Water agglomerates on Fe3O4(001)
Matthias Meier, Jan Hulva, Zdeněk Jakub, Jiří Pavelec, Martin Setvin, Roland Bliem, Michael Schmid, Ulrike Diebold, Cesare Franchini, and Gareth S. Parkinson
Determining the structure of water on metal oxide surfaces is a key step toward a molecular-level understanding of dissolution, corrosion, geochemistry, and catalysis, but hydrogen bonding and large, complex unit cells present a major challenge to modern theory. Here, we utilize state-of-the-art experimental techniques to guide a density functional theory (DFT)-based search for the minimum-energy configurations of water on Fe3O4(001). A subsurface reconstruction dominates adsorption at all coverages. An ordered array of partially dissociated water agglomerates form at low coverage, and these serve to anchor a hydrogen-bonded network. We argue that similar behavior will occur whenever a surface presents a well-spaced array of active sites for dissociation. Given the propensity of metal oxides to undergo surface reconstructions, this is likely often. (See pp. E5642–E5650.)
Simultaneous cancer and tumor microenvironment subtyping using confocal infrared microscopy for all-digital molecular histopathology
Shachi Mittal, Kevin Yeh, L. Suzanne Leslie, Seth Kenkel, Andre Kajdacsy-Balla, and Rohit Bhargava
Cancer alters both the morphological and the biochemical properties of multiple cell types in a tissue. Generally, the morphology of epithelial cells is practical for routine disease diagnoses. Here, infrared spectroscopic imaging biochemically characterizes breast cancer, both epithelial cells and the tumor-associated microenvironment. Unfortunately, conventional spectral analyses are slow. Hence, we designed and built a laser confocal microscope that demonstrates a high signal-to-noise ratio for confident diagnoses. The instrument cuts down imaging time from days to minutes, making the technology feasible for research and clinical translation. Finally, automated human breast cancer biopsy imaging is reported in ∼1 hour, paving the way for routine research into the total tumor (epithelial plus microenvironment) properties and rapid, label-free diagnoses. (See pp. E5651–E5660.)
Exceptionally high levels of lead pollution in the Balkans from the Early Bronze Age to the Industrial Revolution
Jack Longman, Daniel Veres, Walter Finsinger, and Vasile Ersek
A detailed record of historical lead (Pb) pollution from a peat bog in Serbia provides a unique view on the extent and timing of Balkan mining and metallurgy. Evidence of the earliest European environmental pollution is followed by large-scale and sustained increases in the amount of anthropogenically derived Pb after 600 BCE, through the Roman/Byzantine periods, and into the medieval period. Occasional evidence of drops in pollution output reflects the disruptive socioeconomic impact of periods of turmoil. Our data show a trend significantly different to records in western Europe, where Pb pollution decreases dramatically after the collapse of the Roman Empire. These results suggest metal-rich southeastern Europe should be considered a more major player in environmental metal pollution through time. (See pp. E5661–E5668.)
Targeting the potent Beclin 1–UVRAG coiled-coil interaction with designed peptides enhances autophagy and endolysosomal trafficking
Shuai Wu, Yunjiao He, Xianxiu Qiu, Wenchao Yang, Wenchao Liu, Xiaohua Li, Yan Li, Han-Ming Shen, Renxiao Wang, Zhenyu Yue, and Yanxiang Zhao
Beclin 1 is an essential autophagy protein. Through its coiled-coil domain, Beclin 1 recruits two modulators, Atg14L and UVRAG, to form Atg14L- or UVRAG-containing Beclin 1–Vps34 subcomplexes responsible for Vps34-dependent membrane trafficking processes including autophagy and endosomal trafficking. Our structural study of the Beclin 1–UVRAG coiled-coil complex reveals a strengthened interface to maintain potent Beclin 1–UVRAG interaction. This potency is essential for UVRAG to outcompete Atg14L and enhance Vps34-dependent endosomal trafficking. Our designed peptides can target the Beclin 1 coiled-coil domain, promote Atg14L– and UVRAG–Beclin 1 interaction, induce autophagy, and significantly enhance endolysosomal degradation of the EGF receptor. Our results testify to the feasibility of targeting Beclin 1 to regulate both autophagy and endosomal trafficking. (See pp. E5669–E5678.)
