bluuebunny/crossref_metadata_2025_split huggingface dataset:
Tracking of antibiotic resistance transfer and rapid plasmid evolution in a hospital setting by Nanopore sequencing
Authors: Peter Silke , Bosio Mattia , Gross Caspar , Bezdan Daniela , Gutierrez Javier , Oberhettinger Philipp , Liese Jan , Vogel Wichard , Dörfel Daniela , Berger Lennard , Marschal Matthias , Willmann Matthias , Gut Ivo , Gut Marta , Autenrieth Ingo , Ossowski Stephan 
Published: May 2019

AbstractBackgroundInfection of patients with multidrug-resistant (MDR) bacteria often leave very limited or no treatment options. The transfer of antimicrobial resistance genes (ARG) carrying plasmids between bacterial species by horizontal gene transfer represents an important mode of expansion of ARGs. Here, we evaluated the application of Nanopore sequencing technology in a hospital setting for monitoring the transfer and rapid evolution of antibiotic resistance plasmids within and across multiple species.ResultsIn 2009 we experienced an outbreak with an extensively multidrug resistant P. aeruginosa harboring the carbapenemase enzyme blaIMP-8, and in 2012 the first Citrobacter freundii and Citrobacter werkmanii harboring the same enzyme were detected. Using Nanopore and Illumina sequencing we conducted a comparative analysis of all blaIMP-8 bacteria isolated in our hospital over a 6-year period (n = 54). We developed the computational platforms pathoLogic and plasmIDent for Nanopore-based characterization of clinical isolates and monitoring of ARG transfer, comprising de-novo assembly of genomes and plasmids, polishing, QC, plasmid circularization, ARG annotation, comparative genome analysis of multiple isolates and visualization of results. Using plasmIDent we identified a 40 kb plasmid carrying blaIMP-8 in P. aeruginosa and C. freundii, verifying that plasmid transfer had occurred. Within C. freundii the plasmid underwent further evolution and plasmid fusion, resulting in a 164 kb mega-plasmid, which was transferred to C. werkmanii. Moreover, multiple rearrangements of the multidrug resistance gene cassette were detected in P. aeruginosa, including deletions and translocations of complete ARGs.ConclusionPlasmid transfer, plasmid fusion and rearrangement of the multidrug resistance gene cassette mediated the rapid evolution of opportunistic pathogens in our hospital. We demonstrated the feasibility of tracking plasmid evolution dynamics and ARG transfer in clinical settings in a timely manner. The approach will allow for successful countermeasures to contain not only clonal, but also plasmid mediated outbreaks.
Development of the first low nanomolar Liver Receptor Homolog-1 agonist through structure-guided design
Authors: Mays Suzanne G. , Flynn Autumn R. , Cornelison Jeffery L. , Okafor C. Denise , Wang Hongtao , Wang Guohui , Huang Xiangsheng , Donaldson Heather N. , Millings Elizabeth J. , Polavarapu Rohini , Moore David D. , Calvert John W. , Jui Nathan T. , Ortlund Eric A. 
Published: May 2019

ABSTRACTAs a key regulator of metabolism and inflammation, the orphan nuclear hormone receptor, Liver Receptor Homolog-1 (LRH-1), has potential as a therapeutic target for diabetes, nonalcoholic fatty liver disease, and inflammatory bowel diseases. Discovery of LRH-1 modulators has been difficult, in part due to the tendency for synthetic compounds to bind unpredictably within the lipophilic binding pocket. Using a structure-guided approach, we exploited a newly-discovered polar interaction to lock agonists in a consistent orientation. This enabled the discovery of the first low nanomolar LRH-1 agonist, one hundred times more potent than the best previous modulator. We elucidate a novel mechanism of action that relies upon specific polar interactions deep in the LRH-1 binding pocket. In an organoid model of inflammatory bowel disease, the new agonist increases expression of LRH-1-conrolled steroidogenic genes and promotes anti-inflammatory gene expression changes. These studies constitute major progress in developing LRH-1 modulators with potential clinical utility.
Ultrastructural details of mammalian chromosome architecture
Authors: Krietenstein Nils , Abraham Sameer , Venev Sergey V. , Abdennur Nezar , Gibcus Johan , Hsieh Tsung-Han S. , Parsi Krishna Mohan , Yang Liyan , Maehr René , Mirny Leonid A. , Dekker Job , Rando Oliver J. 
Published: May 2019

