nils-herrmann/clean_scirepeval huggingface dataset:
Resident microbial communities inhibit growth and antibiotic-resistance evolution of Escherichia coli in human gut microbiome samples
Venue: PLoS biology

Countering the rise of antibiotic-resistant pathogens requires improved understanding of how resistance emerges and spreads in individual species, which are often embedded in complex microbial communities such as the human gut microbiome. Interactions with other microorganisms in such communities might suppress growth and resistance evolution of individual species (e.g., via resource competition) but could also potentially accelerate resistance evolution via horizontal transfer of resistance genes. It remains unclear how these different effects balance out, partly because it is difficult to observe them directly. Here, we used a gut microcosm approach to quantify the effect of three human gut microbiome communities on growth and resistance evolution of a focal strain of Escherichia coli. We found the resident microbial communities not only suppressed growth and colonisation by focal E. coli but also prevented it from evolving antibiotic resistance upon exposure to a beta-lactam antibiotic. With samples from all three human donors, our focal E. coli strain only evolved antibiotic resistance in the absence of the resident microbial community, even though we found resistance genes, including a highly effective resistance plasmid, in resident microbial communities. We identified physical constraints on plasmid transfer that can explain why our focal strain failed to acquire some of these beneficial resistance genes, and we found some chromosomal resistance mutations were only beneficial in the absence of the resident microbiota. This suggests, depending on in situ gene transfer dynamics, interactions with resident microbiota can inhibit antibiotic-resistance evolution of individual species.
Visualization of two architectures in class-II CAP-dependent transcription activation
Venue: PLoS biology

Transcription activation by cyclic AMP (cAMP) receptor protein (CAP) is the classic paradigm of transcription regulation in bacteria. CAP was suggested to activate transcription on class-II promoters via a recruitment and isomerization mechanism. However, whether and how it modifies RNA polymerase (RNAP) to initiate transcription remains unclear. Here, we report cryo–electron microscopy (cryo-EM) structures of an intact Escherichia coli class-II CAP-dependent transcription activation complex (CAP-TAC) with and without de novo RNA transcript. The structures reveal two distinct architectures of TAC and raise the possibility that CAP binding may induce substantial conformational changes in all the subunits of RNAP and transiently widen the main cleft of RNAP to facilitate DNA promoter entering and formation of the initiation open complex. These structural changes vanish during further RNA transcript synthesis. The observations in this study may reveal a possible on-pathway intermediate and suggest a possibility that CAP activates transcription by inducing intermediate state, in addition to the previously proposed stabilization mechanism.
Portable sequencing as a teaching tool in conservation and biodiversity research
Venue: PLoS biology

As biodiversity loss continues to accelerate, there is a critical need for education and biomonitoring across the globe. Portable technologies allow for in situ molecular biodiversity monitoring that has been historically out of reach for many researchers in habitat nations. In the realm of education, portable tools such as DNA sequencers facilitate in situ hands-on training in real-time sequencing and interpretation techniques. Here, we provide step-by-step protocols as a blueprint for a terrestrial conservation genetics field training program that uses low-cost, portable devices to conduct genomics-based training directly in biodiverse habitat countries.
A pedagogical approach to science outreach
Venue: PLoS biology

Encouragement of students across all communities through scientific outreach programs is critical to engaging the next generation, exciting young minds to pursue careers in science and medicine. Herein, we present a uniquely structured and widely influential science outreach program. Founded in 2005, the Duke Chemistry Outreach (DCO) employs a pedagogical approach to outreach that aims to teach its audience a new scientific concept, while instilling a pure enjoyment of science. DCO has performed 583 events reaching over 70,000 participants throughout 2,270 hours, with the majority of events in Durham, the surrounding North Carolinian communities, and across 8 other states. The flexibility and diversity of this outreach program creates a framework amendable for others to adopt in both secondary and higher education settings.
Ultralow-frequency neural entrainment to pain
Venue: PLoS biology

