bluuebunny/crossref_metadata_2025_split huggingface dataset:
Reinforcement Learning informs optimal treatment strategies to limit antibiotic resistance
Authors: Weaver Davis T. , King Eshan S. , Maltas Jeff , Scott Jacob G. 
Published: January 2023

ABSTRACTAntimicrobial resistance was estimated to be associated with 4.95 million deaths worldwide in 2019. It is possible to frame the antimicrobial resistance problem as a feedback-control problem. If we could optimize this feedback-control problem and translate our findings to the clinic, we could slow, prevent or reverse the development of high-level drug resistance. Prior work on this topic has relied on systems where the exact dynamics and parameters were knowna priori. In this study, we extend this work using a reinforcement learning (RL) approach capable of learning effective drug cycling policies in a system defined by empirically measured fitness landscapes. Crucially, we show that is possible to learn effective drug cycling policies despite the problems of noisy, limited, or delayed measurement. Given access to a panel of 15β-lactam antibiotics with which to treat the simulatedE. colipopulation, we demonstrate that RL agents outperform two naive treatment paradigms at minimizing the population fitness over time. We also show that RL agents approach the performance of the optimal drug cycling policy. Even when stochastic noise is introduced to the measurements of population fitness, we show that RL agents are capable of maintaining evolving populations at lower growth rates compared to controls. We further tested our approach in arbitrary fitness landscapes of up to 1024 genotypes. We show that minimization of population fitness using drug cycles is not limited by increasing genome size. Our work represents a proof-of-concept for using AI to control complex evolutionary processes.
A paternal signal induces endosperm proliferation upon fertilization in Arabidopsis
Authors: Simonini Sara , Bencivenga Stefano , Grossniklaus Ueli 
Published: January 2023

AbstractIn multicellular organisms, sexual reproduction relies on the formation of highly specialized, differentiated cells, the gametes. At maturity, male and female gametes are quiescent, awaiting fertilization, with their cell cycle being arrested at a precise stage. Failure to establish quiescence leads to unwanted proliferation, abortion of the offspring, and a waste of resources. Upon fertilization, the cell cycle resumes, allowing the newly formed zygote to divide rapidly. Successful development requires that male and female gametes are in the same phase of the cell cycle. The molecular mechanisms that enforce quiescence and reinstate cell division only after fertilization occurs are poorly understood. Here, we describe a sperm-derived signal that induces proliferation of theArabidopsiscentral cell precisely upon fertilization. We show that the mature central cell is arrested in S phase, caused by the activity of the conserved RETINOBLASTOMA RELATED1 (RBR1) protein. Paternal delivery of the core cell cycle component CYCD7;1 triggers RBR1 degradation, thereby stimulating S phase progression. Absence of CYCD7;1 delays RBR1 depletion, S phase reactivation, and central cell division, whereas its constitutive expression triggers proliferation of unfertilized central cells. In summary, we show that CYCD7;1 is a paternal signal that informs the central cell that fertilization occurred, thus unlocking quiescence and ensuring that cell division initiates just at the right time to ensure functional endosperm formation.
BRCA1/BARD1 ubiquitinates PCNA in unperturbed conditions to promote replication fork stability and continuous DNA synthesis
Authors: Salas-Lloret Daniel , García-Rodríguez Néstor , Giebel Lisanne , de Ru Arnoud , van Veelen Peter A. , Huertas Pablo , Vertegaal Alfred C.O. , González-Prieto Román 
Published: January 2023

SUMMARYDeficiencies in the BRCA1 tumor suppressor gene are the main cause of hereditary breast and ovarian cancer. BRCA1 is involved in the Homologous Recombination DNA repair pathway, and, together with BARD1, forms a heterodimer with ubiquitin E3 activity. The relevance of the BRCA1/BARD1 ubiquitin E3 activity for tumor suppression and DNA repair remains controversial and most efforts aimed to identify BRCA1/BARD1 ubiquitination substrates rely on indirect evidence. Here, we observed that the BRCA1/BARD1 ubiquitin E3 activity was not required for Homologous Recombination or resistance to Olaparib. Using TULIP2 methodology, which enables the direct identification of E3-specific ubiquitination substrates, we identified substrates for BRCA1/BARD1. We found that PCNA is ubiquitinated by BRCA1/BARD1 in unperturbed conditions independently of RAD18. PCNA ubiquitination by BRCA1/BARD1 avoids the formation of ssDNA gaps during DNA replication and promotes replication fork stability epistatically to BRCA1 S114 phosphorylation, addressing the controversy about the function of BRCA1/BARD1 E3 activity in Homologous Recombination.
A Multiplexed SEC-MS Approach to Systematically Study the Interplay Between Protein Assembly-States and Phosphorylation Events
Authors: Doron-Mandel Ella , Bokor Benjamin J. , Ma Yanzhe , Street Lena A. , Tang Lauren C. , Abdou Ahmed A. , Shah Neel H. , Rosenberger George A. , Jovanovic Marko 
Published: January 2023

