bluuebunny/crossref_metadata_2025_split huggingface dataset:
Interface-guided phenotyping of coding variants in the transcription factor RUNX1 with SEUSS
Authors: Ozturk Kivilcim , Panwala Rebecca , Sheen Jeanna , Ford Kyle , Payne Nathan , Zhang Dong-Er , Hutter Stephan , Haferlach Torsten , Ideker Trey , Mali Prashant , Carter Hannah 
Published: August 2023

AbstractUnderstanding the consequences of single amino acid substitutions in cancer driver genes remains an unmet need. Perturb-seq provides a tool to investigate the effects of individual mutations on cellular programs. Here we deploy SEUSS, a Perturb-seq like approach, to generate and assay mutations at physical interfaces of the RUNX1 Runt domain. We measured the impact of 115 mutations on RNA profiles in single myelogenous leukemia cells and used the profiles to categorize mutations into three functionally distinct groups: wild-type (WT)-like, loss-of-function (LOF)-like and hypomorphic. Notably, the largest concentration of functional mutations (non-WT-like) clustered at the DNA binding site and contained many of the more frequently observed mutations in human cancers. Hypomorphic variants shared characteristics with loss of function variants but had gene expression profiles indicative of response to neural growth factor and cytokine recruitment of neutrophils. Additionally, DNA accessibility changes upon perturbations were enriched for RUNX1 binding motifs, particularly near differentially expressed genes. Overall, our work demonstrates the potential of targeting protein interaction interfaces to better define the landscape of prospective phenotypes reachable by amino acid substitutions.
The essential genome ofXanthomonas citri
Authors: Wang Xiaolan , Wu Manying , Ge Yifei , Lv Weiwei , Liu Chaoying , Ding Xiaojun , Zhang Yu , Wang Jihua , Zhang Yunzeng , Li Lei , Zhou Xiaofeng 
Published: August 2023

AbstractCitrus canker, caused by the bacteriumXanthomonas citrisubsp.citri, is a devastating disease with significant economic implications for the citrus industry worldwide. Understanding the molecular basis ofXanthomonascell cycle and identifying therapeutic targets is crucial for effective disease management. In this study, we employed hyper-saturated transposon mutagenesis combined with high-throughput sequencing to determine the essential features of theXanthomonas citrigenome at ∼7-bp resolution. Our analysis revealed 525 essential genes, 181 high fitness cost genes, 7 small non-coding RNAs, 25 transfer RNAs, 4 ribosomal RNAs, and the origin of replication. Notably, the use of a newly designed Tn5 transposon with an outward pointinglacpromoter significantly reduced false positives caused by polar effects associated with conventional transposons. Functional enrichment analysis showed that essential genes were significantly enriched in processes related to ribosome biogenesis, energy production and conversion, and membrane metabolism. Interestingly, the distribution of essential genes inX. citrishowed similarities to that of the model organismE. coli, suggesting a conserved mode of genome organization that influences transposon accessibility. Our comprehensive analysis provides valuable target genes for potential therapeutic interventions against citrus canker and other related plant diseases.
Seed or soil: tracing back the plant mycobiota primary sources
Authors: Laurent-Webb Liam , Maurice Kenji , Perez-Lamarque Benoît , Bourceret Amélia , Ducousso Marc , Selosse Marc-André 
Published: August 2023

AbstractPlants host diverse communities of fungi (collectively called the mycobiota) which play crucial roles in their development. The assembly processes of the mycobiota, however, remain poorly understood, in particular, whether it is transmitted by parents through the seeds (vertical transmission) or recruited in the environment (horizontal transmission). Here we attempt to quantify the relative contributions of horizontal and vertical transmission in the mycobiota assembly of a desert shrub,Haloxylon salicornicum,by comparing the mycobiota ofin situbulk soil and seeds to that of (i)in situadult individuals and (ii)in vitro-germinated seedlings in soil collectedin situ, either autoclaved or not. We show that the mycobiota is partially transmitted through the seeds to seedlings. In contrast, root mycobiota of adults are highly similar to that of bulk soil, whereas adult leaf mycobiota remain similar to that of seeds. Thus, the mycobiota is transmitted both horizontally and vertically depending on the plant tissue. Despite discrepancies betweenin situandin vitroapproaches, our result may also suggest a compositional turnover in plant mycobiota during plant development. Understanding the respective contribution of these transmission paths to the plant mycobiota is fundamental to deciphering potential coevolutionary processes between plants and fungi.
hGRAD – a versatile ʹone-fits-allʹ system for the acute depletion of RNA binding proteins in nuclear condensates
Authors: Arnold Benjamin , Riegger Ricarda J. , Okuda Ellen Kazumi , Slišcović Irena , Keller Mario , McNicoll François , Zarnack Kathi , Müller-McNicoll Michaela 
Published: August 2023