Individuality and slow dynamics in bacterial growth homeostasis
Lee Susman, Maryam Kohram, Harsh Vashistha, Jeffrey T. Nechleba, Hanna Salman, and Naama Brenner
Microbial cells go through repeated cycles of growth and division. These cycles are not perfect: the time and size at division can fluctuate from one cycle to the next. Still, cell size is kept tightly controlled, and fluctuations do not accumulate to large deviations. How this control is implemented in single cells is still not fully understood. We performed experiments that follow individual bacteria in microfluidic traps for hundreds of generations. This enables us to identify distinct individual dynamic properties that are maintained over many cycles of growth and division. Surprisingly, we find that each cell suppresses fluctuations with a different strength; this variability defines an “individual” behavior for each cell, which is inherited along many generations. (See pp. E5679–E5687.)
In vitro biomimetic engineering of a human hematopoietic niche with functional properties
Paul E. Bourgine, Thibaut Klein, Anna M. Paczulla, Takafumi Shimizu, Leo Kunz, Konstantinos D. Kokkaliaris, Daniel L. Coutu, Claudia Lengerke, Radek Skoda, Timm Schroeder, and Ivan Martin
The development of an in vitro human bone marrow (BM) tissue appears essential to compile information on human hematopoiesis. Conventional systems fail at both capturing the complexity of the bone marrow niche while allowing the maintenance of functional hematopoietic stem cells (HSCs). Here, we report the development of a human 3D (BM) analogue in a perfusion-based bioreactor system, partially recapitulating structural, compositional, and organizational features of the native human osteoblastic niche environment. The engineered tissue supports the maintenance of some hematopoietic stem and progenitor cell (HSPC) properties. This provides an advanced technological platform of broad fundamental and translational relevance, including the study of human HSPC biology and interactions with their niche, the manipulation of functional human HSPCs, or the identification of factors influencing human hematopoiesis. (See pp. E5688–E5695.)
Spatially modulated ephrinA1:EphA2 signaling increases local contractility and global focal adhesion dynamics to promote cell motility
Zhongwen Chen, Dongmyung Oh, Kabir H. Biswas, Cheng-Han Yu, Ronen Zaidel-Bar, and Jay T. Groves
Cell receptors are neither uniformly distributed nor uniformly activated across plasma membrane. However, very little is known about how their spatial arrangement affects cellular response to receptor signaling. EphA2 is a receptor tyrosine kinase whose activation depends on binding of EphrinA1 ligands on the opposing cell, and EphA2 signaling plays an important role in cancer metastasis. In adherent cells, integrin adhesions are a key element to regulate cell–matrix interactions and cell migration. However, our understanding of how their activities are spatiotemporally coordinated remains superficial. In this study, we combined microfabrication, live imaging, and single-molecule tracking to directly map the signal transduction from ephrinA1:EphA2 complex to integrin adhesions, and revealed a spatially controlled mechanism for EphA2–integrin signaling cross talk. (See pp. E5696–E5705.)
Minimizing ATP depletion by oxygen scavengers for single-molecule fluorescence imaging in live cells
Seung-Ryoung Jung, Yi Deng, Christopher Kushmerick, Charles L. Asbury, Bertil Hille, and Duk-Su Koh
Single-molecule live-cell imaging can answer many biological questions, but a significant obstacle is photobleaching of the dye molecules by dissolved oxygen. An excellent way to improve photostability is to reduce the oxygen content of the solutions using glucose oxidase, an oxygen scavenger. However, for live-cell experiments a critical downside of the scavengers is that ATP levels fall as mitochondria become oxygen-deprived. We show that certain metabolic supplements significantly slow the depletion of ATP by oxygen scavengers and substantially restore ATP-dependent functions like lipid synthesis and receptor endocytosis. This method could be valuable for scientific studies that need to control oxygen tension and intracellular ATP levels in live cells. (See pp. E5706–E5715.)