AbstractA protein’s molecular interactions and post-translational modifications (PTMs), such as phosphorylation, can be co-dependent and reciprocally co-regulate each other. Although this interplay is central for many biological processes, a systematic method to simultaneously study assembly-states and PTMs from the same sample is critically missing. Here, we introduce SEC-MX (Size Exclusion Chromatography fractions MultipleXed), a global quantitative method combining Size Exclusion Chromatography and PTM-enrichment for simultaneous characterization of PTMs and assembly-states. SEC-MX enhances throughput, allows phosphopeptide enrichment, and facilitates quantitative differential comparisons between biological conditions. Applying SEC-MX to HEK293 and HCT116 cells, we generated a proof-of-concept dataset mapping thousands of phosphopeptides and their assembly-states. Our analysis revealed intricate relationships between phosphorylation events and assembly-states and generated testable hypotheses for follow-up studies. Overall, we establish SEC-MX as a valuable tool for exploring protein functions and regulation beyond abundance changes.
Active Learning of the Conformational Ensemble of Proteins using Maximum Entropy VAMPNets
Authors: Kleiman Diego E. , Shukla Diwakar 
Published: January 2023

AbstractRapid computational exploration of the free energy landscape of biological molecules remains an active area of research due to the difficulty of sampling rare state transitions in Molecular Dynamics (MD) simulations. In recent years, an increasing number of studies have exploited Machine Learning (ML) models to enhance and analyze MD simulations. Notably, unsupervised models that extract kinetic information from a set of parallel trajectories have been proposed, including the variational approach for Markov processes (VAMP), VAMPNets, and time-lagged variational autoencoders (TVAE). In this work, we propose a combination of adaptive sampling with active learning of kinetic models to accelerate the discovery of the conformational landscape of biomolecules. In particular, we introduce and compare several techniques that combine kinetic models with two adaptive sampling regimes (least counts and multi-agent reinforcement learning-based adaptive sampling) to enhance the exploration of conformational ensembles without introducing biasing forces. Moreover, inspired by the active learning approach of uncertainty-based sampling, we also present MaxEnt VAMPNet. This technique consists of restarting simulations from the microstates that maximize the Shannon entropy of a VAMPNet trained to perform soft discretization of metastable states. By running simulations on two test systems, the WLALL pentapeptide and the villin headpiece subdomain, we empirically demonstrate that MaxEnt VAMPNet results in faster exploration of conformational landscapes compared to the baseline and other proposed methods.
An AlphaFold2 map of the 53BP1 pathway identifies a direct SHLD3-RIF1 interaction critical for DNA repair activity
Authors: Sifri Chérine , Hoeg Lisa , Durocher Daniel , Setiaputra Dheva 
Published: January 2023

Abstract53BP1 is a chromatin-binding DNA repair protein that promotes DNA double-strand break repair through recruitment of downstream effectors including RIF1, shieldin, and CST. The structural basis of the protein-protein interactions within the 53BP1-RIF1-shieldin-CST pathway that are essential for its DNA repair activity are largely unknown. Here we used AlphaFold2-Multimer (AF2) to predict all possible pairwise combinations of proteins within this pathway and provide structural models of seven previously characterized interactions. This analysis also predicted an entirely novel binding interface between the HEAT-repeat domain of RIF1 and the eIF4E-like domain of SHLD3. Extensive interrogation of this interface through both in vitro pulldown analysis and cellular assays supports the AF2-predicted model and demonstrates that RIF1-SHLD3 binding is essential for shieldin recruitment to sites of DNA damage, and for its role in antibody class switch recombination. Direct physical interaction between RIF1 and SHLD3 is therefore essential for 53BP1-RIF1-shieldin-CST pathway activity.
pTDP-43 levels correlate with cell type specific molecular alterations in the prefrontal cortex ofC9orf72ALS/FTD patients
Authors: Wang Hsiao-Lin V. , Xiang Jian-Feng , Yuan Chenyang , Veire Austin M. , Gendron Tania F. , Murray Melissa E. , Tansey Malú G. , Hu Jian , Gearing Marla , Glass Jonathan D. , Jin Peng , Corces Victor G. , McEachin Zachary T. 
Published: January 2023