AbstractNuclear RNA binding proteins (RBPs) are difficult to study because they often belong to large protein families and form extensive networks of auto- and cross- regulation. They are highly abundant and often localize to condensates with a slow turnover, requiring long depletion times or knockouts that cannot distinguish between direct and indirect or compensatory effects.Here, we developed a system that is optimized for the rapid degradation of nuclear RBPs, called hGRAD. It comes as a ʹone-fits-allʹ plasmid, and integration into any cell line that expresses endogenously GFP-tagged proteins allows an inducible, rapid and complete knockdown. We show that the nuclear RBPs SRSF3, SRSF5, SRRM2 and NONO are completely cleared from nuclear speckles and paraspeckles within two hours. hGRAD works in various cell types, is more efficient than other methods and does not require the expression of exogenous ubiquitin ligases. Combining SRSF5 hGRAD degradation with Nascent-seq uncovered highly dynamic transient transcript changes, compensatory mechanisms and that SRSF5 promotes transcript stability.
Rapid and automated design of two-component protein nanomaterials using ProteinMPNN
Authors: de Haas Robbert J. , Brunette Natalie , Goodson Alex , Dauparas Justas , Yi Sue Y. , Yang Erin C. , Dowling Quinton , Nguyen Hannah , Kang Alex , Bera Asim K. , Sankaran Banumathi , de Vries Renko , Baker David , King Neil P. 
Published: August 2023

AbstractThe design of novel protein-protein interfaces using physics-based design methods such as Rosetta requires substantial computational resources and manual refinement by expert structural biologists. A new generation of deep learning methods promises to simplify protein-protein interface design and enable its application to a wide variety of problems by researchers from various scientific disciplines. Here we test the ability of a deep learning method for protein sequence design, ProteinMPNN, to design two-component tetrahedral protein nanomaterials and benchmark its performance against Rosetta. ProteinMPNN had a similar success rate to Rosetta, yielding 13 new experimentally confirmed assemblies, but required orders of magnitude less computation and no manual refinement. The interfaces designed by ProteinMPNN were substantially more polar than those designed by Rosetta, which facilitatedin vitroassembly of the designed nanomaterials from independently purified components. Crystal structures of several of the assemblies confirmed the accuracy of the design method at high resolution. Our results showcase the potential of deep learning-based methods to unlock the widespread application of designed protein-protein interfaces and self-assembling protein nanomaterials in biotechnology.
Time-resolved X-ray solution scattering unveils the sequence of events leading to human Hb heme capture byStaphylococcus aureusIsdB
Authors: Bei Omar De , Marchetti Marialaura , Guglielmo Stefano , Gianquinto Eleonora , Spyrakis Francesca , Campanini Barbara , Bettati Stefano , Levantino Matteo , Ronda Luca 
Published: August 2023

AbstractInfections caused byStaphylococcus aureusdepend on its ability to acquire nutrients. One essential nutrient is iron, which is obtained from the heme of the human host hemoglobin (Hb) through a protein machinery called Iron-regulated Surface Determinant (Isd). IsdB is the protein in charge of heme extraction from Hb, which is the first step of the chain of events leading to iron transfer to the bacterium cell interior. In order to elucidate the molecular events leading from the formation of the initial IsdB:Hb complex to heme extraction, we have performed a time-resolved X-ray solution scattering (TR-XSS) investigation combined with a rapid mixing triggering approach. We succeeded in defining the stoichiometry of IsdB:Hb binding and in describing the kinetics of the subsequent structural changes. The presented approach is potentially applicable to unveil the complex kinetic pathways generated by protein-protein interaction in different biological systems.
Molecular mechanism of bacteriophage tail contraction-structure of an S-layer-penetrating bacteriophage
Authors: Wilson Jason S. , Fortier Louis-Charles , Fagan Robert P. , Bullough Per A. 
Published: August 2023