Automatically identifying, counting, and describing wild animals in camera-trap images with deep learning
Mohammad Sadegh Norouzzadeh, Anh Nguyen, Margaret Kosmala, Alexandra Swanson, Meredith S. Palmer, Craig Packer, and Jeff Clune
Motion-sensor cameras in natural habitats offer the opportunity to inexpensively and unobtrusively gather vast amounts of data on animals in the wild. A key obstacle to harnessing their potential is the great cost of having humans analyze each image. Here, we demonstrate that a cutting-edge type of artificial intelligence called deep neural networks can automatically extract such invaluable information. For example, we show deep learning can automate animal identification for 99.3% of the 3.2 million-image Snapshot Serengeti dataset while performing at the same 96.6% accuracy of crowdsourced teams of human volunteers. Automatically, accurately, and inexpensively collecting such data could help catalyze the transformation of many fields of ecology, wildlife biology, zoology, conservation biology, and animal behavior into “big data” sciences. (See pp. E5716–E5725.)
Directed evolution of multiple genomic loci allows the prediction of antibiotic resistance
Ákos Nyerges, Bálint Csörgő, Gábor Draskovits, Bálint Kintses, Petra Szili, Györgyi Ferenc, Tamás Révész, Eszter Ari, István Nagy, Balázs Bálint, Bálint Márk Vásárhelyi, Péter Bihari, Mónika Számel, Dávid Balogh, Henrietta Papp, Dorottya Kalapis, Balázs Papp, and Csaba Pál
Antibiotic development is frequently plagued by the rapid emergence of drug resistance. However, assessing the risk of resistance development in the preclinical stage is difficult. By building on multiplex automated genome engineering, we developed a method that enables precise mutagenesis of multiple, long genomic segments in multiple species without off-target modifications. Thereby, it enables the exploration of vast numbers of combinatorial genetic alterations in their native genomic context. This method is especially well-suited to screen the resistance profiles of antibiotic compounds. It allowed us to predict the evolution of resistance against antibiotics currently in clinical trials. We anticipate that it will be a useful tool to identify resistance-proof antibiotics at an early stage of drug development. (See pp. E5726–E5735.)
Cell-intrinsic regulation of murine epidermal Langerhans cells by protein S
Yaara Tabib, Nora S. Jaber, Maria Nassar, Tal Capucha, Gabriel Mizraji, Tsipora Nir, Noam Koren, Itay Aizenbud, Avraham Maimon, Luba Eli-Berchoer, Asaf Wilensky, Tal Burstyn-Cohen, and Avi-Hai Hovav
Langerhans cells (LCs) are the exclusive antigen-presenting cells of the epidermis, capable of mounting immunity and tolerance. LCs maintain themselves locally by self-renewing throughout life, a process that is regulated by both LCs and keratinocytes. Nevertheless, the mechanisms underlying this process are not clearly understood. Using targeted genetic ablation, we demonstrate that lack of protein S (PROS1) in keratinocytes, but not in LCs, results in reduced numbers of terminally developed LCs. This is due to increased apoptosis of LCs and associated with altered expression of cytokines involved in tissue homeostasis. Furthermore, PROS1 also down-regulates LC differentiation from bone marrow precursors. Together, these identify PROS1 as a regulator of LC development and homeostasis. (See pp. E5736–E5745.)
Gastrointestinal stromal tumor enhancers support a transcription factor network predictive of clinical outcome
Matthew L. Hemming, Matthew A. Lawlor, Rhamy Zeid, Tom Lesluyes, Jonathan A. Fletcher, Chandrajit P. Raut, Ewa T. Sicinska, Frédéric Chibon, Scott A. Armstrong, George D. Demetri, and James E. Bradner
Enhancers are regulatory regions in DNA that govern gene expression and orchestrate cellular phenotype. We describe the enhancer landscape of gastrointestinal stromal tumor (GIST), identifying established and unique GIST-associated genes that characterize this neoplasm. Focusing on transcriptional regulators, we identify a core group of transcription factors underlying GIST biology. Two transcription factors, BARX1 and HAND1, have mutually exclusive enhancers and expression in localized and metastatic GIST, respectively. HAND1 is necessary to sustain GIST proliferation and KIT expression, and binds to enhancers of GIST-associated genes. The relative expression of BARX1 and HAND1 is predictive of clinical behavior in GIST patients. These results expand our understanding of gene regulation in this disease and identify biomarkers that may be helpful in diagnosis and treatment. (See pp. E5746–E5755.)