Nervous systems exploit regularities in the sensory environment to predict sensory input, adjust behavior, and thereby maximize fitness. Entrainment of neural oscillations allows retaining temporal regularities of sensory information, a prerequisite for prediction. Entrainment has been extensively described at the frequencies of periodic inputs most commonly present in visual and auditory landscapes (e.g., >0.5 Hz). An open question is whether neural entrainment also occurs for regularities at much longer timescales. Here, we exploited the fact that the temporal dynamics of thermal stimuli in natural environment can unfold very slowly. We show that ultralow-frequency neural oscillations preserved a long-lasting trace of sensory information through neural entrainment to periodic thermo-nociceptive input as low as 0.1 Hz. Importantly, revealing the functional significance of this phenomenon, both power and phase of the entrainment predicted individual pain sensitivity. In contrast, periodic auditory input at the same ultralow frequency did not entrain ultralow-frequency oscillations. These results demonstrate that a functionally significant neural entrainment can occur at temporal scales far longer than those commonly explored. The non-supramodal nature of our results suggests that ultralow-frequency entrainment might be tuned to the temporal scale of the statistical regularities characteristic of different sensory modalities.
Precise genomic mapping of 5-hydroxymethylcytosine via covalent tether-directed sequencing
Venue: PLoS biology

5-hydroxymethylcytosine (5hmC) is the most prevalent intermediate on the oxidative DNA demethylation pathway and is implicated in regulation of embryogenesis, neurological processes, and cancerogenesis. Profiling of this relatively scarce genomic modification in clinical samples requires cost-effective high-resolution techniques that avoid harsh chemical treatment. Here, we present a bisulfite-free approach for 5hmC profiling at single-nucleotide resolution, named hmTOP-seq (5hmC-specific tethered oligonucleotide–primed sequencing), which is based on direct sequence readout primed at covalently labeled 5hmC sites from an in situ tethered DNA oligonucleotide. Examination of distinct conjugation chemistries suggested a structural model for the tether-directed nonhomologous polymerase priming enabling theoretical evaluation of suitable tethers at the design stage. The hmTOP-seq procedure was optimized and validated on a small model genome and mouse embryonic stem cells, which allowed construction of single-nucleotide 5hmC maps reflecting subtle differences in strand-specific CG hydroxymethylation. Collectively, hmTOP-seq provides a new valuable tool for cost-effective and precise identification of 5hmC in characterizing its biological role and epigenetic changes associated with human disease.
Rab5c-mediated endocytic trafficking regulates hematopoietic stem and progenitor cell development via Notch and AKT signaling
Venue: PLoS biology

It is well known that various developmental signals play diverse roles in hematopoietic stem and progenitor cell (HSPC) production; however, how these signaling pathways are orchestrated remains incompletely understood. Here, we report that Rab5c is essential for HSPC specification by endocytic trafficking of Notch and AKT signaling in zebrafish embryos. Rab5c deficiency leads to defects in HSPC production. Mechanistically, Rab5c regulates hemogenic endothelium (HE) specification by endocytic trafficking of Notch ligands and receptor. We further show that the interaction between Rab5c and Appl1 in the endosome is required for the survival of HE in the ventral wall of the dorsal aorta through AKT signaling. Interestingly, Rab5c overactivation can also lead to defects in HSPC production, which is attributed to excessive endolysosomal trafficking inducing Notch signaling defect. Taken together, our findings establish a previously unrecognized role of Rab5c-mediated endocytic trafficking in HSPC development and provide new insights into how spatiotemporal signals are orchestrated to accurately execute cell fate transition.
ORANGE: A CRISPR/Cas9-based genome editing toolbox for epitope tagging of endogenous proteins in neurons
Venue: PLoS biology