AbstractRepeat expansions in theC9orf72gene are the most common genetic cause of amyotrophic lateral sclerosis and familial frontotemporal dementia (ALS/FTD). To identify molecular defects that take place in the dorsolateral frontal cortex of patients withC9orf72ALS/FTD, we compared healthy controls withC9orf72ALS/FTD donor samples staged based on the levels of cortical phosphorylated TAR DNA binding protein (pTDP-43), a neuropathological hallmark of disease progression. We identified distinct molecular changes in different cell types that take place during FTD development. Loss of neurosurveillance microglia and activation of the complement cascade take place early, when pTDP-43 aggregates are absent or very low, and become more pronounced in late stages, suggesting an initial involvement of microglia in disease progression. Reduction of layer 2-3 cortical projection neurons with high expression of CUX2/LAMP5 also occurs early, and the reduction becomes more pronounced as pTDP-43 accumulates. Several unique features were observed only in samples with high levels of pTDP-43, including global alteration of chromatin accessibility in oligodendrocytes, microglia, and astrocytes; higher ratios of premature oligodendrocytes; increased levels of the noncoding RNA NEAT1 in astrocytes and neurons, and higher amount of phosphorylated ribosomal protein S6. Our findings reveal previously unknown progressive functional changes in major cell types found in the frontal cortex ofC9orf72ALS/FTD patients that shed light on the mechanisms underlying the pathology of this disease.
Virulence and genomic diversity among clinical isolates of ST1 (BI/NAP1/027)Clostridioides difficile
Authors: Dong Qiwen , Lin Huaiying , Allen Marie-Maude , Garneau Julian R. , Sia Jonathan K. , Smith Rita C. , Haro Fidel , McMillen Tracy , Pope Rosemary L. , Metcalfe Carolyn , Burgo Victoria , Woodson Che , Dylla Nicholas , Kohout Claire , Sundararajan Anitha , Snitkin Evan S , Young Vincent B. , Fortier Louis-Charles , Kamboj Mini , Pamer Eric G. 
Published: January 2023

AbstractClostridioides difficile (C. difficile), a leading cause of nosocomial infection, produces toxins that damage the colonic epithelium and results in colitis that varies from mild to fulminant. Variation in disease severity is poorly understood and has been attributed to host factors (age, immune competence and intestinal microbiome composition) and/or virulence differences betweenC. difficilestrains, with some, such as the epidemic BI/NAP1/027 (MLST1) strain, being associated with greater virulence. We tested 23 MLST1(ST1)C. difficileclinical isolates for virulence in antibiotic-treated C57BL/6 mice. All isolates encoded a complete Tcd pathogenicity locus and achieved similar colonization densities in mice. Disease severity varied, however, with 5 isolates causing lethal infections, 16 isolates causing a range of moderate infections and 2 isolates resulting in no detectable disease. The avirulent ST1 isolates did not cause disease in highly susceptible Myd88-/-or germ-free mice. Genomic analysis of the avirulent isolates revealed a 69 base-pair deletion in the N-terminus of thecdtRgene, which encodes a response regulator for binary toxin (CDT) expression. Genetic deletion of the 69 base-paircdtRsequence in the highly virulent ST1 R20291C. difficilestrain rendered it avirulent and reduced toxin gene transcription in cecal contents. Our study demonstrates that a natural deletion withincdtRattenuates virulence in the epidemic ST1C. difficilestrain without reducing colonization and persistence in the gut. Distinguishing strains on the basis ofcdtRmay enhance the specificity of diagnostic tests forC. difficilecolitis.
Spliceosomal mutations decouple 3′ splice site fidelity from cellular fitness
Authors: Roy Kevin R. , Gabunilas Jason , Neutel Dean , Ai Michelle , Samson Joyce , Lyu Guochang , Chanfreau Guillaume F. 
Published: January 2023