ABSTRACTOver the past decade, 3C-related methods, complemented by increasingly detailed microscopic views of the nucleus, have provided unprecedented insights into chromosome folding in vivo. Here, to overcome the resolution limits inherent to the majority of genome-wide chromosome architecture mapping studies, we extend a recently-developed Hi-C variant, Micro-C, to map chromosome architecture at nucleosome resolution in human embryonic stem cells and fibroblasts. Micro-C maps robustly capture well-described features of mammalian chromosome folding including A/B compartment organization, topologically associating domains (TADs), and cis interaction peaks anchored at CTCF binding sites, while also providing a detailed 1-dimensional map of nucleosome positioning and phasing genome-wide. Compared to high-resolution in situ Hi-C, Micro-C exhibits substantially improved signal-to-noise with an order of magnitude greater dynamic range, enabling not only localization of domain boundaries with single-nucleosome accuracy, but also resolving more than 20,000 additional looping interaction peaks in each cell type. Intriguingly, many of these newly-identified peaks are localized along stripe patterns and form transitive grids, consistent with their anchors being pause sites impeding the process of cohesin-dependent loop extrusion. Together, our analyses provide the highest resolution maps of chromosome folding in human cells to date, and provide a valuable resource for studies of chromosome folding mechanisms.
Frontoparietal connectivity correlates with working memory performance in multiple sclerosis
Authors: Figueroa-Vargas Alejandra , Cárcamo Claudia , Henríquez-Ch Rodrigo , Zamorano Francisco , Ciampi Ethel , Uribe Reinaldo , Vásquez Macarena , Aboitiz Francisco , Billeke Pablo 
Published: May 2019

AbstractWorking Memory (WM) impairment is the most common cognitive deficit of Multiple Sclerosis (MS) patients. However, evidence of its neurobiological mechanisms is scarce. Here we recorded electroencephalographic activity of twenty patients with relapsing-remitting MS and minimal cognitive deficit, and 20 healthy control (HC) subjects while they solved a WM task. In spite of similar performance, the HC group demonstrated both a correlation between temporoparietal theta activity and memory load, and a correlation between medial frontal theta activity and successful memory performances. MS patients did not show theses correlations leading significant differences between groups. Moreover, cortical connectivity analyses using granger causality and phase-amplitude coupling between theta and gamma revealed that HC group, but not MS group, presented a load-modulated progression of the frontal-to-parietal connectivity. This connectivity correlated with working memory capacity in MS groups. This early alterations in the oscillatory dynamics underlaying working memory could be useful for plan therapeutic interventions
Testing implications of the polygenic model for the genetic analysis of loci identified through genome-wide association
Authors: Zhang Wenyu , Reeves R. Guy , Tautz Diethard 
Published: May 2019

AbstractIt has been proposed that many loci with no significant association in GWA studies can nonetheless contribute to the phenotype through modifier interactions with the core genes, implying a polygenic determination of quantitative traits. We have tested this hypothesis by using Drosophila pupal phenotypes. We identified candidate genes for pupal length determination in a GWA and show for disrupted versions of the genes that most are indeed involved in the phenotype, presumably forming a core pathway. We then randomly chose genes below the GWA threshold and found that three quarters of them had also an effect on the trait. We further tested the effects of these knockout lines on an independent behavioral pupal trait (pupation site choice) and found that a similar, but non-correlated fraction of them had a significant effect as well. Our data thus confirm the prediction that a large number of genes can influence independent quantitative traits.Impact statementQuantitative traits are similarly likely influenced by randomly picked loci as by loci identified in a genome-wide association study.
DNA accessibility is not the primary determinant of chromatin-mediated gene regulation
Authors: Chereji Răzvan V. , Eriksson Peter R. , Ocampo Josefina , Clark David J. 
Published: May 2019