A stress-induced response complex (SIRC) shuttles miRNAs, siRNAs, and oligonucleotides to the nucleus
Daniela Castanotto, Xiaowei Zhang, Jessica Alluin, Xizhe Zhang, Jacqueline Rüger, Brian Armstrong, John Rossi, Arthur Riggs, and C. A. Stein
The deregulation of miRNA function is critical in the pathogenesis of cancer and other diseases. miRNAs and other noncoding RNAs (ncRNAs) tightly regulate gene expression, often in the cell nucleus. Heretofore, there has been no understanding that there exists a general shuttling mechanism that brings miRNAs, in addition to therapeutic oligonucleotides and siRNAs, from the cytoplasm into the nucleus. We have identified this shuttling mechanism, which occurs in response to cell stress. Nuclear imported miRNAs are functional, can potentially alter gene expression, and participate in cell stress response mechanisms. This shuttling mechanism can be augmented to target specific RNAs, including miRNA sponges, and long ncRNAs like Malat-1, which have been implicated in promoting tumor metastasis. (See pp. E5756–E5765.)
PHIP as a therapeutic target for driver-negative subtypes of melanoma, breast, and lung cancer
David de Semir, Vladimir Bezrookove, Mehdi Nosrati, Altaf A. Dar, Clayton Wu, Julia Shen, Christopher Rieken, Meenakshi Venkatasubramanian, James R. Miller III, Pierre-Yves Desprez, Sean McAllister, Liliana Soroceanu, Robert J. Debs, Nathan Salomonis, Dirk Schadendorf, James E. Cleaver, and Mohammed Kashani-Sabet
The development of targeted therapies represents a major advance in cancer therapy, with its most prominent examples including agents targeting HER2 in breast cancer, EGFR and ALK in lung cancer, and BRAF in melanoma. However, a key challenge confronting such precision medicine approaches concerns the presence of subtypes of each of these three common, lethal solid tumors that lack identified molecular drivers, and are thus not amenable to targeted therapies. Our studies identify a role for pleckstrin homology domain-interacting protein (PHIP) in promoting the progression of “driver-negative” subtypes of these common solid tumors. In addition, they demonstrate a physical interaction between PHIP and an activating histone modification, thereby identifying PHIP as a rational target for the therapy of these solid tumor subtypes. (See pp. E5766–E5775.)
ABC transporter content diversity in Streptococcus pneumoniae impacts competence regulation and bacteriocin production
Charles Y. Wang, Nisha Patel, Wei-Yun Wholey, and Suzanne Dawid
The opportunistic pathogen Streptococcus pneumoniae (pneumococcus) participates in horizontal gene transfer through genetic competence and produces antimicrobial peptides called “bacteriocins.” Here, we show that the competence and bacteriocin-related ABC transporters ComAB and BlpAB share the same substrate pool, resulting in bidirectional crosstalk between competence and bacteriocin regulation. We also clarify the role of each transporter in bacteriocin secretion and show that, based on their transporter content, pneumococcal strains can be separated into a majority opportunist group that uses bacteriocins only to support competence and a minority aggressor group that uses bacteriocins in broader contexts. Our findings will impact how bacteriocin regulation and production is modeled in the many other bacterial species that use ComAB/BlpAB-type transporters. (See pp. E5776–E5785.)
Comprehensive skin microbiome analysis reveals the uniqueness of human skin and evidence for phylosymbiosis within the class Mammalia
Ashley A. Ross, Kirsten M. Müller, J. Scott Weese, and Josh D. Neufeld
Skin forms a critical protective barrier between a mammal and its external environment. Baseline data on the mammalian skin microbiome elucidates which microorganisms are found on healthy skin and provides insight into mammalian evolutionary history. To our knowledge, this study represents the largest existing mammalian skin microbiome survey. Our findings demonstrate that human skin is distinct, not only from other Primates, but from all 10 mammalian orders sampled. Identifying significant similarities between branching of mammalian phylogenetic trees and relatedness trees for their corresponding microbial communities raises the possibility that mammals have experienced coevolution between skin microbiota and their corresponding host species. (See pp. E5786–E5795.)