AbstractThe fidelity of splice site selection is thought to be critical for proper gene expression and cellular fitness. In particular, proper recognition of 3′-splice site (3′SS) sequences by the spliceosome is a daunting task considering the low complexity of the 3′SS consensus sequence YAG. Here we show that inactivating the near-essential splicing factor Prp18p results in a global activation of alternative 3′SS, many of which harbor sequences that highly diverge from the YAG consensus, including some highly unusual non-AG 3′SS. We show that the role of Prp18p in 3′SS fidelity is promoted by physical interactions with the essential splicing factors Slu7p and Prp8p and synergized by the proofreading activity of the Prp22p helicase. Strikingly, structure-guided point mutations that disrupt Prp18p-Slu7p and Prp18p-Prp8p interactions mimic the loss of 3′SS fidelity without any impact on cellular growth, suggesting that accumulation of incorrectly spliced transcripts does not have a major deleterious effect on cellular viability. These results show that spliceosomes exhibit remarkably relaxed fidelity in the absence of Prp18p, and that new 3′SS sampling can be achieved genome-wide without a major negative impact on cellular fitness, a feature that could be used during evolution to explore new productive alternative splice sites.
A Single Laccase Acts as a Key Component of Environmental Sensing in a Broad Host Range Fungal Pathogen
Authors: Westrick Nathaniel M. , Dominguez Eddie G. , Hull Christina M. , Smith Damon L. , Kabbage Mehdi 
Published: January 2023

AbstractSecreted laccases are important enzymes on an ecological scale for their role in mediating plant-fungal interactions, but their function in fungal pathogenesis has yet to be elucidated. Ascomycete laccases have been primarily associated with cell wall melanin deposition, and laccase mutants in ascomycete species often demonstrate reduced pigmentation. In this study, a putatively secreted laccase,Sslac2, was characterized from the broad host-range plant pathogenSclerotinia sclerotiorum, which is largely unpigmented and is not dependent on melanogenesis for plant infection. Of the seven putative laccases in theS. sclerotiorumgenome,Sslac2was the only one found to be highly upregulated during pathogenesis of soybeans and was additionally found to be induced during growth on solid surfaces. Gene knockouts ofSslac2demonstrate wide ranging developmental defects, including abolished sclerotial formation, and are functionally non-pathogenic on unwounded tissue. While these mutants demonstrated a clear radial growth defect, enhanced growth was observed in liquid culture, likely due to altered hydrophobicity and thigmotropic responsiveness.Sslac2mutants were also unable to respond to a host environment, and accordingly unable to differentiate penetration structures, respond appropriately to chemical stress, or produce the key virulence factor oxalic acid. Transmission and scanning electron microscopy of WT and mutant strains show apparent differences in extracellular matrix structure that may explain the inability of the mutant to respond to environmental cues. TargetingSslac2using host-induced gene silencing significantly improved resistance toS. sclerotiorum, suggesting that fungal laccases could be a valuable target of disease control. Collectively, we identified a laccase critical to the development and virulence of the broad host-range pathogenS. sclerotiorumand propose a potentially novel role for fungal laccases in modulating environmental sensing.
Microbiome derived acidity protects against microbial invasion inDrosophila
Authors: Barron Alexander J. , Lesperance Danielle N. A. , Doucette Jeremy , Calle Sthefany , Broderick Nichole A. 
Published: January 2023

AbstractMicrobial invasions underlie host-microbe interactions that result in microbial pathogenesis and probiotic colonization. While these processes are of broad interest, there are still gaps in our understanding of the barriers to entry and how some microbes overcome them. In this study, we explore the effects of the microbiome on invasions of foreign microbes inDrosophila melanogaster. We demonstrate that gut microbesLactiplantibacillus plantarumandAcetobacter tropicalisimprove survival during invasion of a lethal gut pathogen and lead to a reduction in microbial burden. Using a novel multi-organism interactions assay, we report thatL. plantaruminhibits the growth of three invasive Gram-negative bacteria, whileA. tropicalisprevents this inhibition. A series ofin vitroandin vivoexperiments revealed that inhibition byL. plantarumis linked to its ability to acidify both internal and external environments, including culture media, fly food, and the gut itself, whileA. tropicalisdiminishes the inhibition by quenching acids. We propose that acid produced by the microbiome serves as an important gatekeeper to microbial invasions, as only microbes capable of tolerating acidic environments can colonize the host. The methods described herein will add to the growing breadth of tools to study microbe-microbe interactions in broad contexts.
Bidirectional multiciliated cell extrusion is controlled by Notch driven basal extrusion and Piezo 1 driven apical extrusion
Authors: Ventrella Rosa , Kim Sun K. , Sheridan Jennifer , Grata Aline , Bresteau Enzo , Hassan Osama , Suva Eve E. , Walentek Peter , Mitchell Brian 
Published: January 2023

