were ready to help me whenever I asked. The research group atmosphere was very collegial and stimulating. We had regular group meetings, which showed that Sid knew where everybody was in his or her work. He stimulated all of us to think problems through and come up with solutions. He offered clues but never forced an answer or a solution. He fostered debate and exchange of ideas. He encouraged us to be creative.
Sid was also very generous in acknowledging his coworkers and giving credit for the work accomplished in his laboratory. A case in point, when the news came of his being awarded the Nobel Prize, he had the entire team present at the news conference and share in the excitement. After he came back from Stockholm, where he had received the Prize, he let each one of us look at and handle the medal. He also gave each one of us a small Christmas tree ornament, which we still cherish and shall treasure even more so from now on.
Sid encouraged cohesion of the team even outside the laboratory. Often we had gatherings and meals at various restaurants in town and parties at some of the more senior members’ homes. These were great occasions for cross-pollination of ideas, scientific and otherwise. They also helped us get to know each other better.
Sid followed my progress until I graduated in 1991, even though I had to take on another project in the Steitz lab and spend more time there. Furthermore, he was interested in the progress of his students and postdoctoral researchers after they had graduated and left his group. He thus kept up with my progress through my years as a postdoctoral fellow and junior faculty. Along with the late Tom Steitz, Sid was one of my intellectual fathers and a towering one at that. He was a role model to me in practically the same way, I must say, as my own father, who was also a professor (of economics) before he retired and who passed away late last year. I am grateful to Sid for being an exemplar and for his generosity, and regret that I can no longer say this to him in person.
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TALES OF THE UNEXPECTED IN SIDNEY ALTMAN'S LABORATORY
Sid at 60 with me at his residence in Hamden, CT, during the Altman lab reunion of August 1999.
(Photo courtesy of Anthony Forster.)
My time in Sid Altman's laboratory from January 1988 to December 1990 was inspiring, eventful, frustrating, and life-changing, replete with important lessons from the great man himself. In my personal account of this period and Sid's earlier breakthroughs, I minimize repetition of what has been written so eloquently before, particularly by Sid, Ben Stark and Bill McClain.
I began my postdoc full of optimism as a 25-yr-old Aussie who had already discovered the hammerhead ribozyme structure and had moved to the lab of his choice in the United States with support from Anna Fuller Fund and (later) Jane Coffin Childs fellowships. But the start of my work at Yale proved challenging. There was the fortuitous distraction of meeting my future wife on the first day, and the theft of my handwritten draft of the double-hammerhead manuscript from a locked car (which I instantly learned was not atypical for New Haven!). And my first project, cleverly laid out by Sid, proved too difficult technically for me. The plan was to synthesize small hairpin substrates for RNase P that contained a unique phosphorothioate at the cleavage site, courtesy of a unique nucleobase in the template strand, using phosphorothioate NTPs kindly supplied by Fritz Eckstein. Ideally, these substrates would then act as competitive inhibitors and enable determination of the cleavage stereochemistry and the specificity of cation catalytic cofactors. I thought the easiest way would be for the Altman laboratory to adopt Olke Uhlenbeck's technique of directly transcribing synthetic oligodeoxyribonucleotides with T7 RNA polymerase, but my transcriptions were poor in comparison with ones that I had performed as a graduate student at Adelaide University. So, I asked my PhD advisor, Bob Symons, to kindly mail me some oligos. Based on direct experimental comparisons, I determined that the oligo synthesis core facility at Yale Chemistry was not synthesizing their oligos properly. Inspired by the whirlwind progress with standard (plasmid) transcription templates by Cecilia Guerrier-Takada, the most accomplished member in the Altman group, I next decided to learn DNA cloning to make substrates by cloning oligos. However, that also did not go well. To make matters worse, RNAs that incorporated thioNTPs led to puzzling results, consistent with a later publication revealing that special precautions were required to prevent oxidation during gel purification, handling, and storage (Milligan and Uhlenbeck 1989). Furthermore, RNase P cleavage was shifted to a site adjacent to the intended one (similar to Kahle et al. 1993).
During this frustrating period, my advisor also seemed less happy than expected, especially for someone that I had bet a case of beer on as a future Nobel laureate. A contributing factor may have been the jostling for credit. For example, early prizes for ribozymes were awarded exclusively to Tom Cech, who had published self-splicing two years and then one year prior to Sid publishing Cecilia's discovery of enzymatic RNA (Guerrier-Takada et al. 1983); also see Altman, RNA Society Newsletter of January 2009. Or Sid's service as Dean of Yale College (1985–1989) might have been taking a toll. More concerning was that the Altman laboratory had just found that their 1986 publication reconstituting RNase P activity from partially purified HeLa RNA and Escherichia coli C5 protein was artifactual due to the ability of nonspecific RNA to activate small amounts of M1 RNA contaminating excess purified C5 protein. This finding spurred me to manually deduce potential secondary structures of eukaryotic RNase P RNAs that bore partial similarities with Norman Pace's prokaryotic structures to further strengthen the case that the Altman laboratory had indeed cloned the bona fide HeLa RNase P RNA (Bartkiewicz et al. 1989). But my new structures, which only shared sequence and secondary structural homology at the ends of the RNase P RNAs, raised doubts about another Altman laboratory 1986 publication on the ability of M1 RNA to tolerate large terminal deletions, albeit with much lower activities. These doubts ultimately proved correct (see discussion of Guerrier-Takada and Altman 1992). I later added more homology between the structures, even extending it to RNase MRP RNA. Sid generously shared our submitted manuscript with the RNase MRP discoverer, David Clayton, but he quickly wrote a letter to the editor of Cell rebutting our submission. Fortunately, the editor published our manuscript (Forster and Altman 1990a) and not the rebuttal.
Given the difficulties of my first experimental project, I proposed a new one in mid-1988: to target any RNA for cleavage by RNase P by hybridizing the target with a complementary “external guide sequence” (EGS) RNA terminating with the CCA sequence conserved in tRNAs. Sid really liked this idea and made it the main subject of a patent application that he was previously writing with Cecilia on the ability of RNase P to cleave viruses (ultimately awarded as US patent 5,168,053). Then he had Cecilia, me and himself sign over our rights to Yale, cofounded Innovir Laboratories with an exclusive license to the IP with old friend Hugh Robertson and became a paid consultant. My own payoff was limited to doing the work for a publication (Forster and Altman 1990b), but this did give me tremendous satisfaction as it ultimately became Sid's second-most-highly cited research publication and he featured it prominently in his 1989 Nobel lecture. This work took some time, as I wanted to show that RNase P could cleave at a deoxyribonucleotide linkage (which it did) and make sure that the enzymes were saturated with both target and EGS RNAs to obtain reliable kinetic parameters (the affinities were weak for M1 RNA alone: KM = 4.6 µM; kcat = 4.3 min−1 for HinfI pAT1 with 29-mer; pAT1 hairpin gave 4.6 μM (230x published) and 8.6 min−1; ironically, these values were never published as one reviewer insisted we take out all the kinetic data!).
The most notable event in my middle year was Sid sharing the 1989 Nobel Prize in Chemistry with Tom Cech “for their discovery of catalytic properties of RNA.” Given how thoroughly the Nobel Committee investigates discoveries (e.g., by soliciting written testimonies), this award clarified credit for the research community. The motivation text for the prize included: “All enzymes were considered to be proteins… 1978… was the first time that an RNA molecule had been shown to be necessary for a catalytic reaction.” The Altman laboratory had been more modest at the time, citing the well-known ribosome precedent (which admittedly was thought of, not as an enzyme, but rather as a huge multienzyme complex requiring many RNA and protein factors) and four early obscure papers of enzymes copurifying with an RNA (cited in Kole and Altman 1979). But two of these four papers reported resistance to inactivation by ribonuclease; one was only published in Russian and none had panned out, so it seems that the Nobel Committee got it right. The serendipitous, heretical discovery of an RNA subunit, for which Sid gave full credit to his PhD student, Ben Stark, was rejected by Nature before publication elsewhere (Stark et al. 1978; ironically highlighted in Nature by Travers 1978). That breakthrough was a real slog in the face of huge experimental challenges and scientific skepticism (see Ben Stark's article in this tribute) not to mention fierce competition (Guthrie 1980 and other competitors cited in the introduction of Kole and Altman 1981). Sid suffered ridicule at conferences but held his ground with determination and soon bolstered it with reconstitution of the enzyme (Kole and Altman 1979). Knowledge that a ribonucleoprotein (RNP) enzyme estimated at the time to contain some 80% of its weight as RNA (Stark et al. 1978; later refined to 90%) changed the prevailing view that the RNA portions of all RNPs, including the ribosome, were mainly scaffolds.
Three years after the publication of Stark et al. (1978), Tom Cech presented self-splicing RNA at conferences and endured less resistance than had Sid (Cech 1989, Nobel lecture), presumably because of Sid's earlier RNP work. And, coming full circle two years after that, Cecilia's serendipitous discovery of the first true catalytic RNA in a negative-control experiment was instantly accepted thanks to Tom Cech's work. Remarkably, all experiments and writing for the manuscript (Guerrier-Takada et al. 1983) were completed and submitted in just under one month and published within three months! They concluded, “If proteins were relative latecomers in the evolution of macromolecules, then primeval manipulations of nucleic acids may have been carried out entirely or predominantly by catalytic nucleic acids themselves.” That statement and publication were the real nailing of the prior existence of an RNA world in the origin of life. Self-splicing might not have come extremely early in evolution as it is largely confined to only one of the three kingdoms of life, and life depends on trans-acting enzymes that remain active at the end of the reaction. RNase P appeared to be a universal RNP, was a true enzyme and provided a second example of a ribozyme. With respect to catalytic RNAs acting in trans, the Nobel Committee noted an important application: “Catalytic RNA will probably provide a new tool for gene technology, with potential to create a new defense against viral infections… ribozymes will probably be used as gene shears” (trans versions of the hammerhead). Unfortunately, we're still not there yet.
A general take-home message for me from these unexpected breakthroughs was that scientific assumptions should not be cherished too dearly, especially when confronted by strong data with proper controls. As a further example from the field and an ironic twist, the assumption mentioned above that all RNase P enzymes would contain RNA components was overturned by Peter Gegenheimer's lab (Wang et al. 1988; see Peter Gegenheimer's article in this tribute); that finding also faced considerable skepticism. Invalidating assumptions may seem to be a rather minor feature of everyday science. However, looking back through my own papers, a fifth of them went against an assumption at the time, and those papers were my more impactful ones!
My final year in Sid's lab (1990) ended with my best insight in the RNase P/RNase MRP field, with the benefit of hindsight, which ironically I never published. Based on several lines of evidence (e.g., with a deletion of G291G292 in M1 RNA; see Kirsebom and Altman 1989), I strongly favored pairing of the conserved penultimate tRNA sequence RCC with M1 RNA's 5′-G292G293U294 loop acting as an “internal guide sequence.” Testing this model was my top priority if I stayed in the RNase P field, but, disappointingly, my assistant professor job search didn't turn out as desired. So, with my work visa having expired and having taken the MCAT exam as a backup, early next year (1991) I was admitted into the M.D. training program of Harvard. Leif Kirsebom (who overlapped with me in Sid's laboratory in 1988 and who ultimately recruited me to Uppsala University) independently proposed this same pairing between M1 RNA and substrates and, more importantly, proved it (Kirsebom and Svärd 1994).