Stimulation-induced increases in cerebral blood flow and local capillary vasoconstriction depend on conducted vascular responses
Changsi Cai, Jonas C. Fordsmann, Sofie H. Jensen, Bodil Gesslein, Micael Lønstrup, Bjørn O. Hald, Stefan A. Zambach, Birger Brodin, and Martin J. Lauritzen
Pericytes are located at the outside wall of capillaries. However, whether and how pericytes are involved in the regulation of blood flow in brain capillaries is still debated. We report that capillary vascular responses are mostly initiated and peak at near-arteriole capillaries. These vascular responses are conducted along capillaries at a speed of 5–20 µm/s. Conducted vascular responses in brain capillaries appear to involve pericytes, the mural cells of microvessels, and may be a novel modulator of vascular function in the brain. (See pp. E5796–E5804.)
Rapid, experience-dependent translation of neurogranin enables memory encoding
Kendrick J. Jones, Sebastian Templet, Khaled Zemoura, Bozena Kuzniewska, Franciso X. Pena, Hongik Hwang, Ding J. Lei, Henny Haensgen, Shannon Nguyen, Christopher Saenz, Michael Lewis, Magdalena Dziembowska, and Weifeng Xu
De novo protein synthesis is critical for memory formation. We found that protein synthesis during acquisition is transiently required for contextual memory formation. We identified one candidate gene, Nrgn (encoding protein neurogranin, Ng) with enhanced translation upon novel-context exposure, and found that experience-dependent translation of Ng in the hippocampus is required for contextual memory formation. Furthermore, fragile-X mental retardation protein interacts with the 3′UTR of the Nrgn mRNA, which is required for activity-dependent translation of Ng in the synaptic compartment and contextual memory formation. Together, these results indicate that experience-dependent and acute translation of Ng in the hippocampus during memory acquisition enables durable context memory encoding. (See pp. E5805–E5814.)
Soluble epoxide hydrolase plays a key role in the pathogenesis of Parkinson’s disease
Qian Ren, Min Ma, Jun Yang, Risa Nonaka, Akihiro Yamaguchi, Kei-ichi Ishikawa, Kenta Kobayashi, Shigeo Murayama, Sung Hee Hwang, Shinji Saiki, Wado Akamatsu, Nobutaka Hattori, Bruce D. Hammock, and Kenji Hashimoto
Parkinson’s disease (PD) is a chronic and progressive movement disorder; however, the precise mechanisms of its etiology remain largely unknown. Soluble epoxide hydrolase (sEH) plays a key role in the inflammation associated with PD pathogenesis. The sEH inhibitor or deletion of the sEH gene protected against MPTP-induced neurotoxicity in mouse brain. Furthermore, expression of the sEH protein (or mRNA) was higher in the striatum of MPTP-treated mice, patients with dementia of Lewy bodies (DLB), and neurons from iPSCs of a PD patient with PARKIN mutations. Interestingly, treatment with sEH inhibitor protected against apoptosis in human PARK2 iPSC-derived dopaminergic neurons. Our findings indicate that sEH inhibitors or epoxy fatty acids mimics may be promising prophylactic or therapeutic drugs for PD. (See pp. E5815–E5823.)
A shared cis-regulatory module activates transcription in the suspensor of plant embryos
Kelli F. Henry, Anhthu Q. Bui, Tomokazu Kawashima, and Robert B. Goldberg
Little is known about how genes are expressed in different plant embryo regions. We tested the hypothesis that shared cis-regulatory motifs control the transcription of genes specifically in the suspensor. We carried out functional studies with the Scarlet Runner Bean (SRB) GA 20-oxidase gene that encodes a gibberellic acid (GA) hormone biosynthesis enzyme and is expressed specifically within the suspensor. We show that cis-regulatory motifs required for GA 20-oxidase transcription within the suspensor are the same as those required for suspensor-specific transcription of the SRB G564 gene, although motif number, spacing, and order differ. These cis-elements constitute a control module that is required to activate genes in the SRB suspensor and may form part of a suspensor regulatory network. (See pp. E5824–E5833.)