The correct subcellular distribution of proteins establishes the complex morphology and function of neurons. Fluorescence microscopy techniques are invaluable to investigate subcellular protein distribution, but they suffer from the limited ability to efficiently and reliably label endogenous proteins with fluorescent probes. We developed  Resource for the Application of Neuronal Genome Editing, which mediates targeted genomic integration of epitope tags in rodent dissociated neuronal culture, in organotypic slices, and in vivo. ORANGE includes a knock-in library for in-depth investigation of endogenous protein distribution, viral vectors, and a detailed two-step cloning protocol to develop knock-ins for novel targets. Using ORANGE with (live-cell) superresolution microscopy, we revealed the dynamic nanoscale organization of endogenous neurotransmitter receptors and synaptic scaffolding proteins, as well as previously uncharacterized proteins. Finally, we developed a mechanism to create multiple knock-ins in neurons, mediating multiplex imaging of endogenous proteins. Thus, ORANGE enables quantification of expression, distribution, and dynamics for virtually any protein in neurons at nanoscale resolution.
Crosslinking-guided geometry of a complete CXC receptor-chemokine complex and the basis of chemokine subfamily selectivity
Venue: PLoS biology

Chemokines and their receptors are orchestrators of cell migration in humans. Because dysregulation of the receptor-chemokine system leads to inflammation and cancer, both chemokines and receptors are highly sought therapeutic targets. Yet one of the barriers for their therapeutic targeting is the limited understanding of the structural principles behind receptor-chemokine recognition and selectivity. The existing structures do not include CXC subfamily complexes and lack information about the receptor distal N-termini, despite the importance of the latter in signaling, regulation, and bias. Here, we report the discovery of the geometry of the complex between full-length CXCR4, a prototypical CXC receptor and driver of cancer metastasis, and its endogenous ligand CXCL12. By comprehensive disulfide cross-linking, we establish the existence and the structure of a novel interface between the CXCR4 distal N-terminus and CXCL12 β1-strand, while also recapitulating earlier findings from nuclear magnetic resonance, modeling and crystallography of homologous receptors. A cross-linking–informed high-resolution model of the CXCR4-CXCL12 complex pinpoints the interaction determinants and reveals the occupancy of the receptor major subpocket by the CXCL12 proximal N terminus. This newly found positioning of the chemokine proximal N-terminus provides a structural explanation of CXC receptor-chemokine selectivity against other subfamilies. Our findings challenge the traditional two-site understanding of receptor-chemokine recognition, suggest the possibility of new affinity and signaling determinants, and fill a critical void on the structural map of an important class of therapeutic targets. These results will aid the rational design of selective chemokine-receptor targeting small molecules and biologics with novel pharmacology.
YTHDF2 promotes mitotic entry and is regulated by cell cycle mediators
Venue: PLoS biology

The N6-methyladenosine (m6A) modification regulates mRNA stability and translation. Here, we show that transcriptomic m6A modification can be dynamic and the m6A reader protein YTH N6-methyladenosine RNA binding protein 2 (YTHDF2) promotes mRNA decay during cell cycle. Depletion of YTHDF2 in HeLa cells leads to the delay of mitotic entry due to overaccumulation of negative regulators of cell cycle such as Wee1-like protein kinase (WEE1). We demonstrate that WEE1 transcripts contain m6A modification, which promotes their decay through YTHDF2. Moreover, we found that YTHDF2 protein stability is dependent on cyclin-dependent kinase 1 (CDK1) activity. Thus, CDK1, YTHDF2, and WEE1 form a feedforward regulatory loop to promote mitotic entry. We further identified Cullin 1 (CUL1), Cullin 4A (CUL4A), damaged DNA-binding protein 1 (DDB1), and S-phase kinase-associated protein 2 (SKP2) as components of E3 ubiquitin ligase complexes that mediate YTHDF2 proteolysis. Our study provides insights into how cell cycle mediators modulate transcriptomic m6A modification, which in turn regulates the cell cycle.
Single-cell mapping reveals new markers and functions of lymphatic endothelial cells in lymph nodes
Venue: PLoS biology