Through this retrospective, I hope to have illustrated the highly stimulating scientific environment of the Altman laboratory and also several of Sid's most endearing traits: integrity, a strong work ethic, ability to identify important problems, determination in the face of setbacks and criticism, ability to discard scientific bias when challenged by strong experimental data, high productivity promoted by fast and efficient action, and a well-grounded ego (even after winning a Nobel!). Sid and I became closer as the years progressed (see photo taken at Sid's residence in Hamden, Connecticut) and he will be greatly missed.
I thank Altman lab alumni Venkat Gopalan, Sergei Kazakov, Leif Kirsebom and Ben Stark for many helpful comments on the manuscript.
REFERENCES
Bartkiewicz M, Gold H, Altman S. 1989. Identification and characterization of an RNA molecule that copurifies with RNase P activity from HeLa cells. Genes Dev
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Sid Altman and the author at a lab meeting in 804 Kline Biology Tower, Yale University (May 1990). This conference room was used for different meetings (until the mid-1990s) and located two doors from Sid's office.
(Photo courtesy of Sergei Kazakov.)
When I arrived in Sidney Altman's laboratory in April 1990 as a postdoc, my first project was to map the binding sites of Mg2+ cofactors in M1 RNA, the catalytic subunit of Escherichia coli RNase P. M1 RNA was the first discovered RNA that has enzymatic activity allowing multiple turnover cleavage of precursor tRNA (pre-tRNA) substrates in the presence of Mg2+ and some other divalent metal cations (Guerrier-Takada et al. 1983). Since no crystal structure of M1 RNA was available at that time, Sid wanted to test if the location of essential Mg2+ cations in M1 RNA could be determined by metal-induced cleavage within the Mg2+- binding sites in M1 RNA alone and in a complex with a pre-tRNA.
This project proved difficult due to several challenges. First, it is hard to distinguish between structural and catalytic roles of Mg2+ in ribozymes. Second, X-ray crystallography data could contradict results of (bio)chemical experiments. For example, specific interactions between divalent metal cations (M2+) and RNA determined in solution for the hairpin ribozyme by chemical interference methods were not found in subsequent crystal structures, whereas original X-ray crystallography experiments for the hammerhead ribozyme determined two functionally important Mg2+ in its catalytic center. However, as demonstrated in later biochemical experiments and crystal structure analysis, neither of these ribozymes employ Mg2+ catalysis but rather use acid-base catalysis by RNA functional groups. Third, Mg2+ cations do not cleave the ribozyme under optimal reaction conditions. Although replacing Mg2+ by other metal cations that can rapidly cleave RNA (e.g., Zn2+, Pb2+ or Eu3+) is a commonly used approach to map the metal-binding sites, the results of such mapping are often misleading because of different binding sites within folded RNA and different interactions with RNA residues for Mg2+ compared to other metals. Fourth, metal cations cannot cleave the RNA at some metal-binding pockets. For example, only one of the three site-specifically bound Pb2+ cations observed in tRNAPhe crystals engenders effective cleavage, whereas the other two are inactive, presumably because of an unfavorable structure of these Pb2+-binding sites or inflexibility of specific phosphodiester linkages that restrict their ability to undergo conformational changes necessary for cleavage.
To circumvent these problems, we adapted a trick previously used for analysis of Mg2+ binding sites in tRNA by detecting Mg2+-induced RNA cleavage at mild alkaline pH. We found reaction conditions that preserve the catalytically active structure of M1 RNA and allowed fast, accurate processing of its pre-tRNA substrates at pH 9.5 even while revealing slow, site-specific metal-induced cleavages in the ribozyme and substrate RNAs. The key innovation was a functional approach using analysis of site-specific RNA cleavage induced not only by Mg2+ but also by different metal cations that can either support catalytic activity of M1 RNA (e.g., Mn2+, Ca2+, Ba2+, Sr2+) or inhibits it (e.g., Zn2+, Co2+, Ni2+, Cd2+, Zn2+, Co2+, Ni2+, Cu2+, Fe2+, and Eu3+) at pH 7.5 and/or pH 9.5. We found that there were different cleavage patterns for metal cations that can cleave M1 RNA but cannot support its catalytic activity and for M2+ that can do both. From these results and a comparison of cleavage of M1 RNA with that of a deletion mutant derivative, we identified two different centers for binding of metal ions in M1 RNA that are important for the processing of E. coli pre-tRNATyr. There is also a center for the binding of metal ions in the substrate, close to the site of cleavage by M1 RNA. Finally, we proposed a mechanism of the catalysis involving two Mg2+ cations in the M1 RNA catalytic center as described in Kazakov and Altman (1991). Our proposed functional importance of two metal cations bound near the catalytic center of M1 RNA has passed the test of time and now been confirmed by follow-up studies. This model also predated a seminal publication (Steitz and Steitz 1993) describing “a general two-metal-ion mechanism for catalytic RNA” and was credited therein. I consider it as another proof of the high standard and the fundamental attribute of research publications that came from Sidney Altman's laboratory.
Other exciting aspects of our laboratory were Sid himself and the talented people who worked and collaborated with him. The lab was a happy and inspiring place to work. Sid generously encouraged many projects related to RNA catalysis that were proposed by his postdocs. Among many events that happened during my time in Sid's lab, I share here one that was remarkable and funny. One of the intriguing features of M1 RNA was its ability to cleave many pre-tRNA substrates having different sequences and carry it out most efficiently in a form of M1 RNA dimers (Guerrier-Takada et al. 1986). My crazy hypothesis was that RNase P promotes dimerization of mature tRNA–pre-tRNA molecules where the mature tRNA acts either as an external guide sequence or as a catalytic RNA motif. I tested it and observed perfect (RNase P-like) cleavage of 32P-labeled pre-tRNAPhe by bulk tRNA purified by Sigma from E. coli. Sid questioned this finding and asked me to use northern blotting to test for the presence of M1 RNA in the purified bulk tRNA sample (from Sigma). Indeed, I found a trace of M1 RNA! Ironically, Sid had suffered a similar fate when skeptics did not believe that M1 RNA isolated from E. coli carried catalytic activity, but it rather was contaminated by a “true” protein enzyme. To lessen my disappointment, Sid gifted me what appeared to be his Nobel Prize medal. When I refused to take it, he smiled and disclosed that it was not real but a wrapped chocolate replica. I still have and cherish this gift as a priceless piece of my memory about Sid.
REFERENCES
Guerrier-Takada C, Gardiner K, Marsh T, Pace N, Altman S. 1983. The RNA moiety of ribonuclease P is the catalytic subunit of the enzyme. Cell
35: 849–857. 10.1016/0092-8674(83)90117-4 [DOI] [PubMed] [Google Scholar]
Guerrier-Takada C, Haydock K, Allen L, Altman S. 1986. Metal ion requirements and other aspects of the reaction catalyzed by M1 RNA, the RNA subunit of ribonuclease P from Escherichia coli. Biochemistry
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Steitz TA, Steitz JA. 1993. A general two-metal-ion mechanism for catalytic RNA. Proc Natl Acad Sci
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RNA. 2022 Nov;28(11):1393–1429.
SIDNEY ALTMAN: CHALLENGING AUTHORITY AND BEING AN ERUDITE SCHOLAR
The world lost one of the best biochemists and molecular biologists and I lost my mentor who was the most influential and helpful in my scientific career. Sid was not only an exceptionally talented scientist, but also a great educator and trained several generations of scientists who are currently continuing his scientific legacy all over the world. It can never be overstated how much I have gained and benefited from being trained under his guidance as a postdoctoral fellow in the 1990s. Among all the legacies that Sid left for us, what I appreciate the most are two of his profound characteristics––challenging authority and being an erudite scholar.
Challenging authority: Sid trained his students and postdocs to have creative thinking and an independent intellectual spirit, which, he told us, are the most important components in science. He always advocated healthy disbelief in the words of authority figures and keeping an open mind. To this end, he started by instilling in us the equality of mentors and students academically and encouraging us to challenge his thoughts. My postdoctoral work with Sid was on how human RNase P recognizes its substrates. Tony Forster, a postdoctoral fellow two years ahead of me in the Altman lab, had proposed an external guide sequence (EGS) concept that any RNA can be cleaved by E. coli RNase P if an EGS RNA base-paired with it in cells. My study on substrate recognition by human RNase P made it possible to extend the EGS-induced cleavage of RNA into human cells, therefore establishing a specific gene silencing method. I found that a custom-designed EGS that base-pairs with an mRNA to form the secondary structure equivalent to the acceptor stem, D stem–loop and variable loop of tRNA can induce a cleavage on the mRNA by human RNase P. The variable loop that forms a bulge in the mRNA-EGS double helix is critical for the cleavage efficiency. To reveal the role of the variable loop in RNase P recognition and obtain the knowledge important for us to design EGSs with the maximal efficiency in RNase P-mediated mRNA cleavage, I investigated the contribution of the variable loop in RNase P recognition. I hypothesized that the variability of the loop may render a substrate with a different acceptor stem and D stem–loop the best fit for a tertiary structure that RNase P recognizes. Then the question was how we determine the size and sequence of the variable loop in a particular EGS to achieve maximal efficiency. After several failures with “rational design,” I realized that we are not smart enough and we should seek help from higher powers. I came up with a bold and novel idea—according to the principle of “survival of the fittest” in Darwin's evolution theory: Can this principle be applied to molecular biology to create a selective pressure to screen out the most effective biomolecules in a test tube using molecular evolution? I consulted with Sid for advice. His first reaction was that he didn't think the idea would come to fruition. He was worried that I was wasting a lot of time for nothing. I went back and thought about it for another two weeks and made further modifications to the experimental design. I went back and talked to Sid again about the plan. He noticed that my design was improved and more mature than the first version, but he still thought it was not feasible. But this time, he didn't stop me from trying the idea. He said to me—“My feeling is that this experiment will not work, but I would like to see you prove that you are right and I am wrong.” After a few weeks, the experiment achieved better outcomes than expected. It was my turn to present at the lab meeting the next day. At the lab meeting, I started with the original idea and explained the experimental process. Before showing my results, I was constantly being criticized and questioned by others, including Sid. In the last few minutes, I showed my results: After evolution in the test tube, the biological activity of the selected molecule was increased a hundredfold. Sid was excited about this work. He then helped me publish the work in Science. Since I became an independent investigator, I have been copying Sid's style—open myself to the ideas of any student and coworker and encourage them to challenge my thoughts.
