bluuebunny/crossref_metadata_2025_split huggingface dataset:
VirtuousPocketome: A Computational Tool for Screening Protein-ligand Complexes to Identify Similar Binding Sites
Authors: Pallante Lorenzo , Cannariato Marco , Androutsos Lampros , Zizzi Eric A. , Bompotas Agorakis , Hada Xhesika , Grasso Gianvito , Kalogeras Athanasios , Mavroudi Seferina , di Benedetto Giacomo , Theofilatos Konstantinos , Deriu Marco A. 
Published: December 2023

ABSTRACTProtein residues within binding pockets play a critical role in determining the range of ligands that can interact with a protein, influencing its structure and function. Identifying structural similarities in proteins offers valuable insights into their function and activation mechanisms, aiding in predicting protein-ligand interactions, anticipating off-target effects, and facilitating the development of therapeutic agents. Numerous computational methods assessing global or local similarity in protein cavities have emerged, but their utilization is impeded by complexity, impractical automation for amino acid pattern searches, and an inability to evaluate the dynamics of scrutinized protein-ligand systems. Here, we present a general, automatic and unbiased computational pipeline, namedVirtuousPocketome, aimed at screening huge databases of proteins for similar binding pockets starting from an interested protein-ligand complex. We demonstrate the pipeline’s potential by exploring a recently-solved human bitter taste receptor, i.e. the TAS2R46, complexed with strychnine. We pinpointed 145 proteins sharing similar binding sites compared to the analysed bitter taste receptor and the enrichment analysis highlighted the related biological processes, molecular functions and cellular components. This work represents the foundation for future studies aimed at understanding the effective role of tastants outside the gustatory system: this could pave the way towards the rationalization of the diet as a supplement to standard pharmacological treatments and the design of novel tastants-inspired compounds to target other proteins involved in specific diseases or disorders. The proposed pipeline is publicly accessible, can be applied to any protein-ligand complex, and could be expanded to screen any database of protein structures.
A cyclic peptide toolkit reveals mechanistic principles of peptidylarginine deiminase IV (PADI4) regulation
Authors: Bertran M. T. , Walmsley R. , Cummings T. , Valle Aramburu I. , Benton D. J. , Assalaarachchi J. , Chasampalioti M. , Swanton T. , Joshi D. , Federico S. , Okkenhaug H. , Yu L. , Oxley D. , Walker S. , Papayannopoulos V. , Suga H. , Christophorou M. A. , Walport L. J. 
Published: December 2023

AbstractPeptidylarginine deiminase IV (PADI4) deregulation promotes the development of autoimmunity, cancer, atherosclerosis and age-related tissue fibrosis. Genetic or pharmacological PADI4 inhibition is therapeutically efficacious in mice, but no clinically relevant inhibitors currently exist. PADI4 additionally mediates immune responses and cellular reprogramming, although the full extent of its physiological roles is unexplored. Despite detailed molecular knowledge of PADI4 activationin vitro, we lack understanding of its regulation within cells, largely due to lack of appropriate systems and tools. Here, we developed and applied a set of potent and selective PADI4 modulators. Using the mRNA- display-based RaPID system, we screened >1012cyclic peptides for high-affinity, conformation-selective binders. We report PADI4_3, an inhibitor specific for the active conformation of PADI4; PADI4_7, an inert binder, which we functionalised for the isolation and study of cellular PADI4; and PADI4_11, a first-in-class activator. Using a newly developed method for the quantification of cellular PADI4 activity, we show that PADI4_3 and PADI4_11 are effective in cells. Structural studies with PADI4_11 reveal an allosteric binding mode that may reflect the mechanism that promotes cellular PADI4 activation. This work offers new understanding of PADI4 regulation and a toolkit for the study and modulation of PADI4 across physiological and pathophysiological contexts.
Fixed-target time-resolved crystallography at XFELs: the scourge of light contamination but reduced sample consumption
Authors: Gotthard Guillaume , Flores-Ibarra Andrea , Carrillo Melissa , Kepa Michal W , Mason Thomas J , Stegmann Dennis P. , Olasz Bence , Pachota Magdalena , Dworkowski Florian , Ozerov Dmitry , Pedrini Bill F. , Padeste Celestino , Beale John H , Nogly Przemyslaw 
Published: December 2023