AbstractXenopusembryos are covered with a complex epithelium containing numerous multiciliated cells (MCCs). During late stage development there is a dramatic remodeling of the epithelium that involves the complete loss of MCCs. Cell extrusion is a well-characterized process for driving cell loss while maintaining epithelial barrier function. Normal cell extrusion is typically unidirectional whereas bidirectional extrusion is often associated with disease (e.g. cancer). We describe two distinct mechanisms for MCC extrusion, a basal extrusion driven by Notch signaling and an apical extrusion driven by Piezo1. Early in the process there is a strong bias towards basal extrusion, but as development continues there is a shift towards apical extrusion. Importantly, receptivity to the Notch signal is age-dependent and governed by the maintenance of the MCC transcriptional program such that extension of this program is protective against cell loss. In contrast, later apical extrusion is regulated by Piezo 1 such that premature activation of Piezo 1 leads to early extrusion while blocking Piezo 1 leads to MCC maintenance. Distinct mechansms for MCC loss underlie the importance of their removal during epithelial remodeling.Summay StatementCell extrusion typically occurs unidirectionally. We have identified a single population of multiciliated cells that extrudes bidirectionally: Notch-driven basal extrusion and Piezo 1-mediated apical extrusion.
Periosteal skeletal stem cells can migrate into the bone marrow and support hematopoiesis after injury
Authors: Marchand Tony , Akinnola Kemi E. , Takeishi Shoichiro , Maryanovich Maria , Pinho Sandra , Saint-Vanne Julien , Birbrair Alexander , Lamy Thierry , Tarte Karin , Frenette Paul S. , Gritsman Kira 
Published: January 2023

SUMMARYSkeletal stem cells have been isolated from various tissues, including periosteum and bone marrow, where they exhibit key functions in bone biology and hematopoiesis, respectively. The role of periosteal skeletal stem cells in bone regeneration and healing has been extensively studied, but their ability to contribute to the bone marrow stroma is still under debate. In the present study, we characterized a whole bone transplantation model that mimics the initial bone marrow necrosis and fatty infiltration seen after injury. Using this model and a lineage tracing approach, we observed the migration of periosteal skeletal stem cells into the bone marrow after transplantation. Once in the bone marrow, periosteal skeletal stem cells are phenotypically and functionally reprogrammed into bone marrow mesenchymal stem cells that express high levels of hematopoietic stem cell niche factors such as Cxcl12 and Kitl. In addition, usingex vivoandin vivoapproaches, we found that periosteal skeletal stem cells are more resistant to acute stress than bone marrow mesenchymal stem cells. These results highlight the plasticity of periosteal skeletal stem cells and their potential role in bone marrow regeneration after bone marrow injury.
Piconewton forces mediate GAIN domain dissociation of the latrophilin-3 adhesion GPCR
Authors: Zhong Brian L. , Lee Christina E. , Vachharajani Vipul T. , Südhof Thomas C. , Dunn Alexander R. 
Published: January 2023

AbstractLatrophilins are adhesion G-protein coupled receptors (aGPCRs) that control excitatory synapse formation. aGPCRs, including latrophilins, are autoproteolytically cleaved at their GPCR-Autoproteolysis Inducing (GAIN) domain, but the two resulting fragments remain associated on the cell surface. It is thought that force-mediated dissociation of the fragments exposes a peptide that activates G-protein signaling of aGPCRs, but whether GAIN domain dissociation can occur on biologically relevant timescales and at physiological forces is unknown. Here, we show using magnetic tweezers that physiological forces dramatically accelerate the dissociation of the latrophilin-3 GAIN domain. Forces in the 1-10 pN range were sufficient to dissociate the GAIN domain on a seconds-to-minutes timescale, and the GAIN domain fragments reversibly reassociated after dissociation. Thus, mechanical force may be a key driver of latrophilin signaling during synapse formation, suggesting a physiological mechanism by which aGPCRs may mediate mechanically-induced signal transduction.