Be an erudite scholar: Sid was trained as a physicist but developed an interest in biology and chemistry. His interdisciplinary training laid the foundation for his enormous scientific achievements. Sid repeatedly made his point to his students that he never believed in genius; there are only two factors for success in science: willingness to learn and hard work. My parents came from China to New Haven for my wedding where they met Sid. When Sid left the wedding, he pulled me over and said, “Please make sure to bring your parents to my office and I would like to chat with them.” In Sid's office, my father, a professor of Chinese Literature at Shandong University, China, asked Sid a question—“Should a university offer an interdisciplinary curriculum and train students to be erudite?” He mentioned that in Chinese universities, all students are assigned a narrow specialty from day one in college and all colleges were run like technical skill schools. Sid replied: “Creativity is based on knowledge.” Then he said—“When I was the Dean of Yale College, I proposed and made a change in Yale college curriculum to have students conversant in all disciplinary areas: non-science major students are required to take some science courses, and science major students must take humanity courses.” Such requirement was initiated at Yale and later adapted by many colleges including Harvard, MIT, Columbia University, and UPenn. Sid was the most knowledgeable and erudite scientist I have ever met. Sid often ate lunch with lab members in the cafeteria on the top floor of the Kline Biology Tower, where all kinds of topics could be brought up to the table—history of science, cultures, politics, religions, and sports. For whatever topic, Sid was always able to provide deep insights, reflecting on his wide knowledge and profound thoughts. Everyone who knew Sid was impressed by this attribute. A few years ago, I brought my whole family to visit Sid in New Haven. When my son, a high schooler at the time, told Sid his dream of becoming a US Navy officer, Sid told him the story of Hyman Rickover, about his Navy experience, his struggle with racial discrimination in the Navy being Jewish, and his achievement to become the father of the nuclear Navy and a four-star admiral. Inspired by Sid's account on Admiral Rickover's life, my son pursued this goal and is now an active-duty surface warfare officer on a guided-missile destroyer.
Before leaving Yale, I sat down with Sid and asked him for advice on my future research directions. He said earnestly—“Your scientific training is mainly in biochemistry. I think that most people in the field of biochemistry do not understand the real questions in biology. I hope you find real questions in biology to work with.” After joining the University of Pennsylvania, Sid's words “real question in biology” have guided me to expand my scientific specialties from RNA biochemistry to herpes virology, and to cancer biology. The last time when I saw Sid in 2019, I showed Sid my recent discovery—some pediatric osteosarcomas may be caused by a tumor virus.
1Division of Infectious Diseases, School of Public Health, University of California, Berkeley, California; Program in Comparative Biochemistry, University of California, Berkeley, California
1Division of Infectious Diseases, School of Public Health, University of California, Berkeley, California; Program in Comparative Biochemistry, University of California, Berkeley, California
(Left) Pew Scholar Meeting, Mexico, March 2002. (Right) Lab reunion and 80th birthday celebration with family, colleagues, and former students, Yale University, June 2019.
(Photos courtesy of Fenyong Liu.)
I first met Sidney Altman in spring 1992 when I was interviewing for a postdoctoral position in his laboratory. I subsequently worked as his trainee from 1992 to 1995 before joining as faculty at the University of California at Berkeley. When I arrived in his laboratory, Sid encouraged me to explore the use of RNase P and its associated external guide sequence (EGS) as a tool for practical application, a research direction that I have continued to pursue to this day. We worked together to develop RNase-P catalytic RNA for antiviral applications, using herpes simplex virus 1 as the model system. Sid had tremendous insight and deep understanding of science and research; I benefited profoundly from his advice and guidance throughout my career.
Even after three decades, there are some aspects of my time in his laboratory that clearly stand out to me. Sid cared deeply for his trainees and students and mentored them closely; he talked with me regarding my results and upcoming experiments frequently, despite his busy schedule for travel, teaching, community service, and social engagement. Moreover, Sid created a wonderful culture and research environment in his laboratory. I learned from all the researchers and students in his laboratory, many of whom became my lifelong friends and collaborators. Sid also introduced me to his colleagues at Yale and beyond for further guidance on virology experiments. This larger network, which he facilitated, allowed me to perform many experiments in different laboratories, where the appropriate biosafety containment facilities were available. His mentorship and the environment he created was a paradise to explore science and start a career.
Since my departure from Yale in 1995, Sid continued to be a major influence on my professional and personal development. I recall numerous occasions when I sought his counsel, and he was always warm, kind, friendly, and generously offering his help. He always set aside some time to talk to me about my nonacademic activities whenever I visited him at Yale, as well as during the times where we met in meetings or traveled together (see photo, left). Sid always inquired about how my family and I were doing in our numerous email exchanges and phone calls, advising me to spend more time with my parents and discussing with me about the education of my children. His warmness, kindness, and love had a profound impact on me. Sid was, is, and always will be, a role model to me as a person who is always willing to help and to contribute to society.
At our last in-person meeting on December 24, 2021, his focus was on science and research. Sid discussed with me about my ongoing projects and asked me to send him recent preprints and reprints, which I did soon after. For his part, he sent me a manuscript on a related topic that he was working on in January 2022. The example Sid set as a scientist and educator continues to inspire me.
For the last few months after his passing, I have remained in shock and grief. I received many condolence messages from colleagues and friends around the world, including many of my own students and trainees, who have not met or known Sid. Sid touched many lives; his legacy as a scientist and educator will continue to inspire generations of scientists to come. I am profoundly impacted by and will forever remember his benevolence and empathy.
The memory of my final farewell to him, just a few months prior, remains as if it were yesterday. During the funeral, I had the privilege of meeting Sid's family and expressing my condolences in person, overwhelmed with emotion myself. Lifting his casket as one of his pallbearers, I felt this was the least I could do to honor a man who was a mentor, colleague, and friend throughout my academic career and in my times of need. As I was shoveling soil over the casket of such a great man with a lifelong influence on me, I whispered, “Farewell, my beloved mentor, thank you for all your help and support. I love you and may you rest in peace.”
I still remember the day I approached my doctoral supervisor with a short list of names of scientists with whom I wished to do my postdoctoral stint. The list included the names of stars in molecular biology. Raymond Kaempfer took a look at the list and pointed to a specific name: Sidney Altman. He said that he would directly write an email to Sid to see if he had room for me. It took a few seconds for me to grasp the word “room” seen on the close computer screen. To my joy, it took a short time also to receive a hopeful reply from the award-winning laureate from Yale University. Ray reiterated that I would do fine working with Sid. Back then, I did not imagine that small and big developments would take place in the near future. I found myself in an airliner landing in the JFK airport before moving to New Haven. Two generous and courageous postdoctoral fellows, Paul Eder and Adam Ruth, from Sid's team, picked me up from the snow-covered limousine station of New Haven and dropped me in the front of a reserved apartment in Humphrey Street. I had been told by the landlord that Sid asked him to keep, if possible, the apartment available for his foreign students and colleagues.
After introducing me to some of his team members, including Cecilia Guerrier-Takada and Donna Wesolowski, Sid advised me that I should be careful of the cold in the winter and repeated that as long as I could feel my nose and toes I should be fine. He, who lived and worked in a moshav as a young chap, was familiar with the climate of the arid land I came from and knew that its people are not used to severe snow squalls. To me, the expression of personal concern was a true sign that I would be in good hands. Indeed, as a postdoc and later as a principal investigator, I had a close personal and professional relationship with Sid. In a social occasion held for Sid in 2009, I was approached by a generous lady who whispered that among the many postdoctoral fellows that Sid had, I was one of a few with whom he had collaborated actively for many years. Sid was blessed with the highest standards of integrity and ethics as well as a passion for science, knowledge and education. In one gathering, I heard him mention often that hard work can bring success and that it does not matter what job you do for living, as long as you do it professionally.
As to science, Sid was familiar with our work on human RNase P, including the initial attempts to reconstitute the in vitro endoribonucleolytic cleavage of the 5' leader sequence of precursor tRNA by this ribonuclease. He was helpful to our scientific progress and keen in winning joint grants from the United States–Israel Binational Science Foundation starting from 2001. He asked that my group receive the entire allocated funds in support of our early research of human RNase P.
Even though I was requested to write personal reflections on my mentor, writing about the landmark discovery of the first RNA enzyme by Sid is unavoidable. The finding that M1 RNA is a genuine enzyme had been accomplished following preceding studies, including that of Kole and Altman in 1981, in which it had been demonstrated that the protein subunit, termed C5, of E. coli RNase P is inactive by its own in catalysis and that its interacting partner, M1 RNA, must be the one that carries out the endoribonucleolytic cleavage reaction reconstituted in vitro. Kole and Altman wrote: “The absence of any demonstrable hydrolytic activity of the C5 protein is striking and implies that the M1 RNA must be involved in activating the catalytic mechanism of the RNase P complex.” Here Sid already made the verdict that M1 RNA is the subunit critical for the reconstituted enzymatic activity. Shortly afterward, he adjusted the conditions of substrate cleavage by including higher concentrations of divalent ions that activated the M1 RNA itself in substrate cleavage assays, which were masterfully executed by Cecilia Guerrier-Takada. Short of a resemblance or reminders of known chemical reactions, Sid tackled the unparalleled case of an RNA that functions as a genuine trans-acting catalyst by following the strict Michaelis-Menten parameters of substrate recognition, catalytic efficiency and turnover formulated and applied for many decades to describe protein enzymes only. The discovery of M1 RNA as a catalyst had not been immediately acknowledged by the entire scientific community, as some reasoned that enzymes are made of polypeptides and not polynucleotides. Together with the discovery of the self-cleaving group I intron by Thomas Cech, the terminology of ribozymes was coined and the possible roles of RNA, alone or in combination with other polymers, were presented as the first replicating genomes of living cells on earth.
Sid visited Israel several times. Once he told me that he spent a few months working in exhausting farming in a moshav, Nahlal, before pursuing his academic studies in the United States. Later, as an independent principal investigator at Yale, he had ties with the Weizmann Institute of Science and collaboration with his close friend Raymond Kaempfer. In 2013, Sid received an honorary doctorate from the Hebrew University of Jerusalem. In this several millennia old city, he liked its unique cuisines and enjoyed its cold weather at night. The last time I met Sid was in the King David Hotel of Jerusalem in 2019 before the start of the coronavirus pandemic. He advised me that a person should take care of himself and enjoy life.
RNA. 2022 Nov;28(11):1393–1429.
IN MEMORY OF PROFESSOR SIDNEY ALTMAN: INSPIRING PEOPLE TO THE FULL
Photos of Sid with the authors. (Left) Sid and Ge during a press interview in the lab at Yale University in 2003. (Middle) Sid and Taijiao during Sid's visit to Beijing in 2006. (Right) Sid and Li during a lab reunion in the Department of Molecular Cellular and Developmental Biology at Yale University in 2015.
(Photo courtesy of Ge [left], Taijiao [middle], and Li [right].)
Sidney (Sid) Altman is recognized globally for the discovery of an RNA catalyst in RNase P. For the three of us, however, our time with him showcased his influential role as an inspiring mentor. Here, we reflect on our training with Sid, which encouraged us to pursue different career paths after leaving his laboratory.
All three of us graduated from the Shanghai Institute of Biochemistry and Cell Biology (SIBCB), Chinese Academy of Sciences (CAS). SIBCB is prominent for its contributions to biochemistry, molecular biology and cell biology having made an early mark through the synthesis of crystalline bovine insulin in 1960s. Trained as graduate students in three different SIBCB labs, all three of us had strong experience in RNA molecular biology, and importantly, were fascinated by the complexity and diversity of RNA. Luckily, we got an opportunity to work with Sid as postdoctoral fellows (Taijiao 1999–2002; Ge 2003–2004; Li 2004–2006), focusing on different aspects pertaining to the characterization of RNase P and use of this enzyme as a tool to regulate gene expression by the external guide sequence (EGS) technology. Although we had been involved in different projects there, we had all done the same very first experiment at Sid's laboratory: Test the catalytic activity of Escherichia coli RNase P RNA subunit!