AbstractX-ray free electron laser (XFEL) light sources have allowed for the rapid growth of time-resolved structural experiments, which provide crucial information on the function of biological machines and their mechanisms. We set out to commission the SwissMX fixed-target sample delivery system at the SwissFEL Cristallina experimental station using the PSI developed MISP-chip for pump-probe time-resolved experiments. To characterise the system, we used the light-sensitive protein crystals of the Light-Oxygen-Voltage domain 1 (LOV1) fromChlamydomonas reinhardtii. Using different experimental settings, the adjacent-well light contamination was carefully assessed, indicating that it is crucial to control the light scattering from solid supports otherwise significant contamination can occur. However, our results show that, after the initial experiments and parameter refinement, the opaque MISP-chips are suitable for pump-probing a light-sensitive protein. This crystallographic experiment also probed the sub-millisecond structural dynamics of the LOV1 and indicated that at Δt=10 μs the covalent thioether bond is already established between the reactive Cys57 and FMN cofactor. This experiment validated the crystals to be suitable for in-depth follow up studies of the still poorly understood signal transduction mechanism. Importantly, the fixed-target delivery system also permitted a tenfold reduction in protein sample consumption compared to the most successful system used at XFEL, the high-viscosity extruder. This development creates the prospect of an exciting increase in XFEL project throughput for the field.
Mucosal washes are useful for sampling intestinal mucus-associated microbiota despite low biomass
Authors: Martinez-Medina Jennifer N , Ghazisaeedi Fereshteh , Kramer Catharina , Ziegler Jörn F , McParland Victoria , Siegmund Britta , Jarquín-Díaz Víctor Hugo , Fulde Marcus , Forslund Sofia K 
Published: December 2023

AbstractExploring the dynamic relationship between mucus-associated microbiota and host health is pivotal, yet prevalent studies using stool samples may not accurately represent these bacteria. Here, we explored mucus-associated microbiota in the gastrointestinal tract of mice and the terminal ileum in humans, using three different sample types: mucosal washes, scraping, and intestinal content in mice and biopsies and mucosal washes in humans. We employed DNA quantification and 16S rRNA sequencing to assess how comparable the information yielded from different sample types, evaluating findings relative to expectations from state-of-the-art and under controlled benchmarks.Mucosal washes in mice exhibited higher bacterial DNA and lower host DNA contamination than scraping samples. Similarly, in humans, washes surpassed biopsies in bacterial yield. Despite variations in read counts, microbiota diversity and composition remained remarkably similar between methods in both species, faithfully reflecting expected genotypic and phenotypic differences.We conclude that washes reduce host contamination without inducing substantial compositional bias when sampling mucosal microbiota. Our findings emphasize mucosal washes as alternatives to biopsies in humans and scrapings in mice, providing insights for improving result transferability across hosts. Our research underscores the importance of considering the mucus-associated microbiota to track host-microbiome interactions closer to their actual interface surface.
Myrmecophytism as a driver of macroevolutionary patterns: perspectives from the Southeast AsianMacarangaant-plant symbiosis
Authors: Dixit Nadi M , Guicking Daniela 
Published: December 2023

AbstractMyrmecophytes utilise defensive services offered by obligate ant partners in a novel means of survival in tropical habitats. Although much is known about the ecology of myrmecophytism, there aren’t enough empirical examples to demonstrate whether it substantially influences evolutionary patterns in host plant lineages. In this study, we make use of the species-richMacaranga(Euphorbiaceae) ant-plant symbiosis distributed in Sundaland to delve into the evolutionary dynamics of myrmecophytism in host plants. We generated the most comprehensive dated phylogeny of myrmecophyticMacarangatill date using sequences derived from genotyping-by-sequencing (GBS), a next-generation sequencing (NGS) technique. With this in hand, we traced the evolutionary history of myrmecophytism inMacarangausing parametric biogeography and ancestral state reconstruction. Diversification rate analysis methods were employed to determine if myrmecophytism enhanced diversification rates in the genus. Our results demonstrate that myrmecophytism is plastic and easily lost unless it is overly specialised. Ancestral state reconstruction supported a single origin of myrmecophytism inMacaranga∼18 mya on Borneo followed by multiple losses. Diversification rate analysis methods did not yield sufficient evidence to support the hypothesis that myrmecophytism enhanced diversification rates inMacaranga; we found that topographical features on Borneo may have played a more direct role in the divergence of clades instead. Through this comprehensive investigation of the evolution of myrmecophytism inMacaranga, our study also provides evidence that a key innovation may not necessarily enhance diversification rates. In fact, we hypothesise that overly specialised cases of myrmecophytism may even be an evolutionary dead end.
Aberrant Wnt activation in recurrent genetically variant human pluripotent stem cells impairs cardiomyocyte differentiation and phenotype
Authors: Wing Theodore , Price Christopher J. , Stavish Dylan , Laing Owen , Riley Jack J. , Lam Alan , Oh Steve , Atlasi Yaser , Barbaric Ivana 
Published: December 2023