Lymph nodes (LNs) are highly organized secondary lymphoid organs that mediate adaptive immune responses to antigens delivered via afferent lymphatic vessels. Lymphatic endothelial cells (LECs) line intranodal lymphatic sinuses and organize lymph and antigen distribution. LECs also directly regulate T cells, mediating peripheral tolerance to self-antigens, and play a major role in many diseases, including cancer metastasis. However, little is known about the phenotypic and functional heterogeneity of LN LECs. Using single-cell RNA sequencing, we comprehensively defined the transcriptome of LECs in murine skin-draining LNs and identified new markers and functions of distinct LEC subpopulations. We found that LECs residing in the subcapsular sinus (SCS) have an unanticipated function in scavenging of modified low-density lipoprotein (LDL) and also identified a specific cortical LEC subtype implicated in rapid lymphocyte egress from LNs. Our data provide new, to our knowledge, insights into the diversity of LECs in murine LNs and a rich resource for future studies into the regulation of immune responses by LN LECs.
Neurophysiological evidence for crossmodal (face-name) person-identity representation in the human left ventral temporal cortex
Venue: PLoS biology

Putting a name to a face is a highly common activity in our daily life that greatly enriches social interactions. Although this specific person–identity association becomes automatic with learning, it remains difficult and can easily be disrupted in normal circumstances or neurological conditions. To shed light on the neural basis of this important and yet poorly understood association between different input modalities in the human brain, we designed a crossmodal frequency-tagging paradigm coupled to brain activity recording via scalp and intracerebral electroencephalography. In Experiment 1, 12 participants were presented with variable pictures of faces and written names of a single famous identity at a 4-Hz frequency rate while performing an orthogonal task. Every 7 items, another famous identity appeared, either as a face or a name. Robust electrophysiological responses were found exactly at the frequency of identity change (i.e., 4 Hz / 7 = 0.571 Hz), suggesting a crossmodal neural response to person identity. In Experiment 2 with twenty participants, two control conditions with periodic changes of identity for faces or names only were added to estimate the contribution of unimodal neural activity to the putative crossmodal face-name responses. About 30% of the response occurring at the frequency of crossmodal identity change over the left occipito-temporal cortex could not be accounted for by the linear sum of unimodal responses. Finally, intracerebral recordings in the left ventral anterior temporal lobe (ATL) in 7 epileptic patients tested with this paradigm revealed a small number of “pure” crossmodal responses, i.e., with no response to changes of identity for faces or names only. Altogether, these observations provide evidence for integration of verbal and nonverbal person identity-specific information in the human brain, highlighting the contribution of the left ventral ATL in the automatic retrieval of face-name identity associations.
BigBrain 3D atlas of cortical layers: Cortical and laminar thickness gradients diverge in sensory and motor cortices
Venue: PLoS biology

Histological atlases of the cerebral cortex, such as those made famous by Brodmann and von Economo, are invaluable for understanding human brain microstructure and its relationship with functional organization in the brain. However, these existing atlases are limited to small numbers of manually annotated samples from a single cerebral hemisphere, measured from 2D histological sections. We present the first whole-brain quantitative 3D laminar atlas of the human cerebral cortex. It was derived from a 3D histological atlas of the human brain at 20-micrometer isotropic resolution (BigBrain), using a convolutional neural network to segment, automatically, the cortical layers in both hemispheres. Our approach overcomes many of the historical challenges with measurement of histological thickness in 2D, and the resultant laminar atlas provides an unprecedented level of precision and detail. We utilized this BigBrain cortical atlas to test whether previously reported thickness gradients, as measured by MRI in sensory and motor processing cortices, were present in a histological atlas of cortical thickness and which cortical layers were contributing to these gradients. Cortical thickness increased across sensory processing hierarchies, primarily driven by layers III, V, and VI. In contrast, motor-frontal cortices showed the opposite pattern, with decreases in total and pyramidal layer thickness from motor to frontal association cortices. These findings illustrate how this laminar atlas will provide a link between single-neuron morphology, mesoscale cortical layering, macroscopic cortical thickness, and, ultimately, functional neuroanatomy.
Every fifth published metagenome is not available to science
Venue: PLoS biology