While our research objectives were different, we had an overarching sentiment of Sid's complete devotion to research and striving for the best. Such an assessment was shaped by his open-door policy for discussion when we encountered unexpected results or needed help to solve problems. Sid's own enthusiasm for the work was evident by the fact that he would come to our lab benches regularly to learn of our results first-hand (seephoto, left). Hard copies of our manuscript drafts were edited carefully. He made sure that we wrote concisely without compromising the main message and ensured that redundancies were eliminated. These exchanges were formative especially for non-native English speakers.
Transitioning from graduate students to postdocs is never easy, a difficulty heightened when one crosses borders to a new country with different norms. However, our research and lives were fruitful and joyful because Sid and all lab members were supportive and ensured our professional and personal well-being. Sid liked to have a cup of coffee during morning breaks. Quite often, Sid also joined us during lunch time to share with us some scientific and non-scientific developments he thought interesting. Sometimes, Sid gave us a ride across New Haven, and he would narrate stories associated with some historical buildings at Yale. While we were so happy to learn about the past, we could also appreciate his pride of Yale, where he had worked during his entire independent career.
Around the time we joined Sid's laboratory, life science research was continuously booming largely due to the advent of new technologies, the completion of the human genome project, and the subsequent observation of genome-wide expression of noncoding sequences. Accordingly, we had opportunities to explore different research directions with the guidance of Sid at his laboratory and obtained his full support when choosing different career paths. After leaving his laboratory, we maintained ties with Sid and found opportunities to meet him in person. For example, Taijiao showed Sid around in Beijing when he was invited to China for academic activities (see photo, middle) and Li went to Yale for the lab reunion to share research in progress with Sid and many other lab alumni (see photo, right). During our conversations, Sid's sharp intellect was always evident. He would convey crisp thoughts on projects that we were focusing on, and importantly, gave positive feedback on our achievements. This encouragement, which sparkled in his eyes, inspired us to move forward. After learning that we worked very hard with little time to spend with our children, he shared his joy when he spent time with his children and grandchildren and suggested to us to schedule more time with our families. Such counsel strengthened our personal ties to him.
In recent times, despite his illness, Sid was still optimistic and even talked about travel to China. Due to the COVID19 pandemic, we had no chance to meet Sid in person after the lab reunion in the summer of 2019. In early April 2022, even while we were locked down due to the outbreak of omicron in different cities in China, we were saddened to learn of Sid's demise. At such difficult times, we all take recourse by reliving our cherished memories. Sid's integrity and dedication to science and mentoring remain useful beacons for us. While we deeply miss his enduring support and inspiration, we recognize that we were extremely fortunate to have worked with him.
I met Sidney Altman soon after he came to Yale from the MRC Laboratory of Molecular Biology, Cambridge, UK, where he initiated with John Smith and Sydney Brenner studies on tRNA precursors in prokaryotes. When he first came to Yale, I went up to the 8th floor of the Kline Biology Tower and saw a person walking down the hall that I presumed was Altman and, knowing that he was originally from Montreal (near where I grew up), I hollered a greeting in French Canadian. I was in turn greeted by a long reply in perfect French-Canadian dialect, the nature of which I will not repeat here. This exchange was the beginning of our 50-year friendship.
Initially, I must say that I was not particularly interested in Sid's project on tRNAs and the RNases that might be processing them, until his papers in the late 70s showed that there was a definite RNA moiety involved in the enzymatic processing of the tRNA. This information came to me through Sid's graduate student, Ben Stark, who was friendly with students in my own laboratory (see Ben Stark's contribution in this tribute) and Peter Rae, a new Assistant Professor who had joined us from the University of Chicago. Ben's PhD project was to purify bacterial RNase P, and during one of his meetings with his thesis committee, in which Rae was a member, Ben related his difficulties in the enzyme purification because of what seemed to be an “anionic contaminant.” Rae suggested that Ben treat the crude enzyme preparation with a nonspecific ribonuclease in case the “contaminant” was RNA and offered a suggestion to also inactivate it given the subsequent need to assay RNase P. In hindsight, this suggestion seemed reasonable, but I suspect, at the time, the idea of treating a crude preparation of RNase P with ribonuclease probably engendered some chuckles. The rest is history, of course, as this experiment established the requirement of an RNA for RNase P activity.
As the papers of Sid Altman and his colleagues began to be published on the RNA moiety of RNase P, he had a hard time convincing his colleagues, not only those working on RNA at Yale, but internationally, that he had an RNA essential for enzymatic activity on a tRNA precursor substrate. These were hard times for Sid, as he was also having a difficult time getting his papers published and getting invited to international meetings on RNA (even those being organized, in part, by colleagues from Yale!). RNA researchers did not believe his findings.
One morning (in 1988), as I was having breakfast in Stony Creek, Connecticut, not far from New Haven, and was reading the New York Times, I read an article about the awarding of that year's Louisa Gross Horwitz Prize, an international award often leading to a Nobel Prize. It was shared by Phillip Sharp (MIT) and Thomas Cech (University of Colorado), the latter being recognized for his discovery of an RNA enzyme (“ribozyme”). As I read, I hoped that Sid would be a co-recipient of this award for his work on RNase P, also a ribozyme, but still having difficulty being accepted. Sid was not a recipient. As you might imagine, he was not happy about his omission. Later that day, I gathered all of Sid Altman's publications on RNase P and drafted a rather long letter to the Horwitz Prize Selection Committee at Columbia University letting them know, quite directly, that they had made a mistake in not giving the award to both Cech and Altman. I did not expect a reply, but I did, indeed, get a very nice response from the Horwitz Selection Committee Chair, thanking me for sending the material I had sent them and letting me know “they would look into it.” That reply was more than I expected. I heard nothing further until the Nobel Prizes were announced in 1989, in which both Altman and Cech received the award for discovering catalytic RNAs. The day that occurred, the Press was swarming the Kline Biology Tower, and when interviewed I was ready with some of the material about Altman's research, which I had previously sent to the Horwitz Prize committee! I was quite happy that it all worked out for Sid.
My best times with Sid were skating at the Yale Ingalls Hockey rink across the street from the Kline Biology Tower. It was open to public skating for a couple of hours around lunch time. Sid was an excellent skater and a very good hockey player, having done so in high school in Montreal and at MIT, where he played on their varsity team. I had grown up with hockey in my hometown on the New York State-Canadian border, so we had a lot in common with the sport and knew the key hockey players, especially those with the Montreal Canadiens (Sid's favorite team!). You could tell Sid was a hockey player, just by the way he skated even recreationally, as the experts have a certain “swing” when they skate. Sid also played at Yale with a pick-up hockey team of graduate students and faculty, one of the latter being Fred Richards, Chair of Molecular Biophysics and Biochemistry at Yale, who was also an excellent hockey player.
Academically, there were many things about Sid Altman that were outstanding. He was a perfectionist with his research, an excellent teacher, and later a very good Dean of Yale College. As Dean, Sid helped reorganize the science requirements for Yale undergraduates. But, possibly, of even more importance was his oversight of the appointment of new faculty to the Department of Biology (renamed later as Molecular, Cellular, and Developmental Biology) while he was the department's chair. He had a “nose” for high-quality researchers in important research areas, and the people he helped hire invariably progressed to tenure in the department. I cannot say that for any other departmental chair in the five decades I have been in the department except for one person, Clement Markert, who was the first chair when the Kline Biology Tower was built, and who hired Sid Altman!
My friend Sid's most remarkable characteristic was his brutal honesty. I illustrate with a few stories based on some recent professional and personal interactions.
In late 2017, after reading the draft of one of my papers, Sid wrote, “Thank you for sending me your manuscript. To begin with, I find that rereading the text reminded me very much of the last manuscript you sent. In the movie Amadeus, the king says to Mozart after hearing his first opera, ‘Too many notes. Too many notes.’ We can substitute “words” for “notes” …
In the midst of the COVID pandemic, I started working on coronaviruses. As I had done many times before, I wanted to share the results of my experiments with Sid who always offered direct and helpful insights. One morning, I met Sid, who was appropriately garbed in mask and gloves and, of course, in his favorite NY Mets baseball jacket. I brought along a photocopy of the results of an experiment comparing the effects of codon optimization on the expression of nucleocapsid proteins encoded by the N genes of CoV2 and several endemic coronaviruses. Sid looked carefully at the paper, then folded up the paper and put it in his pocket. “I want to share this with my friends in Russia. This is a lot better than most of the crap you bring me!”
For those close to Sid, the above candor was typical. Such bluntness was perhaps shaped in part by Sydney Brenner, his role model. Sid once shared with me a story from his time at the MRC Laboratory of Molecular Biology, Cambridge, UK. A visiting postdoctoral fellow asked Brenner whether he could present a seminar. Brenner responded, “We don't give seminars; we just talk.” The postdoc persisted and Brenner agreed to let him give a seminar to lab members about his work. After the seminar, the postdoc asked Brenner whether he thought the work was publishable. Brenner responded, “Maybe you could get it published in the Jewish Chronicle” (the newsletter of the Jewish community in London).
Sid was always direct and sparing in his praise. For many years, I headed the Pediatric Infectious Disease Section at the Yale New Haven Hospital. One of my duties was to find labs for the physicians-in-training to conduct research. Sid agreed to take on one of our fellows who had been a Rhodes Scholar before attending Medical School, where he earned an MD/PhD. Sid put him to work on a collaborative project using RNase P to knock down a Salmonella gene that was important in pathogenesis. After three years, the fellow had published his work and was on his way to a junior faculty job at another university. The fellow, Sid, and I had lunch at Mory's, a venerable New Haven establishment since 1849. As a way of saying adieu and evaluating his research, Sid said to the fellow, “I think you will be a very good pediatrician!”
Sid took great pride in his teaching. In late April 2020, I wrote to Sid saying that I was reviewing grant applications for a small foundation that funds young investigators. One of them was studying lung cancer. In the opening sentence of her application describing her career goals, one applicant wrote, “As an enthusiastic Yale undergraduate, I took a seminar course titled ‘Novel Functions of Proteins and Nucleic Acids.’ Professor Sidney Altman opened my creative mind to endless possibilities and challenged me to cultivate my own scientific theories.” I forwarded this remark to Sid. He responded, “I am so glad to hear that my teaching had some effect. The chair of my department called me in one day and said my teaching was terrible. I vehemently disagreed with him. My chair is a ‘learn everything by rote’ teacher whose goal is high scores on the MCATs.”
Sid and I shared enjoyment of ethnic Jewish foods. Once in a while, I took Sid out to lunch at Katz's, a local delicatessen, where he very much enjoyed the chopped chicken liver. On one occasion, we were at Katz's when it had just reopened after a fire and was particularly crowded. The manager, standing directly at the back of our table, was shouting instructions to the wait staff and arranging seating for other customers. Sid requested the manager to quiet down but he did not. I said to the manager, “Do you know who that is? My friend is a Nobel laureate!” The manager said, “I don't care.” Sid later wrote back to me: “Thanks for your support with the loud-mouthed ‘maître d’ at Katz's. I find generally that nobody outside of the scientific world has any regard for or knowledge of honors and they pay no attention to it. Any mention of honors is met with puzzlement.” Sid took these affairs in his stride.