AbstractHuman pluripotent stem cell (hPSC)-derived cardiomyocytes have emerged as powerful tools for disease modelling and cell therapy. The production of cardiomyocytes from hPSCs typically requires expanding large numbers of hPSCs and maintaining them in culture for extended periods of time. This in turn predisposes hPSCs to the acquisition of non-random genetic changes, including recurrent gains of chromosome 1q. Here, we show that gain of chromosome 1q in hPSCs affects both the efficiency of differentiation to cardiomyocytes and phenotype of the differentiated cells. Mechanistically, we show that aberrant activation of the Wnt signalling pathway underpins the skewed differentiation of variant 1q hPSCs. Collectively, our data demonstrates that the presence of genetically variant cells in cultures is a significant concern for production of hPSC-derived cardiomyocytes for research or clinical applications. Further, our results suggest new approaches for removing genetically variant cells for future clinical applications.
The impact of PCR duplication on RNAseq data generated using NovaSeq 6000, NovaSeq X, AVITI and G4 sequencers
Authors: Zajac Natalia , Vlachos Ioannis S , Sajibu Sija , Opitz Lennart , Wang Shuoshuo , Chittur Sridar V , Mason Christopher E. , Knudtson Kevin L , Ashton John M , Rehrauer Hubert , Aquino Catharine 
Published: December 2023

AbstractRNA sequencing (RNA-seq) is a powerful technology for gene expression and functional genomics profiling. Expression profiles generated using this approach can be impacted by the methods utilised for cDNA library generation. Selection of the optimal parameters for each step during the protocol are crucial for acquisition of high-quality data. Polymerase chain reaction (PCR) amplification of transcripts is a common step in many RNA-seq protocols and, if not optimised, high PCR duplicate proportions can be generated, resulting in the inflation of transcript counts and introduction of bias. In this study, we investigate the impact of input amount and PCR cycle number on the PCR duplication rate and on the RNA-seq data quality using a broad range of inputs (1 ng -1,000 ng) for RNA-seq library preparation with unique molecular identifiers (UMIs) and sequencing the data on four different short-read sequencing platforms: Illumina NovaSeq 6000, Illumina NovaSeq X, Element Biosciences AVITI, and Singular Genomics G4. Across all platforms, samples of input amounts greater than 125 ng had a negligible PCR duplication rate and the number of PCR cycles did not have a significant effect on data quality. However, for input amounts lower than 125ng we observed a strong negative correlation between input amount and the proportion of PCR duplicates; between 34% and 96% of reads were discarded via deduplication. Fortunately, UMIs were effective for removingin silicoPCR duplicates without removing valuable biological information. Removal of PCR duplicates resulted in more comparable gene expression obtained from the different PCR cycles. Data generated with each of the four sequencing platforms presented similar associations between starting material amount and the number of PCR cycles on PCR duplicates, a similar number of genes detected, and comparable gene expression profiles. However, the sequencers using conversion kits for Illumina libraries (AVITI, G4) exhibited lower adapter dimer abundance across all input amounts, but also a higher PCR duplication rate in very low input amounts (<15ng). Overall, this study showed that the choice of input amount and number of PCR cycles are important parameters for obtaining high-quality RNA-seq data across all sequencing platforms. UMI deduplication is an effective way to remove PCR duplicates, improving the data quality and removing any variation caused by the conversion kits.
A Sensitivity Analysis of Methodological Variables Associated with Microbiome Measurements
Authors: Forry Samuel P. , Servetas Stephanie L. , Dootz Jennifer N. , Hunter Monique E. , Kralj Jason G. , Filliben James J. , Jackson Scott A. 
Published: December 2023

