Profile of Jennifer Lippincott-Schwartz

Even with a microscope, our eyes typically fail to see the bustling activity inside cells, the constant flux known as the steady state. Jennifer Lippincott-Schwartz, elected to the National Academy of Sciences in 2008, has made it her career to reveal the dynamic nature of cells and their organelles.

Her work has debunked notions that organelles, from lysosomes to the Golgi apparatus, are static entities. Most recently, she has focused on cellular sorting and recycling systems.

In her inaugural article (1), Lippincott-Schwartz, a tenured investigator at the National Institutes of Health (NIH, Bethesda, MD), explores how the ubiquitin tag, once thought to be a 1-way ticket to the proteasome, can also, in concert with the p62 protein, efficiently ship proteins and other targets to the autophagosome. The work has implications for improving researchers' understanding of neurodegenerative diseases characterized by the accumulation of cellular debris.

A Grounding in Philosophy

Lippincott-Schwartz was born in 1952 in Manhattan, Kansas, where her father was a professor of chemistry. Her family soon moved to College Park, Maryland, before settling on a farm in northern Virginia for her teenage years. Lippincott-Schwartz had always been interested in science, owing to her early exposure to her father's work.

“We had a periodic table hanging in the kitchen,” she recounted. Moving to the farm, however, where her family kept horses and other animals, awakened her interest in biology.

When the time came for college, Lippincott-Schwartz picked Swarthmore College (Swarthmore, PA). “I assumed that I would be a biology major,” she said, but her alma mater's strong focus on a liberal arts education changed her plans. The intellectual stimulation and interesting people that she found in the biology department had equals in other areas.

She ended up majoring in psychology and philosophy. “When I finished college, I was pretty confused as to what I wanted to do,” she explained. “I felt that I wasn't ready to make any decisions long term.”

Lippincott-Schwartz decided to do something completely different and teach abroad. She taught high school physics, chemistry, and biology at a girls school in Tigoi, Kenya, where her experiences cemented a decision to study biology.

“I saw the problems all around,” she said. In addition to lack of treatment for endemic diseases like malaria, there was a lack of understanding of how to grow crops efficiently. “I quickly realized that I did not want to go into psychology,” she said. “I veered back into the biological world.”

Jennifer Lippincott-Schwartz

Lippincott-Schwartz's future husband, Jonathan Schwartz, now a diplomat with the U.S. State Department, accompanied her on the trip. Upon returning to the United States in 1976, the couple moved to Palo Alto, CA. Lippincott-Schwartz took a job at a private boys' high school teaching chemistry and physics and waited until her husband finished law school before she started her own graduate studies.

“It was a contrast at all levels,” she said, explaining that going from an all-girls school in an impoverished country to an all-boys school in a wealthy community brought her “phenomenal perspective” on the disparities of teaching and learning in the sciences.

Seeing Is Knowing

After several years of teaching, Lippincott-Schwartz entered the master's program in biology at Stanford University (Palo Alto, CA). There she worked with Phil Hanawalt on the biochemistry of DNA repair. But not being able to see what she studied bothered her.

“I never looked under a microscope,” she said. “You do biochemical work and you have no concept of how cells are organized. Such organization may be important for how biochemistry works inside cells.”

The experience shaped her choice about which areas to pursue when she entered the doctoral program in biochemistry at Johns Hopkins University (Baltimore, MD) in 1979. Microscopy techniques turned her on to cell biology, and she joined Douglas Fambrough's lab, which was part of the Carnegie Institution of Embryology.

Lippincott-Schwartz remembers her time at Carnegie as “one of the most important parts of my development.” Because the institute had mostly postdoctoral researchers and principal investigators working in a collaborative environment, she found herself in close association with advanced scientists, and the large number of postdocs meant that she had many mentors.

During her doctoral work, Lippincott-Schwartz gained experience with techniques including electron microscopy, light microscopy, and the relatively new tool of monoclonal antibodies. “They were the hot thing at the time,” she said.

She used the tool to track and quantify how proteins move within cells. Using antibodies as tags to follow cellular proteins, she explained, led her to later recognize the potential power of green fluorescent protein (GFP) as a tracking tool. She and Fambrough used monoclonal antibodies to identify and characterize the lysosomal membrane protein LEP100 (2).

At the time, “most people thought that the lysosome was a dead-end organelle, akin to a garbage disposal with its hydrolytic enzymes,” she explained. But the work with LEP100 showed that lysosomal proteins do not solely reside in the lysosome and that lysosomes have pathways for both intake and output (3).