I really loved talking to Sid about experiments, sharing meals, and telling stories. He was a truly great scientist and one of a kind as a friend. Most of all, he spoke the truth as he saw it, a rare trait!
Sid Altman telling the author to take it easy after he paid Sid a compliment at the 2016 Symposium at Yale that was held to commemorate Sid's retirement.
(Photo courtesy of Daniel DiMaio.)
I knew Sid Altman for my entire scientific life. Sid was a lead professor in a course on “Molecular Genetics of Procaryotes” I took as a college senior in 1974. The course was difficult and eye-opening and inspiring, with a roster of amazing scientists as instructors including Sid, Joan Steitz, Bill Summers and Charles Radding. This course convinced me to become a molecular biologist, so Sid literally changed my life. I talked to Sid only once during the term, when I picked up my final exam. There was a stack of blue examination books on a wooden chair outside his office, and I saw that Sid was sitting at his desk. I knocked on the door and introduced myself as a student in his class, blue book in hand. Sid looked at me and scowled, and it was clear that the last thing he wanted to do was to talk to me, so I quickly thanked him, mumbled that it was the best course I had ever taken, and turned to leave, but he stopped me and said, “Sorry, students usually come to complain about a grade.” He then invited me into his office, and we talked for a few minutes about my impressions of the course, my nascent interest in molecular biology, and my future plans.
When I joined the Yale faculty almost 10 years later, I reintroduced myself to Sid and we became friends and collaborated on a few minor projects. I learned that my first encounter with Sid was not atypical: He could be gruff and distant—and he was always brutally honest. But he also had a soft side once he knew you. I remember calling him the day he won the Nobel Prize to congratulate him, and as we wrapped up the conversation, he said, “But, Dan, we're still friends, aren't we?”
For several years, Sid was Dean of Yale College. Many thought this was an odd assignment, given Sid's brusque exterior. But it actually was an inspired choice, because he had the single most important job qualification, a deep concern for students and their education. My family and I got to know Sid socially, and he joined us a few times for holiday meals or dinner on the patio. On these occasions he was invariably gracious and displayed his knowledge and interest in a wide range of topics. Six years ago, Sid asked me to emcee his retirement symposium (see photo), which brought together friends who had known Sid far longer than I had, including Matt Meselson, Mark Ptashne, and my own former postdoctoral mentor, Tom Maniatis. The respect and affection for Sid from this group of luminaries was palpable.
Over the years, Sid's passion for science and his rigorous approach to it never waned. Whenever we met, he unfailingly asked me what was going on in my lab and urged me to send him copies of my papers. After Sid's health started to decline, I would visit him at his apartment, and he would talk at length about his childhood in Montreal, current events and politics, his family, and, not infrequently, baseball. (He was a staunch Mets fan and I am a stalwart supporter of the Red Sox, so we shared an opinion of the New York Yankees.) At our last conversation, he told me that when he was a young child, he was going with an uncle to see the Montreal Royals, the local minor league team, and he was excited because he was going to see a Black baseball player for the first time. Someone named Jackie Robinson. But when he arrived at the ballpark, he was disappointed because Robinson wasn't in the lineup. He learned the next day that Robinson had been called up to the Brooklyn Dodgers to become the first Black major league player in the modern baseball era. Sid grudgingly admitted, almost 75 years later, that it probably was OK that he had missed seeing Robinson play, given the circumstances.
Sid Altman made a discovery that fundamentally changed our understanding of biology. On a more personal level, he altered the trajectory of my life and became a lifelong friend.
Beginning and end of a letter from Sid, written in 1984, demonstrating his commitment to providing strong support to a new junior faculty member.
(Photo courtesy of John Carlson.)
Sid Altman was a titanic figure in the RNA community, in Yale College, and in Yale's Department of Molecular, Cellular and Developmental Biology, where I am on the faculty. While Sid is widely remembered for his discovery of a novel catalyst, I will always remember Sid himself as a catalyst. He catalyzed my transition from naïve postdoctoral fellow to faculty member.
Sid was the chair who hired me. I was exceptionally fortunate he was Chair, because in retrospect I was an extremely risky hire. I was proposing to enter, as a beginning assistant professor, uncharted territory in a field in which I had no experience whatsoever. Most Chairs would have balked. Sid provided solid, unwavering support of the highest caliber. He said he would do everything in his power to provide an environment in which I could succeed (see photo of letter). And he meant it.
As it turns out, Sid ascended from Chair to Dean of Yale College shortly before I arrived at Yale, but he continued to consider me as his hire and to take a serious interest in my welfare. Despite the extraordinary demands of his new position, he regularly invited me to lunch. These lunches were unforgettable. They were often at an unassuming café just off campus called the Educated Burgher. Usually we were joined by Mike Snyder, who was also hired by Sid at about the same time, and who is now Chair of Genetics at Stanford.
At these lunches, Sid quickly focused on the essential. “What's happening in your lab?” He wanted to know. Although my work was far afield from his, he asked probing questions. He wanted detail. He was interested. And after a few months I realized that there were three reasons why I needed to design successful experiments: (i) to satisfy my curiosity; (ii) to assure my academic survival; and (iii) to have something to tell Sid at the Educated Burgher.
As it turned out, my laboratory's progress was excruciatingly slow. So it was all the more reassuring to feel that I had the attention, respect, and support of such a towering figure as Sid Altman. His confidence was most welcome during times when my own was wavering.
Sid also had a great influence on us, as assistant professors, in establishing a standard of rigor that we could try to emulate. The Nobel Prize only confirmed what we already knew: Sid was a remarkable scientist whose expectations were high.
Finally, I'd like to emphasize that Sid was always very cordial toward me. We were colleagues for 30 years, and I benefited a great deal from our many constructive conversations. I especially appreciated his eloquence, his broad intellectual interests, and his insight.
I feel deeply fortunate to have had Sid as a senior colleague in the Kline Biology Tower. It was an honor to have been so close to such extraordinary science, and I am very grateful to him for his catalytic role in helping me through a very challenging transition in my career.
(Left) Yale Press Conference, October 12, 1989. (Right) Relaxing in Stockholm, December 9, 1989, “YALE IS A GREAT PLACE FOR RESEARCH.”
(Photos courtesy of William McClain.)
Sidney Altman and I began our 53-year collaboration and friendship in 1969 as two Sydney Brenner–Francis Crick postdocs at the MRC Laboratory of Molecular Biology, Cambridge, England. These longstanding ties allow me to shed light on the decisive attributes of Sid, a complex and distinguished individual.
Serendipity plays a role in all scientific discoveries, but persistence and a strong work ethic are critical for success. Due to Sid's (and Tom Cech's) groundbreaking work, the idea that RNA was a key player at the origin of life has gained support. However, how did we get here? What innovations were the drivers for Sid's discovery? Perhaps a simple technique proved critical and inspired many advances. As described elsewhere in the perspective on the history of RNase P (McClain et al. 2010), Sid proposed isolating acridine-induced tRNATyr suppressor-negative mutants. He was using the Brenner model system where mutant phages overproduced tRNAs. During these investigations, it became clear that there were mutants that accumulated tRNA species with more nucleotide residues than the mature form. However, Sid was disappointed that he did not consistently and reproducibly observe these precursors.
Sid then summoned several postdocs in our lab (at the LMB) to discuss this complication. Perhaps the precursor tRNAs were labile and underwent rapid and complete degradation during cell harvest and RNA isolation. Reflection and discussion culminated in the suggestion to skip the cell-pelleting step and add phenol directly to the liquid culture. Sid immediately pursued this direction and designed a fast RNA extraction technique. Albeit simple, Sid fine-tuned the procedure in ways that would carry him to the Nobel Prize. This new method soon found widespread use in bacterial RNA processing research. For example, after establishing my group at the University of Wisconsin, we benefited tremendously from this approach. We were able to identify and sequence several phage T4-encoded RNA species containing two tandem tRNA precursors, that is, a multicistronic tRNA, a novel finding at that time (McClain et al. 1972; Guthrie et al. 1973; Barrell et al. 1974).
Back to England! Hugh Robertson soon joined Sid and me at the LMB and remained a very dear colleague to us until his untimely demise in 2005. Hugh brought his RNase III purification expertise and became involved with Sid in efforts to purify RNase P. Their very first report (now categorized as a JBC classic) formed a vital step toward a better understanding of the enzyme. Sid's eminent conservative approach never allowed him to overstate research findings and implications. When Hugh and Sid observed that the enzyme contains RNA with little protein, they concluded with what now seems a prophetic statement: “In light of these [purification] properties, it is possible that the active form of RNase P, which must have a strong negative charge, could be associated with some nucleic acid” (Robertson et al. 1972). Despite this cautionary language, this carefully stated conclusion garnered intense disbelief and dislike for Sid by a few well-connected workers, a sentiment only heightened after his nascent laboratory group made additional discoveries late in the 1970s at Yale University.
In the mid-70s, Benjamin Stark, a graduate student working with Sid at Yale, demonstrated that RNA is an essential component of RNase P (see Ben Stark's contribution in this tribute). This conclusion caused more consternation among several RNA biologists, and unfortunately, some were overly influential. With the lack of speaking invitations at conferences and new funds to support his research, Sid faced some of the most challenging times in his career. His family's love and support were vital during these periods. Sid's tenacity paid off in follow-up work by Cecilia Guerrier-Takada, an exceptional postdoc in Sid's laboratory. Her persuasive discovery using an in vitro transcribed RNase P RNA revealed that it exhibits multiple-turnover kinetics (Guerrier-Takada et al. 1983; Guerrier-Takada and Altman 1984), a hallmark enzyme property. At this juncture and with mounting independent evidence, refuting the enzymatic properties of RNA was no longer possible. In 1989, the Royal Swedish Academy of Sciences tacked on their approval.
Sid was a careful experimentalist, an assessment based on our numerous joint studies. Before his Nobel Prize, we collaborated to devise small RNA helices and model substrates to define RNase P's substrate recognition and cleavage properties (McClain et al. 1987). Such model substrates proved vital in studies of diverse enzymes that act on tRNAs (e.g., tRNA synthetases). Additional experimental work with Sid focused on phage T4 tRNAs. This work uncovered precursor tRNAs without the 3′-CCAOH sequences of mature tRNAs (Barrell et al. 1974) but required by RNase P (Seidman and McClain 1975; Guerrier-Takada et al. 1984), yet another novelty.
We should remember Sid's admirable administrative duties at Yale University as they coincided with his scientific achievements. He was the Chairperson of the Department of Biology (1983–1985) and Dean of Yale College (1985–1989). I recall reading an article in the Yale Daily News, a student newspaper, lauding Sid for instituting an undergraduate science requirement. I am sure it was not an easy task to incorporate such duties while performing cutting-edge science. However, Sid was relatively modest about his academic services at Yale.