AbstractViruses that infect bacteria (bacteriophages or phages) attach to the host cell envelope, inject their genetic material into the host cytosol and either persist as prophage or hijack the host machinery to produce progeny virions. Attachment is mediated through phage receptor binding proteins that are specific for different host cell surface molecules. A subset of phage, the myoviruses, possess contractile tails, the outer sheath of which contracts upon receptor binding, driving an inner tail tube through the cell envelope and delivering the phage genome into the host cytosol. The molecular details of phage tail contraction and mode of cell envelope penetration have remained poorly understood and were completely unknown for any phage infecting bacteria enveloped by a proteinaceous S-layer. Here we reveal the extended and contracted atomic structures of an intact contractile-tail phage that binds to and penetrates the protective S-layer of the Gram positive human pathogenClostridioides difficile. Surprisingly, we find no evidence of the intrinsic enzymatic domains that other phages exploit in cell wall penetration, suggesting that sufficient energy is released upon tail contraction to penetrate the S-layer and the thick cell wall without enzymatic activity. However, it is also notable that the tail sheath subunits move less than those studied in related contractile injection systems such as the model phage T4. Instead, the unusually long tail length and flexibility upon contraction likely contribute towards the required free energy release for envelope penetration. Our results show that the principles of phage contraction and infection as determined in the model system of T4 are not universal. We anticipate that our structures will form a strong foundation to engineerC. difficilephages as therapeutics, and highlight important adaptations made in order to infect S-layer containing pathogens.
Air pollution modifies key colonization factors of the beneficial bee gut symbiontSnodgrassella alviand disrupts the bumblebee (Bombus terrestris) gut microbiome
Authors: Sampson Hannah R. , Allcock Natalie , Mallon Eamonn B. , Ketley Julian M. , Morrissey Julie A. 
Published: August 2023

SummaryAir pollution is the world’s largest environmental health risk. Particulate pollutants, a major component of air pollution, are detrimental to human health and a significant risk to wildlife and ecosystems globally. Black carbon, a by-product of fossil fuel and biomass burning, is a key constituent of air pollution with levels continuing to increase worldwide. Here we describe the effects of black carbon on the beneficial gut microbiome of an important global insect pollinator, the buff-tailed bumblebee (Bombus terrestris). Our data shows that exposure to black carbon particulates alters the biofilm structure, gene expression and initial adhesion of beneficial bee gut coloniser,Snodgrassella alvi in vitro.Additionally, our results show that black carbon disrupts adultBombus terrestrisgut microbiome composition, a vital component to bee health. Exposure to black carbon increased bees’ viable gut bacteria and significantly altered the abundance of beneficial core bacteriaGilliamellaandBombilactobacillusin the microbiome. These findings demonstrate that exposure to black carbon air pollution has direct, measurable effects on bees’ beneficial gut commensal bacteria and microbiome. Together these data highlight that particulate pollutants are an underexplored risk for the health of insect pollinators.
YfmR is a translation factor that prevents ribosome stalling and cell death in the absence of EF-P
Authors: Hong Hye-Rim , Prince Cassidy R. , Tetreault Daniel D. , Wu Letian , Feaga Heather A. 
Published: August 2023

AbstractProtein synthesis is performed by the ribosome and a host of highly conserved elongation factors. Elongation factor P (EF-P) prevents ribosome stalling at difficult-to-translate sequences, particularly polyproline tracts. In bacteria, phenotypes associated withefpdeletion range from modest to lethal, suggesting that some species encode an additional translation factor that has similar function to EF-P. Here we identify YfmR as a translation factor that is essential in the absence of EF-P inB. subtilis. YfmR is an ABCF ATPase that is closely related to both Uup and EttA, ABCFs that bind the ribosomal E-site and are conserved in more than 50% of bacterial genomes. We show that YfmR associates with actively translating ribosomes and that depleting YfmR from Δefpcells causes severe ribosome stalling at a polyproline tractin vivo. YfmR depletion from Δefpcells was lethal, and caused reduced levels of actively translating ribosomes. Our results therefore identify YfmR as an important translation factor that is essential inB. subtilisin the absence of EF-P.SignificanceTranslation is one of the most ancient and energetically demanding processes that occurs in the cell. Ribosomes constitute more than 60% of cellular mass in actively growing cells, and ribosomes are a major target of antimicrobials and chemotherapeutics. Here, we identify YfmR as a translation factor that is essential in the absence of EF-P. YfmR is a member of the ABCF family of ATPases whose role in translation is only beginning to be understood. Given the broad distribution of ABCFs from bacteria to fungi, we expect our results to have implications for understanding translation elongation in diverse organisms.
Nonstructural protein 1 of swine arterivirus PRRSV downregulates promyelocytic leukemia nuclear bodies and promotes viral replication
Authors: Su Chia-Ming , Han Mingyuan , Everett Roger , Yoo Dongwan 
Published: August 2023