AbstractThe experimental methods employed during metagenomic sequencing analyses of microbiome samples significantly impact the resulting data and typically vary substantially between laboratories. In this study, a full factorial experimental design was used to compare the effects of a select set of methodological choices (sample, operator, lot, extraction kit, variable region, reference database) on the analysis of biologically diverse stool samples. For each parameter investigated, a main effect was calculated that allowed direct comparison both between methodological choices (bias effects) and between samples (real biological differences). Overall, methodological bias was found to be similar in magnitude to real biological differences, while also exhibiting significant variations between individual taxa, even between closely related genera. The quantified method biases were then used to computationally improve the comparability of datasets collected under substantially different protocols. This investigation demonstrates a framework for quantitatively assessing methodological choices that could be routinely performed by individual laboratories to better understand their metagenomic sequencing workflows and to improve the scope of the datasets they produce.
Mechanism of Dimer Selectivity and Binding Cooperativity of BRAF Inhibitors
Authors: Clayton Joseph , Romany Aarion , Matenoglou Evangelia , Gavathiotis Evripidis , Poulikakos Poulikos I. , Shen Jana 
Published: December 2023

AbstractAberrant signaling of BRAFV600Eis a major cancer driver. Current FDA-approved RAF inhibitors selectively inhibit the monomeric BRAFV600Eand suffer from tumor resistance. Recently, dimer-selective and equipotent RAF inhibitors have been developed; however, the mechanism of dimer selectivity is poorly understood. Here, we report extensive molecular dynamics (MD) simulations of the monomeric and dimeric BRAFV600Ein the apo form or in complex with one or two dimer-selective (PHI1) or equipotent (LY3009120) inhibitor(s). The simulations uncovered the unprecedented details of the remarkable allostery in BRAFV600Edimerization and inhibitor binding. Specifically, dimerization retrains and shifts the αC helix inward and increases the flexibility of the DFG motif; dimer compatibility is due to the promotion of the αC-in conformation, which is stabilized by a hydrogen bond formation between the inhibitor and the αC Glu501. A more stable hydrogen bond further restrains and shifts the αC helix inward, which incurs a larger entropic penalty that disfavors monomer binding. This mechanism led us to propose an empirical way based on the co-crystal structure to assess the dimer selectivity of a BRAFV600Einhibitor. Simulations also revealed that the positive cooperativity of PHI1 is due to its ability to preorganize the αC and DFG conformation in the opposite protomer, priming it for binding the second inhibitor. The atomically detailed view of the interplay between BRAF dimerization and inhibitor allostery as well as cooperativity has implications for understanding kinase signaling and contributes to the design of protomer selective RAF inhibitors.
INTEGRATED GLOBAL CHICKEN REFERENCE PANEL FROM 13,187 CHICKEN GENOMES
Authors: Zhu Di , Wang Yuzhan , Qu Hao , Feng Chugang , Zhang Hui , Sheng Zheya , Jiang Yuliang , Nie Qinghua , Chu Suqiao , Shu Dingming , Zhang Dexiang , Fang Lingzhao , Zhao Yiqiang , Wang Yuzhe , Hu Xiaoxiang 
Published: December 2023

ABSTRACTChickens are a crucial source of protein for humans and a popular model animal for bird research. Despite the emergence of imputation as a reliable genotyping strategy for large populations, the lack of a high-quality chicken reference panel has hindered progress in chicken genome research. To address this issue, here we introduce the first phase of the 100 K Global Chicken Reference Panel Project (100 K GCRPP). The project includes 13,187 samples and provides services for varied applications on its website (http://farmrefpanel.com/GCRP/). Currently, two panels are available: a Comprehensive Mix Panel (CMP) for domestication diversity research and a Commercial Breed Panel (CBP) for breeding broilers specifically. Evaluation of genotype imputation quality showed that CMP had the highest imputation accuracy compared to imputation using existing chicken panel in animal SNPAtlas, whereas CBP performed stably in the imputation of commercial populations. Additionally, we found that genome-wide association studies using GCRP-imputed data, whether on simulated or real phenotypes, exhibited greater statistical power. In conclusion, our study indicates that the GCRP effectively fills the gap in high-quality reference panels for chickens, providing an effective imputation platform for future genetic and breeding research.
Adipose Tissue Analysis Toolkit (ATAT) for Automated Analysis of Adipocyte Size and Extracellular Matrix in White Adipose Tissue
Authors: Robino Jacob J. , Plekhanov Alexander P. , Zhu Qingzhang , Jensen Michael D. , Scherer Philipp E. , Roberts Charles T. , Varlamov Oleg 
Published: December 2023