ABSTRACTDNA accessibility is thought to be of major importance in regulating gene expression. We test this hypothesis using a restriction enzyme as a probe of chromatin structure and as a proxy for transcription factors. We measured the digestion rate and the fraction of accessible DNA at all genomicAluI sites in budding yeast and mouse liver nuclei. Hepatocyte DNA is more accessible than yeast DNA, consistent with longer linkers between nucleosomes, and indicating that nucleosome spacing is a major determinant of accessibility. DNA accessibility varies from cell to cell, such that essentially no sites are accessible or inaccessible in every cell.AluI sites in inactive mouse promoters are accessible in some cells, implying that transcription factors could bind without activating the gene. Euchromatin and heterochromatin have very similar accessibilities, suggesting that transcription factors can penetrate heterochromatin. Thus, DNA accessibility is not likely to be the primary determinant of gene regulation.
High-Throughput Single-Molecule R-loop Footprinting Reveals Principles of R-loop Formation
Authors: Malig Maika , Hartono Stella R. , Giafaglione Jenna M. , Sanz Lionel A. , Chedin Frederic 
Published: May 2019

ABSTRACTR-loops are a prevalent class of non-B DNA structures that form during transcription upon reannealing of the nascent RNA to the template DNA strand. R-loops have been profiled using the S9.6 antibody to immunoprecipitate DNA:RNA hybrids. S9.6-based DNA:RNA immunoprecipitation (DRIP) techniques revealed that R-loops form dynamically over conserved genic hotspots. We developed an orthogonal profiling methodology that queries R-loops via the presence of long stretches of single-stranded DNA on the looped-out strand. Non-denaturing sodium bisulfite treatment catalyzes the conversion of unpaired cytosines to uracils, creating permanent genetic tags for the position of an R-loop. Long read, single-molecule PacBio sequencing allows the identification of R-loop ‘footprints’ at near nucleotide resolution in a strand-specific manner on single DNA molecules and at ultra-deep coverage. Single-molecule R-loop footprinting (SMRF-seq) revealed a strong agreement between S9.6-and bisulfite-based R-loop mapping and confirmed that R-loops form from unspliced transcripts over genic hotspots. Using the largest single-molecule R-loop dataset to date, we show that individual R-loops generate overlapping sets of molecular clusters that pile-up through larger R-loop-prone zones. SMRF-seq further established that R-loop distribution patterns are driven by both intrinsic DNA sequence features and DNA topological constraints, revealing the principles of R-loop formation.
Three-dimensional morphological analysis of the dynamic digestive system in the green brittle star
Authors: Wakita Daiki , Naniwa Keisuke , Aonuma Hitoshi 
Published: May 2019

AbstractBrittle stars (Echinodermata: Ophiuroidea) digest a great diversity of food in their stomach, which widely lies in the central disk. As for a possible digestive activity, the green brittle starOphiarachna incrassata(Lamarck, 1816) is known to show a dynamic movement at the disk. This phenomenon would deeply involve the morphological structure of the stomach. However, past anatomical studies have shown the digestive system in two dimensions after wide incision of the body wall anchoring the stomach. This methodology restrains us from understanding how the stomach actually shapes inside a brittle star. We aim to visualize the morphology of brittle stars’ digestive system in a non-destructive and three-dimensional way, with a comparison between a relaxed specimen and a specimen fixed at the very moment of the disk’s movement. Employing X-ray micro-computed tomography (micro-CT) and introducing an instant freezing method with cryogenic ethanol, we found the stomach wholly transformed during the movement. We here brought transparency to thein vivoposition of gut contents to hint the mechanism and digestive function of the movement. Our outcome spotlights a dynamic digestive process in echinoderms and a widely applicable method for probing into its relation with body structure.
When will the cancer start? Elucidating the correlations between cancer initiation times and lifetime cancer risks
Authors: Teimouri Hamid , Kochugaeva Maria , Kolomeisky Anatoly B. 
Published: May 2019