Sid was a warm friend. After Hugh took up a faculty position at Rockefeller University, the three of us met multiple times in New York City to discuss the scientific progress in our respective laboratories and our lives. Hugh was the loquacious one! We were together for memorable milestones. One gathering was at the Four Seasons Restaurant to celebrate Hugh's tenure promotion when a Robertson lab member even delivered a bottle of chilled Veuve Clicquot Champagne to our table! Another was the 1989 Nobel Prize ceremony, where Hugh and I were Sid's guests for the entire week in Stockholm. While there, Sid learned that his undergraduate thesis at MIT in nuclear physics (the nonconservation of parity, performed under the guidance of Professor Lee Grodzins) had received experimental support. Sid told me that this news excited him nearly as much as the King of Sweden placing a Nobel Prize medal in his hand! The festivities and the buoyant atmosphere were made more special by my cherished interactions with Sid's family. The delightful week ended with my finally uncovering the contents (pricey chocolate bars) of Hugh's handbag, which had been an enduring mystery to Sid and me for several years!
I want to stress that Sid took great pride in his heritage. When I last visited Sidney three years ago, we discussed science (as usual) before Sid showed me a photo of some of his ancestors. Sidney was quite proud of his grandfather and uncle, describing them and their place of residence called Chornyi Ostriv (spellings can vary), southeast of Lviv in Ukraine. The recent Russian invasion has likely devastated this place, as is most of Ukraine. I am happy that my dear friend did not have to endure this tragic loss of life and the destruction of a place he treasured.
REFERENCES
Barrell BG, Seidman JG, Guthrie C, McClain WH. 1974. Transfer RNA biosynthesis: the nucleotide sequence of a precursor to serine and proline transfer RNAs. Proc Natl Acad Sci
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Sidney arrived at the MRC Laboratory of Molecular Biology in Cambridge, England, early in 1969. He preceded me there by just a few months, but I assumed that he was an old hand, familiar with the running of the lab, knowing everyone, and well-established in his project. Both of us were postdocs working on the cellular machinery for protein synthesis, albeit from different angles and with different goals. He was studying tRNA biosynthesis in E. coli, taking a mutagenic approach and using molecular analyses that ultimately—and unexpectedly—led to the groundbreaking discovery of the enzymatic activity of RNA. Only later did I learn that he, like me, was a newcomer at the time, still feeling his way into his research project. No one then had an inkling of how far it would take him and the field, of course.
Unlike Sid, I was coaxing a mammalian cell-free translation system to produce proteins under the direction of various cellular and viral messenger RNAs, an extension of my graduate work. We were in different departments. Sid was on the second floor, in a group led by Francis Crick and Sydney Brenner that boasted a dynamic crew of North American postdocs. I was on the third floor, led by Fred Sanger. Fred's group had pioneered the RNA fingerprinting method for RNA analysis that was exploited by Sid and many others on both floors. My small lab was shared with a rotating cast of illustrious visiting scientists working on DNA sequencing (primarily by methods analogous to those used for RNA, but destined to be soon surpassed by Fred's own innovations). The lab was directly opposite the indispensable high-voltage electrophoresis facility. This equipment was housed in a large room reeking of Varsol, a solvent used as a coolant in the electrophoresis tanks. The hallway in between was a gathering point where users of the facility would hang out for a few minutes in the relatively fresh air while waiting for their electrophoresis runs to finish. The X-ray film developer was also nearby, which added to the traffic and supplied occasional excitement when an unexpected or particularly sought-after image was spotted.
That hallway may well have been where I first encountered Sid. Certainly, it was where Hugh Robertson and I chatted frequently. Hugh was a friend and collaborator of Sid who arrived at the LMB at much the same time as Sid and I. Hugh had a background in the translation of viral RNA in a bacterial cell-free system that drew us together. He also had expertise with the nuclease RNase III and its substrate double-stranded RNA, which came into play in our subsequent collaboration. This expertise was instrumental in the purification and characterization of RNase P achieved by Hugh and Sid together with John Smith (Robertson et al. 1972). RNase P is the enzyme that cleaves the tRNATyr precursor which Sid had found. Because of its unusually acidic nature, deduced from its chromatographic behavior, they presciently speculated that RNase P might be associated with nucleic acid.
Over the next decade, Sid published an elegant series of papers confirming this perceptive comment and nailing down the nucleic acid component of the enzyme as an RNA molecule. He and his coworkers progressively documented and expanded on this observation. They identified the RNA, in E. coli and other organisms, showed that it is necessary for RNase P function (Stark et al. 1978), and then—famously—sufficient for tRNA precursor cleavage (Guerrier-Takada et al. 1983). This discovery, together with Tom Cech's discovery of self-splicing RNA, unleashed a paradigm shift: Enzymes do not have to be proteins, they can be RNA molecules.
For his discovery, Sid was awarded the Nobel Prize in Chemistry, an accolade he shared with Tom Cech. Needless to say, Sid's work on RNA did not end there. He went on to further the RNase P story and elucidate the mechanism of RNA catalysis as well as to make contributions in several other RNA-related fields. Moreover, he found the time to serve as chair of his department and even Dean of Yale College, where he had been on the faculty since leaving the LMB in 1971. We met occasionally throughout the years thereafter, often at RNA Processing meetings held at Cold Spring Harbor. My memories are of a quiet, somewhat reserved personality which thinly veiled an incisive intellect and a laconic wit, and a generosity of spirit. He was also resilient, successfully maintaining the research momentum of his group during a severe funding drought that he experienced at a critical juncture.
Looking through the 1972 RNase P purification paper, I was surprised (and somewhat gratified) to find my name mentioned. I was credited with providing rabbit globin for use as a marker, something I had forgotten long ago! It should not have been a surprise, though. More recently, Sid took the time to courteously shepherd my stepdaughter around the campus as a prospective Yale undergrad (she went to a music conservatory instead). A Yale student who took over as guide was clearly in awe of him, remarking with obvious understatement that he was one of the better-known people on campus. Recollections like these inevitably kindle a sense of regret that we did not spend more time together. Still, it is better to remember the pleasure and privilege of knowing such an accomplished, colorful and warm person.
REFERENCES
Guerrier-Takada C, Gardiner K, Marsh T, Pace N, Altman S. 1983. The RNA moiety of ribonuclease P is the catalytic subunit of the enzyme. Cell
35: 849–857. 10.1016/0092-8674(83)90117-4 [DOI] [PubMed] [Google Scholar]
Robertson HD, Altman S, Smith JD. 1972. Purification and properties of a specific Escherichia coli ribonuclease which cleaves a tyrosine transfer ribonucleic acid presursor. J Biol Chem
247: 5243–5251. 10.1016/S0021-9258(19)44963-6 [DOI] [PubMed] [Google Scholar]
Stark BC, Kole R, Bowman EJ, Altman S. 1978. Ribonuclease P: an enzyme with an essential RNA component. Proc Natl Acad Sci
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1Department of Biochemistry and Molecular Biology, Institute of Medical Research Israel-Canada, Faculty of Medicine, The Hebrew University, Jerusalem, Israel
1Department of Biochemistry and Molecular Biology, Institute of Medical Research Israel-Canada, Faculty of Medicine, The Hebrew University, Jerusalem, Israel
Sidney Altman with Raymond Kaempfer in April 2008 at the Yale Club in New York.
(Photo courtesy of Raymond Kaempfer.)
Sid and I first graduated in physics and chemistry, respectively, before our paths crossed in the Meselson laboratory.
“Ray, do you really believe that messenger RNA exists?” This question greeted me when I first entered the laboratory at MIT where I was to do my doctoral research with Boris Magasanik, having earned an undergraduate degree in chemistry from Leiden University in Holland. It was September 1961; Jacob and Monod had just presented their mRNA hypothesis at Cold Spring Harbor. Indeed, earlier in that same year, using heavy isotope transfer experiments in phage-infected bacteria, Brenner, Jacob, and Meselson had provided the first evidence for the existence of an unstable intermediate carrying information from genes to ribosomes for protein synthesis: messenger RNA. This advance was the start of RNA molecular biology. My doctoral thesis was dedicated to studying how T-even phage infection affects cellular mRNA stability. This theme would subsequently create common ground with Sid. Later, once Matt Meselson had moved to Harvard, this topic also motivated me to work with him as a postdoc.
As Sid recounted in The RNA Revolution, a series of in-depth interviews with Altman, published in Italian in 2011: “Matt's laboratory was frequented by many postdocs and some graduates. Among the former were Ray Kaempfer, Marc Rhoades, Toshio Nagata (a senior visitor) and Bob Yuan. The atmosphere of our group was very friendly, but we also worked hard, never wasting time.”
Once at Harvard in the fall of 1966, I was confronted with the question, whether the universal bipartite nature of ribosomes reflected their two subunits as structural building blocks that are permanently associated into functional ribosomes, or whether this could serve a function in protein synthesis, with the ribosomal subunits coming apart. I would often go back to the lab until midnight. It was during this research that Sidney Altman appeared in the Meselson laboratory as a new postdoc in the fall of 1967. He had just completed his thesis on bacteriophage T4 DNA replication in Denver, Colorado. Bob Yuan worked with Matt on isolating a first restriction enzyme. Now, Sid embarked with Matt on a study of phage T4 infection that yielded a T4-induced endonuclease that attacks T4 DNA.
Although we were working on very different projects, Sid and I soon became close friends. In the laboratory, we would also play pranks on him to break his serious mindset, such as tying his chair to his desk with a rope in the early morning before Sid came in. Much in need of social interaction, he would enjoy coming to our house. At one stage, my wife and I went out for dinner often, as she was then heavily pregnant and we knew that once our baby was born, we would no longer be able to do so. Sid joined us one key evening as we went to a restaurant in the Italian section of Boston. It was a tasty yet heavy meal. That night, her contractions started and the baby was born that same morning. It turned out to be a boy. In those days there was no email as yet, nor could one make international phone calls from home. From the Harvard Square post office, I sent a brief telex to my parents who were living in Holland: “See you at the brit. Gideon.” However, in those days the long flights from Holland to America were not yet frequent. My parents could not arrive in time for the circumcision ceremony held eight days later, where my father would have served as the godfather. I thus turned to my best friend, Sid, and he became our son's godfather. Sid recalls this episode in The RNA Revolution: “Ray and I became good friends, and when his wife Miep, a physician, gave birth to their first child (which happened after a heavy Italian dinner loaded with garlic) I was his godfather.”
Prompted by a visit of Sydney Brenner, Matt's close colleague, Sid left for the MRC (Cambridge) in October of 1969, where he was headed for studies on tRNA precursors that would later result in his discovery of catalytic ribonuclease P RNA at Yale. An outstanding doctoral student in my laboratory in Jerusalem, Nayef Jarrous, later joined Sid for successful postdoctoral research on ribonuclease P that earned him an academic position at our Medical School. Sid and I kept meeting regularly throughout his lifetime (see photo). Whenever I came to Yale, I would stay at his house. Whenever Sid came to Israel, he would come to our house and interact with our children. He remained in close contact with me and my family until the very end. He will be sorely missed.
Sid and I first crossed paths in 1971 when he joined the faculty at Yale. I had already been up the road at the University of Connecticut School of Medicine for a couple of years, and it was exciting to now have someone nearby who also was interested in tRNA processing. Sid was already well known in the field for his discovery of the first tRNA precursor while a postdoc at Cambridge, and I was delighted to get to know him. This ultimately led to a friendship of over 45 years.