ABSTRACTPorcine reproductive and respiratory syndrome virus (PRRSV) inhibits the type I interferon (IFN) production and signaling pathways during infection, and the nonstructural protein 1 (nsp1) has been identified as a potent IFN antagonist. Promyelocytic leukemia (PML) protein is a major scaffold protein organizing the PML nuclear bodies (NBs) of the cell and plays a diverse role in maintaining the cellular homeostasis including antiviral response among many other processes. The present study reveals a significant reduction of PML NBs in cells during infection of PRRSV, implicating the negative regulation of PML gene expression by PRRSV. Subsequently, the nsp1β protein was identified as the viral regulator for PML expression. The overexpression of PML isoforms restricted viral replication, while the gene silencing of endogenous PML promoted viral replication. The downregulation of PML expression by PRRSV was post-translational via the ubiquitination-proteasome pathway. Of six isoforms, PML-II and PML-IV exhibited the most potent suppressive activity against viral replication. PRRSV nsp1β bound to PML directly, and this interaction was mediated through the small ubiquitin-like modifier (SUMO)-interacting motifs (SIMs) on nsp1β. Further studies revealed that double mutations in SIM1 and SIM4 abolished the binding of nsp1β to PML and prevented the PML degradation. The PML downregulation by nsp1 was common in other arteriviruses including equine arteritis virus, murine lactate dehydrogenase elevating virus, and simian hemorrhagic fever virus. Our study unveils the evolutionary conservation of the viral immune evasion strategy employed by arteriviruses, which promotes their replication by targeting PML for downregulation.IMPORTANCE OF THE STUDYPorcine reproductive and respiratory syndrome (PRRS) is an economically significant disease in the swine industry worldwide. One of the immunological hallmarks in virus-infected animals is the suppression of type I interferon response during an early-stage infection, leading to the consequence of adaptive immunity and viral persistence. In the present study, we report that the nsp1-beta protein of PRRS virus degrades the promyelocytic leukemia (PML) protein and downregulates PML nuclear body (NB) formation. The PML downregulation by PRRS virus results in enhanced viral replication. The PML downregulation by nsp1 is common in other arteriviruses, unveiling the basic understanding of cell-virus interactions and immune evasion strategies for arteriviruses.
OsNAC2 maintains the homeostasis of immune responses to bacterial blight through the OsEREBP1 in rice
Authors: Zhong Qun , Yu Jiangtao , Wu Yiding , Yao Xuefeng , Meng Xiangzong , Ming Feng 
Published: August 2023

AbstractRice bacterial blight, caused byXanthomonas oryzaepv.Oryzae(Xoo), threatens plant growth and yield. However, the molecular mechanisms underlying rice immunity againstXooremain elusive. Here, we demonstrated down-regulation of a NAC (NAM-ATAF-CUC) transcription factorOsNAC2enhanced resistance to bacterial blight disease in rice. Consistently, salicylic acid (SA) biosynthesis was greatly attenuated when OsNAC2 overexpressed. Furthermore, study demonstrated that OsNAC2 can down-regulated the expression of SA synthesis genes, thus mediating SA signal transmission. Simultaneously, OsNAC2 interacted with OsEREBP1 (AP2/ERF family) in the nucleus, may be bloking the inhibition of OsNAC2 toOsICS1,OsPAL3and so on. Furthermore, OsEREBP1 interacted with OsXb22a in the cytoplasm to exert its positive regulatory effects to bacterial blight. However,OsNAC2-overexpression kept OsEREBP1 in the nucleus and be rapidly degraded in pathogen infection, which adversely affects the interaction of OsEREBP1-OsXb22a. Our results determined that OsNAC2 inhibits the SA signaling and stably interacts with OsEREBP1 to maintain disease resistance. This OsNAC2-OsEREBP1-based homeostatic mechanism provides new insights into rice disease resistance, and it may be useful for improving the disease resistance of important crops through breeding.
A Bright, Photostable Dye that Enables Multicolor, Time Lapse, and Super-Resolution Imaging of Acidic Organelles
Authors: Lesiak Lauren , Dadina Neville , Zheng Shuai , Schelvis Marianne , Schepartz Alanna 
Published: August 2023