While still a student, Lippincott-Schwartz began to develop concepts that would drive her future work. Illuminating the dynamic nature of the lysosome, once perceived as static, taught her to ask what a system at steady state really looks like, with ebbs and flows that are not immediately apparent.

Upon graduating from Hopkins in 1986, Lippincott-Schwartz took a postdoctoral position at NIH with Richard Klausner, who studied the secretory pathway. There, she continued to debunk popular notions of static, unchanging organelles.

The Golgi apparatus is a packaging organelle. It takes proteins built in the endoplasmic reticulum (ER) and enzymatically modifies them for delivery outside the cell or to other intracellular destinations. In Klausner's lab, Lippincott-Schwartz treated mammalian cells with brefeldin A (BFA), halting transport of proteins from the ER to the Golgi. Unexpectedly, treatment with BFA caused complete resorption of the Golgi into the ER, and removing the drug from the system allowed the Golgi to regenerate (4, 5). “This demonstrated that the Golgi is not static, but continually being regenerated,” she explained.

Members of the Lippincott-Schwartz laboratory at NIH.

In addition to showing that the Golgi can be made to completely disappear and then reassemble, the results paved the way for understanding the molecular basis for Golgi maintenance, which relies on the binding of peripheral membrane proteins that are sensitive to BFA (6, 7). Using BFA as a tool to selectively block intracellular traffic patterns also allowed Lippincott-Schwartz et al. to investigate how cells control the organization and distribution of endosomes and lysosomes (8).

Seeing the Light

In 1990, Lippincott-Schwartz became a staff fellow at the National Institute of Child Health and Human Development at NIH, beginning her work with GFP, the visible tracking molecule that would fundamentally change the way she studied the interior activities of cells and their organelles. She had read about GFP in a research article (9).

“I knew instantly that if we tagged target pathway molecules with GFP that it could be very valuable for understanding trafficking pathways and for distinguishing different models of organelle disassembly and reassembly,” she said.

The imaging techniques available up to that point were too limited to elucidate answers to explain the organelle activity. “Dyes used to illuminate the cell, like BODIPY and fluorescein, fade quickly, meaning that the cell cannot be imaged for any length of time,” she explained. “In addition, the dyes often give rise to free radicals during imaging, which can damage the cell.”

GFP was functionally inert and could be targeted to specific proteins, making it an excellent measure. Using the fluorescent protein to study membrane trafficking intermediates, her group found the intermediates existed as μm-sized, globular-tubular elements that trafficked along microtubules (10).

Visualizing mitotic nuclear envelope and Golgi breakdown for the first time in live cells, Lippincott-Schwartz's team discovered that nuclear envelope and some Golgi components redistribute into the ER at the beginning of mitosis and then re-emerge out of the ER at the end of mitosis, pointing to a fundamental role of the ER in the biogenesis and maintenance of these organelles (11, 12).

Even with specific molecule labeling, Lippincott-Schwartz still had to contend with the limitations of the imaging techniques available in the early 1990s.

“Microscopes were not very sophisticated at the time,” she said. Although she could see activity in the cell, “it was a real challenge to document. There were no movies.”

She used videotape to record the images, but even showing the videotapes at meetings proved onerous, requiring the set-up of cumbersome video monitors for the audience. “It was not very satisfying,” she said.

Imaging techniques have since advanced to the current state of the art thanks to companies in the confocal microscope industry maintaining relationships with scientists and soliciting feed-back, she said.

Cellular Traffic Patterns

Throughout the early 1990s, Lippincott-Schwartz continued to focus on understanding how pathways move molecules within a cell.

“When you're looking, you see only a steady state,” she said. “So it doesn't clarify how things are moving.”

Lippincott-Schwartz began to look for ways to perturb the cell system in an effort to observe it rebounding to steady state. To this end, her lab refined photobleaching techniques for use with confocal microscopy. One key technique was fluorescence recovery after photobleaching (FRAP), in which the researcher destroys fluorescence with a burst of intense light, thus creating a dark spot in the sample, and observes the diffusion of fresh fluorophores into the bleached region.

This technique allowed Lippincott-Schwartz to observe the speed at which different molecular pools equilibrated. She explained that researchers assumed that “molecules retained in different organelles were static, anchored in some fashion.”

Her work showed that the molecules moved, and they moved fast. The results changed the perception of how molecules are retained within the ER, Golgi, and plasma membrane, because resident molecules are all freely diffusing (13–15).

Lippincott-Schwartz also used FRAP to look at the membrane turnover rates of peripheral membrane proteins that form skeletal-like coats on membranes to drive vesicle budding.