Have you ever sought to use metagenomic DNA sequences reported in scientific publications? Were you successful? Here, we reveal that metagenomes from no fewer than 20% of the papers found in our literature search, published between 2016 and 2019, were not deposited in a repository or were simply inaccessible. The proportion of inaccessible data within the literature has been increasing year-on-year. Noncompliance with Open Data is best predicted by the scientific discipline of the journal. The number of citations, journal type (e.g., Open Access or subscription journals), and publisher are not good predictors of data accessibility. However, many publications in high–impact factor journals do display a higher likelihood of accessible metagenomic data sets. Twenty-first century science demands compliance with the ethical standard of data sharing of metagenomes and DNA sequence data more broadly. Data accessibility must become one of the routine and mandatory components of manuscript submissions—a requirement that should be applicable across the increasing number of disciplines using metagenomics. Compliance must be ensured and reinforced by funders, publishers, editors, reviewers, and, ultimately, the authors.
Thinking small: Next-generation sensor networks close the size gap in vertebrate biologging
Venue: PLoS biology

Recent advances in animal tracking technology have ushered in a new era in biologging. However, the considerable size of many sophisticated biologging devices restricts their application to larger animals, whereas older techniques often still represent the state-of-the-art for studying small vertebrates. In industrial applications, low-power wireless sensor networks (WSNs) fulfill requirements similar to those needed to monitor animal behavior at high resolution and at low tag mass. We developed a wireless biologging network (WBN), which enables simultaneous direct proximity sensing, high-resolution tracking, and long-range remote data download at tag masses of 1 to 2 g. Deployments to study wild bats created social networks and flight trajectories of unprecedented quality. Our developments highlight the vast capabilities of WBNs and their potential to close an important gap in biologging: fully automated tracking and proximity sensing of small animals, even in closed habitats, at high spatial and temporal resolution.
Over a century of cancer research: Inconvenient truths and promising leads
Venue: PLoS biology

Despite over a century of intensive efforts, the great gains promised by the War on Cancer nearly 50 years ago have not materialized. Since 1999, we have analyzed the lack of progress in explaining and “curing” cancer by examining the merits of the premises that determine how cancer is understood and treated. Our ongoing critical analyses have aimed at clarifying the sources of misunderstandings at the root of the cancer puzzle while providing a plausible and comprehensive biomedical perspective as well as a new theory of carcinogenesis that is compatible with evolutionary theory. In this essay, we explain how this new theory, the tissue organization field theory (TOFT), can help chart a path to progress for cancer researchers by explaining features of cancer that remain unexplainable from the perspective of the still hegemonic somatic mutation theory (SMT) and its variants. Of equal significance, the premises underlying the TOFT offer new perspectives on basic biological phenomena.
Dynamics in the murine norovirus capsid revealed by high-resolution cryo-EM
Venue: PLoS biology

Icosahedral viral capsids must undergo conformational rearrangements to coordinate essential processes during the viral life cycle. Capturing such conformational flexibility has been technically challenging yet could be key for developing rational therapeutic agents to combat infections. Noroviruses are nonenveloped, icosahedral viruses of global importance to human health. They are a common cause of acute gastroenteritis, yet no vaccines or specific antiviral agents are available. Here, we use genetics and cryo-electron microscopy (cryo-EM) to study the high-resolution solution structures of murine norovirus as a model for human viruses. By comparing our 3 structures (at 2.9- to 3.1-Å resolution), we show that whilst there is little change to the shell domain of the capsid, the radiating protruding domains are flexible, adopting distinct states both independently and synchronously. In doing so, the capsids sample a range of conformational space, with implications for maintaining virion stability and infectivity.
A compact Cas9 ortholog from Staphylococcus Auricularis (SauriCas9) expands the DNA targeting scope
Venue: PLoS biology

Compact CRISPR/Cas9 systems that can be packaged into an adeno-associated virus (AAV) hold great promise for gene therapy. Unfortunately, currently available small Cas9 nucleases either display low activity or require a long protospacer adjacent motif (PAM) sequence, limiting their extensive applications. Here, we screened a panel of Cas9 nucleases and identified a small Cas9 ortholog from Staphylococcus auricularis (SauriCas9), which recognizes a simple NNGG PAM, displays high activity for genome editing, and is compact enough to be packaged into an AAV for genome editing. Moreover, the conversion of adenine and cytosine bases can be achieved by fusing SauriCas9 to the cytidine and adenine deaminase. Therefore, SauriCas9 holds great potential for both basic research and clinical applications.