ABSTRACTLysosomes have long been known for their acidic lumen and efficient degradation of cellular byproducts. In recent years it has become clear that their function is far more sophisticated, involving multiple cell signaling pathways and interactions with other organelles. Unfortunately, their acidic interior, fast dynamics, and small size makes lysosomes difficult to image with fluorescence microscopy. Here we report a far-red small molecule, HMSiR680-Me, that fluoresces only under acidic conditions, causing selective labeling of acidic organelles in live cells. HMSiR680-Me can be used alongside other far-red dyes in multicolor imaging experiments and is superior to existing lysosome probes in terms of photostability and maintaining cell health and lyso-some motility. We demonstrate that HMSiR680-Me is compatible with overnight time lapse experiments, as well as time lapse super-resolution microscopy with a fast frame rate for at least 1000 frames. HMSiR680-Me can also be used alongside silicon rhodamine dyes in a multiplexed super-resolution microscopy experiment to visualize interactions between the inner mitochondrial membrane and lysosomes with only a single excitation laser and simultaneous depletion. We envision this dye permitting more detailed study of the role of lysosomes in dynamic cellular processes and disease.
Molecular layer disinhibition unlocks climbing-fiber-instructed motor learning in the cerebellum
Authors: Zhang Ke , Yang Zhen , Gaffield Michael A. , Gross Garrett G. , Arnold Don B. , Christie Jason M. 
Published: August 2023

AbstractClimbing fibers supervise cerebellar learning by providing signals to Purkinje cells (PCs) that instruct adaptive changes to mistakenly performed movements. Yet, climbing fibers are regularly active, even during well performed movements, suggesting that a mechanism dynamically regulates the ability of climbing fibers to induce corrective plasticity in response to motor errors. We found that molecular layer interneurons (MLIs), whose inhibition of PCs powerfully opposes climbing-fiber-mediated excitation, serve this function. Optogenetically suppressing the activity of floccular MLIs in mice during the vestibulo-ocular reflex (VOR) induces a learned increase in gain despite the absence of performance errors. Suppressing MLIs when the VOR is mistakenly underperformed reveled that their inhibitory output is necessary to orchestrate gain-increase learning by conditionally permitting climbing fibers to instruct plasticity induction during ipsiversive head turns. Ablation of an MLI circuit for PC disinhibition prevents gain-increase learning during VOR performance errors which was rescued by re-imposing PC disinhibition through MLI activity suppression. Our findings point to a decisive role for MLIs in gating climbing-fiber-mediated learning through their context-dependent inhibition of PCs.
Ablation of carotid body activity reverses diabesity by improving white and brown adipose tissue sympathetic innervation and metabolism
Authors: Melo Bernardete F. , Sacramento Joana F. , Lavergne Julien , Martins Fátima O. , Rosendo-Silva Daniela , Panzolini Clara , Melo Gonçalo M. , Prego Cláudia S. , Falvey Aidan , Olea Elena , Matafome Paulo , Rocher Asuncion , Prieto-Lloret Jesus , Correia Miguel C. , Blancou Phillipe , Conde Silvia V. 
Published: August 2023

SummaryFinding novel pathological mechanisms that lead to innovative strategies to treat obesity and its associated illness are critically needed. The carotid bodies (CB) are metabolic sensors whose dysfunction contributes to insulin resistance and glucose intolerance development. Herein, we find that the ablation of CB activity, through resection of carotid sinus nerve (CSN) promotes weight loss and restores metabolic function in high fat (HF) rodents, by increasing WAT basal metabolism and by restoring WAT and BAT sympathetic activation. Moreover, we found that CSN resection rescues adipose tissue sympathetic/catecholamine resistance present in obesity states. Additionally, we found that the CB signals integrated in the paraventricular nucleus of the hypothalamus are decreased in obesity states and that CSN denervation in HF animals restore neuronal activity in this region. By inducing energy expenditure via the increase in WAT and BAT metabolism, CB modulation might be used as a therapeutic target for obesity and dysmetabolism.HighlightsAblation of carotid body (CB) activity promotes weight lossCarotid body modulates white and brown adipose tissue metabolismAblation of CB activity rescues adipose tissue sympathetic resistance in obesityCarotid sinus nerve resection restored the altered neuronal activity induced by obesity in the paraventricular nucleus of the hypothalamus