The coat proteins “turn on and off rapidly, on the order of seconds,” she said (16).

Developing a related confocal photobleaching technique called fluorescence loss in photobleaching, Lippincott-Schwartz and colleagues isolated populations of fluorescent proteins associated with specific organelles such as the ER and Golgi to determine whether they exchanged with populations elsewhere in the cell (17–19).

To incorporate the information into kinetic models, Lippincott-Schwartz needed to be able to quantify what she was seeing.

Robert Phair at Integrative BioInformatics (Los Altos, CA) helped her develop ways to simulate and test models of membrane trafficking patterns, based on the residency times and transport rates of proteins in different organelles. Phair, Lippincott-Schwartz, and colleagues used these models to characterize secretory and endocytic trafficking (20, 21).

They put together a model of how the Golgi acts as a transport apparatus, involving membrane partitioning rather than a progressing set of cisterna, as had been the standard model (22). Their approach opened new avenues in understanding the complexities of the Golgi.

A Resolution Revolution

In 2002, under her guidance, Lippincott-Schwartz's postdoctoral fellow George Patterson developed photoactivatable GFP. Tagging this version of GFP to a target molecule installed a switch that allowed a researcher to activate fluorescence with a flash of light (23).

Investigators could now follow molecules through the cell selectively without having to rely on photobleaching.

Patterson and Lippincott-Schwartz tagged lysosomal membrane proteins with photoactivatable GFP. In one experiment, for example, they used light to activate the proteins in a single lysosome. Imaging revealed that the activated proteins move quickly to other lysosomes within the cell.

Photoactivatable GFP also led to an application that Lippincott-Schwartz did not anticipate: the ability to visualize individual molecules at the nanometric scale in a densely populated biological system.

Eric Betzig, now at Howard Hughes Medical Institute's Janelia Farm campus (Ashburn, VA), developed the idea for this kind of “superresolution imaging.” He theorized that switching molecules on 1 at a time and measuring the position of the feedback fluorescence spectrum could show the exact location of a molecule.

“Switching on tens of thousands of molecules, 1 at a time, over the course of an hour, would produce a distribution map of molecules in a fixed cell,” Lippincott-Schwartz explained.

But Betzig had no reagent for isolating single molecules. Photoactivatable GFP provided that function, so when he approached Lippincott-Schwartz in early 2005, they knew that a collaboration might produce something truly revolutionary. Betzig worked in her lab, together with coinventor Harald Hess, to develop a technique they named photoactivation localization microscopy (PALM) (24). For the first time, it became possible to assemble images of organelles at the molecular level.

“The Golgi is not static, but continually being regenerated.”

Lippincott-Schwartz's interest in superresolution microscopy has continued to flourish. Her group introduced single-particle tracking PALM in living cells to track the motions of single molecules in densely packed protein populations typically found in cells (25). The research team also developed a dual-label PALM approach for visualizing the nanometric distribution of 2 populations of molecules simultaneously in cells (26). Continued collaborations with Hess, who invented interferometry PALM (iPALM), have allowed her group to visualize the 3D organization of minute cellular structures and spaces (27).

Targeting for Disposal

The interlinking and overlapping of cellular processes fascinate Lippincott-Schwartz. In her inaugural article (1), she explores how large and small cellular structures are targeted to the autophagosome for destruction.

She first encountered this phenomenon in an experiment in which her group attached red fluorescent protein to ubiquitin, a tag that targets proteins for delivery to the proteasome and thus destruction. They found that the ubiquitin-tagged protein also got targeted to the autophagosome.

Because ubiquitin is typically thought of as the proteasome tag, Lippincott-Schwartz and postdoc Peter Kim sought to investigate this second role of ubiquitin. They found that knocking down p62, a protein that recognizes ubiquitin and attaches it to the autophagosome membrane, causes aggregation of cellular debris in the cytoplasm, an effect seen in neurodegenerative diseases such as Alzheimer's and Huntington's diseases.

Lippincott-Schwartz may be noted for breathing life into seemingly static organelles, but her research on short time scales is motivated by the recognition that biology is essentially a historical study.

“Cells and systems have evolved to be dynamic processes, and this evolutionary principle should guide our inquiries,” she said. Moreover, majoring in philosophy gave her the conviction that knowledge is best built on multiple techniques whenever possible. “Don't rely solely on 1 approach to verify what you found,” she explained.

Openness to different perspectives, whether it be evolutionary or otherwise, is important to Lippincott-Schwartz. She has not forgotten the influence of her exposure to different cultures in Africa. She makes it a priority for members of her family to travel to remote places so they, too, are open to new ways of seeing the world.