Early on, together with Hugh Robertson, Sid discovered RNase P, the enzyme that generated the mature 5′ end of tRNA. It was a running joke between us that only one RNase was required to process the 5′ end of tRNA, whereas I had chosen to work on processing of the 3′ end, and even though multiple RNases acted at the 3′ end, none had the unusual properties of RNase P.
RNase P would occupy the rest of Sid's scientific career. Since his work with RNase P led him more and more into protein purification and enzymology, areas with which I had experience, we had occasion to speak frequently. I remember distinctly discussions in which Sid was frustrated with the inability to remove an RNA component, which initially was believed to be a contaminant. However, upon further examination, it became clear that the RNA copurified with RNase P activity through an extensive purification protocol, and most importantly, inactivation of the RNA component by micrococcal nuclease or pancreatic RNase A led to loss of RNase P activity. This finding led to the inevitable conclusion that RNase P was a ribonucleoprotein and that the RNA component was essential for activity.
This period was a frustrating time for Sid since these ideas were not readily accepted by the community. I remember multiple instances in which Sid expressed his difficulties in getting his work published. Nevertheless, he persisted and gathered more and more data to support his ideas, first separating the protein and RNA of RNase P into inactive components, and then reconstituting the complete ribonucleoprotein and regaining cleavage activity. Finally, in a coup de grace, he was able to show with Cecilia Guerrier-Takada and Norman Pace's lab, that under special conditions, the RNA moiety itself was capable of specific cleavage of a tRNA precursor. The rest is history. I was especially pleased that I was able to get Sid to present this work to my department at UConn in one of his first public announcements of this unprecedented discovery.
Sid and I continued to get together for many years at RNA processing meetings or at International tRNA Workshops and would have meals together and take strolls at the venue. I remember particularly the tRNA Workshop in Cambridge in 2000 in which Sid took me around the town pointing out familiar haunts of his. More recently, Sid took time from his busy schedule as a Nobel Laureate to spend several days at my current institution, the University of Miami, to present a name lecture and speak at length with students and faculty. I will always be grateful for that kindness.
Sid's scientific legacy changed biochemistry forever. His perseverance in pursuing the mechanism of RNase P action, despite the difficulty of the problem and the resistance of the scientific community to the idea that something other than a protein molecule could be an enzyme, serves as a model for what scientific inquiry is all about.
RNA. 2022 Nov;28(11):1393–1429.
RECOLLECTIONS OF MY SCIENTIFIC INTERACTIONS WITH SIDNEY ALTMAN
1CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, Shanghai, China
1CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, Shanghai, China
A photo from the Sidney Altman Symposium, Yale University, March 24, 2016. Front row (left to right): Spyros Artavanis-Tsakonas, Matthew Meselson, Sidney Altman, Daniel DiMaio, and William McClain. Back row (left to right): Ling-Ling Chen (the author), Ronald Breaker, Thomas Maniatis, Roger Kornberg, and Mark Ptashne.
(Photo courtesy of Ling-Ling Chen.)
My first scientific interaction with Sid was in 2008. At that time, it was reported by David Spector's group at the Cold Spring Harbor Laboratory that RNase P performs 3′ processing of two long noncoding RNAs (lncRNAs) to yield tRNA-like cytoplasmic RNAs. These lncRNAs are MALAT1 and the long isoform of NEAT1, which localize to two types of nuclear bodies, nuclear speckles and paraspeckles, respectively. At that time, I was a graduate student with Gordon Carmichael at the University of Connecticut Health Center and focused on unraveling the gene regulation roles of retrotransposon Alu elements and NEAT1 in paraspeckles. Thus, I was very excited to see this unexpected nexus between RNase P and NEAT1 biogenesis. Therefore, I boldly invited Sid to serve as my external thesis committee member as I was preparing for my dissertation defense in early 2009. Sid generously agreed. He drove from New Haven to Farmington on a rainy morning and was fully engaged during my thesis defense. I still distinctly recall Sid asking me many questions during this two-and-a-half hour exam. These included basic biochemical questions such as “Why did the addition of deoxycholic acid to your fractionation buffer allow a better fractionation of cytoplasmic and nuclear RNAs?” as well as more difficult and thought-provoking questions such as “How do you expect NEAT1 to work in paraspeckles?” and “How many functional lncRNAs would you expect to see in the genome?” As with most doctoral candidates who live the moment of their final exam, I do not fully remember how I handled all these questions. However, I clearly recollect that Sid looked serious during the entire defense and that I was nervous! Nevertheless, having this intense exchange with Sid on this specific milestone occasion inspired me to pay attention to details and to ponder about the many open questions in the field.
My graduate study had witnessed one of the most exciting discoveries in molecular biology and genome biology: the widespread expression of the “mRNA-like” lncRNAs that originate from intergenic regions. Motivated by the nonpolyadenylated NEAT1, I asked one question: “Do all lncRNAs look similar to mRNAs?” To address this question, I developed novel assays to explore nonpolyadenylated transcriptomes in Gordon's laboratory, funded by a State of Connecticut Stem Cell Seed grant. In 2010, I was offered an independent position at the Shanghai Institute of Biochemistry and Cell Biology, CAS. I visited Sid in his office (at Kline Biology Tower, Yale) shortly before I relocated to China. Sid was very happy to learn that I was going to have a laboratory in a top institute in my home country, which he had visited several times. He liked my idea of exploring previously under appreciated nonpolyadenylated transcriptomes. I recall his perceptive remarks: “Risky, but potentially interesting. But new findings usually come from seemingly impossible contexts.” He encouraged me to email him if I found anything new related to RNase P.
After having moved back to China, I did not really get a chance to meet with Sid although we exchanged occasional emails. Unfortunately, we did not identify additional lncRNAs that were processed by RNase P, but rather those stabilized by snoRNAs at the ends and circular RNAs. However, I was thrilled and pleasantly surprised to receive an invitation from Dr. Ronald Breaker to speak at the “Sidney Altman Symposium” (March 2016) at Yale that was being organized to honor Sid's retirement. When I learned later that Sid was the one who had suggested my name to Dr. Breaker, I realized that Sid had actually followed my independent research career in Shanghai!
In this symposium, I was the only female and junior scientist speaker. However, I was treated just the same as the other distinguished scientists. I had many opportunities to talk with these leading scientists whose findings I had read in top scientific journals or in textbooks. I cherish the group photo taken at this historic symposium (see photo). When I asked Sid why he chose me to speak at this event, Sid said, “Research has been carried out well by many female scientists. Scientists should be recognized regardless of their gender and geography.” The enormous amount of respect that I received from him and from others at the symposium greatly motivated me to become the best I can be in research.
Although not his direct trainee, I am sure that I am one of the many junior scientists who had Sid's encouragement in their early career stages. He was a man of few words. All our conversations were brief except during my PhD defense! I recognized, however, that he cared deeply for the younger generation of scientists and endeavored to promote them in their scientific journeys. I vividly remember when he said “very good” with a smile and in a light voice right after my defense in 2009 and after my talk at the Sidney Altman Symposium in 2016. These simple words from a great scientist meant so much to me, a junior scientist who dreamt of research excellence.
(Left) Dr. Sidney Altman's visit to Shanghai in 2007. (Left to right) Drs. Mofang Liu, Enduo Wang, and Altman. (Photo courtesy of Enduo Wang.) (Right) Sid Altman and Yong Li at Hamden, Connecticut (2017).
(Photo courtesy of Yong Li.)
Sidney (Sid) Altman's death is a great loss to all who have known him personally and have been touched by his wisdom and kindness. This remembrance is shared by a former postdoc (Y.L.) and a longstanding friend (E.W.). While Sid is well known as a pioneer in RNA catalysis, a supportive mentor, and an amiable colleague, we highlight here his uncommon giving of time and effort to a faraway land.
Sid visited China multiple times and developed a personal friendship with E.W. and many other scientists, and, importantly, enjoyed his interactions with students in different settings and campuses. In 2004, during his visit to Shanghai, he delivered seminars on scientific research and ethics at Fudan University and the Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences. As a very loyal Yale faculty member, it was no surprise that Sid visited Beijing in 2007 to participate in the “Peking University–Yale University Lecture Series” at Peking University, where he gave a lecture on “Life and the properties of a special RNA molecule.” During a subsequent trip to Shanghai, Sid gave three public lectures at the Institute of Biochemistry and Cell Biology and a series of six lectures solely for undergraduates at Fudan University. In all these forums spanning more than a decade, he held active discussions with small and large groups of students, and was always well received by the participants. Since August 2010, he served as a member of the International Expert Advisory Committee of the Institute of Biochemistry and Cell Biology and provided valuable advice on faculty development and institutional advancement.
During the happy hours in Shanghai, Sid enjoyed conversations with many acquaintances, including the insulin pioneer Dr. Youshang Zhang, who was the only student from mainland China to study during the 1960s at the University of Cambridge in England. In addition to developing enduring personal ties (as attested by E.W.), Sid took a great interest in Chinese cuisine and tea. He appreciated the Longjing (Dragon Well) green tea, and took this high-quality tea home.
Despite his preferential status at all scientific venues, Sid never liked any deferential treatment. For example, in 2009, Sid was awarded the Severo Ochoa medal at the 21st IUBMB International Congress on Biochemistry and Molecular Biology in Shanghai. During this meeting, E.W. saw Sid sitting on the floor while attending several talks in a fully packed conference room. Although the meeting organizers had reserved a seat for him in the front row, Sid did not want to disrupt others or distract the speakers. His considerate behavior was a charming hallmark. Sid was a great role model in many aspects.
Sid mentored over ten graduate students and postdoctoral associates from China. One postdoc (Y.L.) obtained his PhD in Biochemistry and Molecular Biology, with a thesis that focused on leucyl-tRNA synthetase and supervised by E.W. and Yinglai Wang. In Sid's lab, Y.L. characterized human RNase P, a multisubunit RNP, using molecular, biochemical, and genomic approaches. The training in the Altman laboratory sharpened Y.L.'s analytical and experimental skills and prepared him for a faculty position at the University of Louisville. Importantly, Y.L. remembers many inspiring conversations. For instance, Sid reminded Yong that breakthrough discoveries may or may not come, yet it is still rewarding to bring small increments with “small papers.” Sid also gave practical advice on giving enthusiastic seminars and using the laser pointer with care!
Another enduring and fond memory (of Y.L.) about Sid concerns cars, a flooded one. When Y.L. got his 10-year-old used car flooded in 2000 (only a few months after he got his driver's license), Sid listened carefully and then provided advice on what he should do in the event of a recurrence: he provided specific instructions on how to take the exhaust pipe out and drain the water. At that moment, Y.L. thought Sid might be a do-it-yourself mechanic!
For all his scientific achievements, Sid remained highly accessible to his trainees and importantly even to students and scientists around the world, including China. Sid was a global citizen who readily gave of himself, unmindful of geographical borders.
RNA. 2022 Nov;28(11):1393–1429.
TARGETING RNA TO FIGHT ANTIBIOTIC-RESISTANT BACTERIA
Russian-American Laboratory, headed by Sidney Altman. House of Scientists, Academgorodok, Novosibirsk, Russia, 2014.