AbstractCancer is a genetic disease that results from accumulation of unfavorable mutations. As soon as genetic and epigenetic modifications associated with these mutations become strong enough, the uncontrolled tumor cell growth is initiated, eventually spreading through healthy tissues. Clarifying the dynamics of cancer initiation is thus critically important for understanding the molecular mechanisms of tumorigenesis. Here we present a new theoretical method to evaluate the dynamic processes associated with the cancer initiation. It is based on a discrete-state stochastic description of the formation of tumors as a fixation of unfavorable mutations in tissues. Using a first-passage analysis the probabilities for the cancer to appear and the times before it happens, which are viewed as fixation probabilities and fixation times, respectively, are explicitly calculated. It is predicted that the slowest cancer initiation dynamics is observed for neutral mutations, while it is fast for both advantageous and, surprisingly, disadvantageous mutations. The method is applied for estimating the cancer initiation times from experimentally available lifetime cancer risks for different types of cancer. It is found that the higher probability of the cancer to occur does not necessary lead to the fast times of starting the cancer. Our theoretical analysis helps to clarify microscopic aspects of cancer initiation processes.
Variability in human drug metabolizing cytochrome P450 CYP2C9, CYP2C19 and CYP3A5 activities caused by genetic variations in cytochrome P450 oxidoreductase
Authors: Velazquez Maria Natalia Rojas , Parween Shaheena , Udhane Sameer S , Pandey Amit V 
Published: May 2019

AbstractA broad spectrum of human diseases are caused by mutations in the NADPH cytochrome P450 oxidoreductase (POR). Cytochrome P450 proteins perform several reactions, including the metabolism of steroids, drugs, and other xenobiotics. In 2004 the first human patients with defects in POR were reported, and over 250 variations in POR are known. Information about the effects of POR variants on drug metabolizing enzymes is limited and has not received much attention. By analyzing the POR sequences from genomics databases, we identified potentially disease-causing variations and characterized these by in vitro functional studies using recombinant proteins. Proteins were expressed in bacteria and purified for activity assays. Activities of cytochrome P450 enzymes were tested in vitro using liposomes prepared with lipids into which P450 and P450 reductase proteins were embedded. Here we are reporting the effect of POR variants on drug metabolizing enzymes CYP2C9, CYP2C19, and CYP3A5 which are responsible for the metabolism of many drugs. POR Variants A115V, T142A, A281T, P284L, A287P, and Y607C inhibited activities of all P450 proteins tested. Interestingly, the POR variant Q153R showed a reduction of 20-50% activities with CYP2C9 and CYP2C19 but had a 400% increased activity with CYP3A5. The A287P is most common POR mutation found in patients of European origin, and significantly inhibited drug metabolism activities which has important consequences for monitoring and treatment of patients. In vitro, functional assays using recombinant proteins provide a useful model for establishing the metabolic effect of genetic mutations. Our results indicate that detailed knowledge about POR variants is necessary for correct diagnosis and treatment options for persons with POR deficiency and the role of changes in drug metabolism and toxicology due to variations in POR needs to be addressed.
Cooperation and spatial self-organization determine ecosystem function for polysaccharide-degrading bacteria
Authors: Ebrahimi Ali , Schwartzman Julia , Cordero Otto X. 
Published: May 2019