AbstractObjectiveThe pathological expansion of white adipose tissue (WAT) in obesity involves adipocyte hypertrophy accompanied by expansion of collagen-rich pericellular extracellular matrix (ECM) and the development of crown-like structures (CLS). Traditionally, WAT morphology is assessed through immunohistochemical analysis of WAT sections. However, manual analysis of large histological sections is time-consuming, and available digital tools for analyzing adipocyte size and pericellular ECM are limited. To address this gap, we developed the Adipose Tissue Analysis Toolkit (ATAT), an ImageJ plugin facilitating analysis of adipocyte size, WAT ECM and CLS.Methods and ResultsATAT utilizes local and image-level differentials in pixel intensity to independently threshold background, distinguishing adipocyte-free tissue without user input. It accurately captures adipocytes in histological sections stained with common dyes and automates the analysis of adipocyte cross-sectional area, total-field, and localized region-of-interest ECM. ATAT allows fully automated batch analysis of histological images using default or user-defined adipocyte detection parameters.ConclusionsATAT provides several advantages over existing WAT image analysis tools, enabling high-throughput analyses of adipocyte-specific parameters and facilitating the assessment of ECM changes associated with WAT remodeling due to weight changes and other pathophysiological alterations that affect WAT function.Study Importance QuestionsWhat is already known about this subject?The manual analysis of large WAT histological sections is very time-consuming, while digital tools for the analysis of WAT are limited.What are the new findings in your manuscript?-ATAT enables fully automated analysis of batches of histological images using either default or user-defined adipocyte detection parameters-ATAT allows high-throughput analyses of adipocyte-specific parameters and pericellular extracellular matrix-ATAT enables the assessment of fibrotic changes associated with WAT remodeling and crown-like structuresHow might your results change the direction of research or the focus of clinical practice?-ATAT is designed to work with histological sections and digital images obtained using a slide scanner or a microscope.-This tool will help basic and clinical researchers to conduct automated analyses of adipose tissue histological sections.
Cysteine String Protein alpha in Extracellular Vesicle Subtypes: a Proteomic Analysis
Authors: Nogueira de Almeida Luiz Gustavo , Armstrong Victoria , Dufour Antoine , Braun Janice E.A. 
Published: December 2023

AbstractCysteine string protein (CSPα /DnaJC5) is a presynaptic J-domain protein (JDP) that prevents neurodegeneration. CSPα/DnaJC5 is reported to facilitate export of distinct, highly oligomeric, disease-causing proteins in addition to wild-type TDP-43, tau and α-synuclein. Yet, detailed mechanistic knowledge of the full CSPα/DnaJC5 secreted proteome is lacking. Understanding the CSPα/DnaJC5 export pathway has implications for a growing number of neurodegenerative diseases. In humans, Leu115Arg or Leu116deletion mutations cause adult-onset neuronal ceroid lipofusinosis (ANCL), a rare neurodegenerative disorder. In the present study, we examined extracelular vesicles (EVs) released from CSPα/DnaJC5 expressing cells. Cells are known to secrete many types of EVs of different sizes and origins into the extracellular space. EV subpopulations were separated by their sedimentation speed and subjected to proteomic analysis. We find that CSPα/DnaJC5 and the CSPα/DnaJC5 mutants, Leu115Arg or Leu116del are enriched in multiple EV subpopulations. The exported protein profile is determined by proteomics. We report that several other J-domain proteins (JDPs), such as DnaJC7, DnaJA1 and DnaJA2 are exported and speculate that export of JDPs may facilitate the secretion of diverse client proteins. Our work provides a platform for further inquiry into the role of secreted CSPα/DnaJC5 and other JDPs in proteostasis.