(Photo courtesy of Marina Zenkova.)
During the past few decades, our knowledge of RNA molecular biology has expanded significantly in part due to Professor Sidney Altman. His inquisitiveness, critical thinking, and hard work were instrumental in his pathbreaking discovery indicating that RNA, in particular the RNA component of bacterial RNase P, is endowed with catalytic activity. His findings, together with those made independently by Tom Cech, overturned the established notion that RNA acts only as a carrier of genetic information. For this important advance, Altman and Cech shared the 1989 Nobel Prize in Chemistry.
In later years of his scientific career, Sidney Altman's attention was focused on an antibacterial strategy centered on the use of RNase P and specific oligonucleotide constructs, external guide sequences (EGSs), for targeted gene inactivation. This work gains significance given the pressing problem of growing antibiotic resistance and the need for new bactericidal therapeutics to eliminate fast-emerging multidrug resistant bacteria. In this regard, Sidney Altman was quite excited about the use of antisense antibiotics that allowed direct targeting of unique RNA sequences of pathogens. However, the work of Sidney Altman and his team, as well as those of numerous other groups, left no doubt that in order to increase the efficiency of antibacterial therapy, it was essential to have nucleic acid analogs that are resistant to intracellular nucleases, exhibit high biological efficiency, and possess superior cell-penetrating ability.
Sidney Altman was aware of the significant achievements of the Siberian Institute of Chemical Biology and Fundamental Medicine (ICBFM) in this field, an organization that conducted pioneering work on the efficient inactivation of RNA target and suppression of gene expression by complementary oligonucleotides. Therefore, when the Russian government established a program to finance leading laboratories managed by established scientists and titans in the field, we approached Sidney Altman to head the Russian-American Laboratory founded at the ICBFM (2013–2015). With genuine openness and enthusiasm, he accepted the invitation even though he understood well all the ground realities and difficulties of organizing a new laboratory from scratch, especially given the challenges of procuring research supplies to Siberia. We were also moved when he flatly refused to accept any payment for his time and effort.
The main goal of this new program was to develop targeted antibacterial and anti viral prototype therapeutics, based on chemically modified oligonucleotide analogs and their conjugates. During his visits to Novosibirsk, Sidney Altman gave valuable advice not only on science but on organizing effective teamwork. He was also a great mentor and role model for the youth of the laboratory. Apart from being an outstanding scholar, Sidney Altman was also someone who charmed all his colleagues with his wit and sense of humor. His presence ensured that young scientists and students were deeply involved in problem-solving discussions. His invaluable contributions, inspiration, and unwavering commitment to science will be felt for many years to come at the Russian-American Laboratory and ICBFM.
Sidney Altman's constant support ensured the success of the Russian-American project. As a result of three years of work, several chemically modified EGS oligonucleotides that were highly stable in biological media were developed. This list includes selectively modified 2′-O-methyl and 2′-fluoro analogs, as well as completely new DNA derivatives (e.g., phosphoryl guanidines), which showed efficient cleavage of a model RNA target by bacterial RNase P. We also obtained exciting data on suppression of growth of various Salmonella enterica strains by peptido-nucleic acid oligomers targeting vital bacterial genes. A pronounced antiviral activity was also confirmed for a morpholino oligonucleotide targeted to the second segment of influenza virus RNA. As expected, delivery turned out to be the biggest challenge in experiments with bacteria. Significant progress has also been achieved in this direction. Conjugates of 2′-OMe-modified RNA and selectively modified phosphoryl guanidine oligonucleotides, when combined with polycationic peptides, penetrated Escherichia coli and Acinetobacter baumannii and inhibited bacterial growth. Another major achievement was the creation of fundamentally new oligonucleotides with unique properties, specifically N-mesylphosphoramidate oligonucleotides, which has opened up new possibilities for inactivation of pathogenic RNAs associated with various human diseases. This type of DNA analog recruits RNase H and has significant advantages over phosphorothioates with respect to RNA affinity, nuclease stability, and specificity in inhibiting carcinogenesis. In some cases, these mesyl oligonucleotides may be an attractive alternative to phosphorothioates.
After the end of the project, Sidney Altman continued to keep in touch with the staff of the Russian-American Laboratory. He was pleased to be kept abreast of the successful functioning of the team and was supportive of our work. Our continued development of antisense drugs to combat bacterial infections and other human diseases is a fitting tribute to the rich legacy that Sidney Altman has left behind at ICBFM.
During the summer of 2017, we visited Sidney Altman in New Haven and recorded long conversations with him over two days. One of us (J.B.L.) was only six years into an independent science career then and was grappling with the question of what makes illustrious scientists “great.” One of us (V.G.) had previously been a postdoctoral fellow with Sid and enjoyed many interactions with him in different contexts. Both of us were passionate about scientific history in general and inspired in our own research by the discovery of catalytic RNAs by Sid and Tom Cech, one of the most important breakthroughs in molecular biology. Therefore, we were eager to hear Sid's views on his discovery and his philosophy about science in general.
Sid—a private person who was never interested in making himself well known—was patient, thoughtful, and expansive even as we covered an array of topics spanning his life, his work, attributes of great scientists, the nature of scientific discovery and the research enterprise, research advising and teaching, and the role of social and cultural contexts on the scientist. From this extensive interview, which was conducted at his home and at his office at Yale, we highlight here select questions and responses to showcase elements that shaped Sid's life and importantly to share some of his perspectives that are likely to be valuable to all researchers. (In some instances, we have collated from different parts of the interview that related to the same question, expanded abbreviations, or provided comments to supplement his responses.)
Q: What appealed to you about science?
A: As a child, I was interested in natural phenomena like thunder and lightning. I suppose I was interested in science. When I was 12 or 13, one of my relatives gave me a book called Explaining the Atom, by Selig Hecht. I was impressed by Mendeleev and his formulation of the periodic table, and importantly his ability to predict the presence and properties of some elements that had not been found. For a 12-year-old, it was amazing that someone could think about something deeply and make suggestions about what might be discovered. So, that fixed me on the path to science, and equally important was my reading about the history of physics in the first part of the 20th century. (Note: Elsewhere, Sid has talked about how the atomic bomb was in the news when he first started school and it was clear to him that science was important. He also mentioned that even before the age of 10 he had read about Einstein, who was presented as a model for anyone interested in studies.)
I always wanted to be a physicist. Due to certain circumstances, I went to MIT instead of McGill. At MIT, I pursued physics and found that fellow students were stimulating and crazy. I heard about interesting things all the time. My teachers were pretty good too. My Senior Honors thesis was completed under the mentorship of Lee Grodzins and it was serious research. These experiences provided a firm interest in science but I should also mention here that there was also nothing to dissuade me from science. My parents had no force in their voices. They never mentioned medical school or the legal profession, which was highly unusual for immigrant families in Montreal since these professions guaranteed good living. I have to praise them for it.
Q: Did you have early scientific heroes? What makes them great scientists?
A: My scientific heroes were Einstein, Bohr, and Rutherford. They were great scientists in every way because they recognized a new era in their fields and took to it quickly. Moreover, they were successful and not influenced by anyone else. Einstein is superb in this regard and was very unusual—everything that he did was imaginative. His prediction of gravitational waves was validated 100 years later by work conducted at the LIGO (the Laser Interferometer Gravitational Wave Observatory)!
Q: Why did you become a scientist? Did you enjoy being a scientist?
A: When I was a teenager, I thought that the life of a scientist is perfect, especially if you are at a university even though I did not know much about the job. I believed that it would be good to be in science because the amount of material that you use in judging another person's productivity is not susceptible to any kind of fiddling. Also, the person who is producing the material cannot cheat. Twenty years later, when I was in my 30's, I found that those concepts were probably true but they were not really true. I realized that people are the same everywhere. There were good scientists who worked hard and did good experiments, but there [are] also nasty people who got credit because of other people's work.
Yes, I enjoyed being a scientist, partly because my work was successful. If my work had not been successful, I may not have enjoyed it so much.
Q: What makes a laboratory a good environment for trainees?
A: Intelligence of the group leader is important, especially how she/he approaches a research problem. The leader must understand that different students have different ways of showing their intelligence. A good leader must recognize that getting a PhD is tough regardless of one's previous record. The advisor should work with the trainee to solve a problem. Rigid mentor-mentee structures should not exist.
Q: Why do you think the MRC Laboratory of Molecular Biology (LMB), Cambridge, has been such a successful research enterprise?
A: The LMB is a special place where everyone worked at the bench. When I was a postdoc, I enjoyed coming to the LMB every day because it was a conglomeration of people who were dedicated to pushing science forward. Virtually, no one worked in a team. When you gathered for coffee or tea in the morning or afternoon, however, you could discuss anything with anyone, including senior scientists. The general attitude was that you get the right people and give them the freedom to do what they want. Max Perutz promoted this approach (like Bell Labs), and he got the necessary funding. However, I do not know how he [or they] always identified great people.
One important aspect of the LMB was that knowledge/technical advances were shared immediately. For example, when I was at the LMB, typically polyacrylamide gels were run with wide lanes to obtain radiolabeled tRNAs and 5S rRNA. I went to the machine shop and had them make for me a 15-well comb for my gels. I was so excited because this is something new. Soon, everyone at the LMB was using this multiwell comb. I only learnt later that Bill Studier had done this six months before me.
It has been hard to replicate the LMB anywhere else. In the US, each individual is her/his champion. The time spent obtaining funds (unlike the LMB) distracts from doing the science.
Q: What is your opinion about teaching in STEM areas and in the laboratory? Can creativity be nurtured? Do you believe that cultural contexts color the nature and scope of research?
A: I do not have a specific teaching philosophy. Classroom material should be presented in the simplest form and lecture content must connect with the students. For example, when I gave my lecture on human genetics at Yale, I would talk about number of chromosomes and diseases like sickle cell anemia. Many African American students came to see me afterwards to remark that this is the first time they had learned about this disease. These students were interested because I spoke about something that affected their personal lives. Lectures need not be made easy but must cover material that students might be interested in.
For graduate training, the key is for students to be sufficiently competent to become independent. Innovation cannot be nurtured but the ability to observe something new and look at things in a different way is key to becoming independent scientists.
In some cultures, there is an emphasis on listening to older people and doing exactly as instructed. It is important to encourage young people everywhere to come up with their own ideas.
Be not afraid of greatness. Some are born great, some achieve greatness, and others have greatness thrust upon them. (Shakespeare)
Within their own spheres, every person seeks to shape her/his trail to success. Our quest to better understand Sid, a great scientist, left us surprised by the simplicity of his responses. The themes highlighted by Sid—imagination, inventiveness, thinking outside the box, and the appreciation of the uniqueness of individuals—were familiar because these traits are exemplified in the best teachers and researchers. These individuals may not be recognized with prizes and awards, but they embody the same elements of “greatness” as Sid. After reflecting on our interview with Sid, the important takeaway message, rather a reminder, is that marshaling the elements of great scientists latent within all of us may offer a path to a more fulfilling career in science.
We are extremely grateful for the meticulous collation and administrative support provided by Ann Marie Micenmacher. This Tribute paper would not have been possible without her help.