AbstractThe recycling of particulate organic matter (POM) by microbes is a key part of the global carbon cycle, one which is mediated by the extracellular hydrolysis of polysaccharides and the production of public goods that can trigger social behaviors in bacteria. Despite the potential importance of these microbial interactions, their role in regulating of ecosystem function remains unclear. In this study, we developed a computational and experimental model system to address this challenge and studied how POM depolymerization rate and its uptake efficiency –two main ecosystem function parameters– depended on social interactions and spatial self-organization on particle surfaces. We found an emergent trade-off between rate and efficiency resulting from the competition between oligosaccharide diffusion and cellular uptake, with low rate and high efficiency being achieved through cell-to-cell cooperation between degraders. Bacteria cooperated by aggregating in cell-clusters of ~10-20μm, where cells were able to share public goods. This phenomenon, which was independent of any explicit group-level regulation, led to the emergence of critical cell concentrations below which degradation did not occur, despite all resources being available in excess. By contrast, when particles were labile and turnover rates were high, aggregation promoted competition and decreased the efficiency of carbon utilization. Our study shows how social interactions and cell aggregation determine the rate and efficiency of particulate carbon turnover in environmentally relevant scenarios.Significance StatementMicroorganisms can cooperate by secreting public goods that benefit local neighbors, however, the impact of cooperation on ecosystem functions remains poorly constrained. We here pair computation and experiment to show that bacterial cooperation mediates the degradation of polysaccharide particles recalcitrant to hydrolysis in aquatic environments. On particle surfaces, cooperation emerges through the self-organization of cells into ~10-20μm clusters that promote cooperative uptake of hydrolysis products. The transition between cooperation and competition in aggregates is mitigated by individual cell behaviors such as motility and chemotaxis, that promote reorganization on the particle surface. When cooperation is required, the degradation of recalcitrant biopolymers can only take place when degraders exceed a critical cell concentration, underscoring the importance of microbial interactions for ecosystem function.
The role of ATP hydrolysis in conformational changes of human P-glycoprotein in living cells
Authors: Futamata Ryota , Ogasawara Fumihiko , Ichikawa Takafumi , Kodan Atsushi , Kimura Yasuhisa , Kioka Noriyuki , Ueda Kazumitsu 
Published: May 2019

AbstractP-glycoprotein (P-gp; also known as MDR1 or ABCB1) is an ATP-driven multidrug transporter that extrudes various hydrophobic toxic compounds to the extracellular space. P-gp consists of two transmembrane domains (TMDs) that form the substrate translocation pathway and two nucleotide-binding domains (NBDs) that bind and hydrolyze ATP. P-gp takes at least two states during transport; the inward-facing (pre-drug transport) conformation, in which the two NBDs are separated and the two TMDs are open to the intracellular side, and the outward-facing (post-drug transport) conformation, in which the NBDs are dimerized and the TMDs are slightly open to the extracellular side. ATP binding and hydrolysis cause conformational changes between the inward-facing and the outward-facing conformations to translocate substrates across the membrane. However, it remains unclear how ATP is used during these conformational changes in living cells. In this study, we investigated the role of ATP binding and hydrolysis during the conformational changes of human P-gp in living cells by using fluorescence resonance energy transfer (FRET). We show that ATP binding causes the conformational change to the outward-facing state and that ATP hydrolysis and subsequent release of γ-phosphate from both NBDs allow the outward-facing state to return to the original inward-facing state.
A bacterial effector protein prevents MAPK-mediated phosphorylation of SGT1 to suppress plant immunity
Authors: Yu Gang , Xian Liu , Xue Hao , Yu Wenjia , Rufian Jose S. , Sang Yuying , Morcillo Rafael J. L. , Wang Yaru , Macho Alberto P. 
Published: May 2019

AbstractNucleotide-binding domain and leucine-rich repeat-containing (NLR) proteins function as sensors that perceive pathogen molecules and activate immunity. In plants, the accumulation and activation of NLRs is regulated by SUPPRESSOR OF G2 ALLELE OF skp1 (SGT1). In this work, we found that an effector protein named RipAC, secreted by the plant pathogen Ralstonia solanacearum, associates with SGT1 to suppress NLR-mediated SGT1-dependent immune responses, including those triggered by another R. solanacearum effector, RipE1. RipAC does not affect the accumulation of SGT1 or NLRs, or their interaction. However, RipAC inhibits the interaction between SGT1 and MAP kinases, and the phosphorylation of a MAPK target motif in the C-terminal domain of SGT1. Such phosphorylation is enhanced upon activation of immune signaling, leads to the release of the interaction between SGT1 and NLRs, and contributes to the activation of NLR-mediated responses. Additionally, SGT1 phosphorylation contributes to resistance against R. solanacearum, and this is particularly evident in the absence of RipAC. Our results shed light onto the mechanism of activation of NLR-mediated immunity, and suggest a positive feedback loop between MAPK activation and SGT1-dependent NLR activation.