bluuebunny/crossref_metadata_2025_split huggingface dataset:
Dynamics of the CD9 interactome during bacterial infection of epithelial cells by proximity labelling proteomics
Authors: Wolverson PA , Fernandes Parreira I , Collins MO , Shaw JG , Green LR 
Published: December 2024

AbstractEpithelial colonisation is often a critical first step in bacterial pathogenesis, however, different bacterial species utilise several different receptors at the cell membrane to adhere to cells. We have previously demonstrated that interference of the human tetraspanin, CD9, can reduce adherence of multiple species of bacteria to epithelial cells by approximately 50%. However, CD9 does not act as a receptor and is responsible for organising and clustering partner proteins commandeered by bacteria for efficient adherence. CD9 can organise numerous host proteins at the cell membrane but the full interactome has not been delineated. Here, using a novel CD9 proximity-labelling model, we demonstrate a vast and diverse CD9 interactome with 845 significantly enriched proteins associated with CD9 over four hours. These putative proximal proteins were associated with various cellular pathways including cell adhesion, ECM-receptor interactions, endocytosis, SNARE interactions and adherens and tight junctions. Significant and known interactors of CD9 were enriched including β1 integrins and major immunoglobulin superfamily members but also included several known bacterial adherence receptors including CD44, CD46 and CD147. We further demonstrate dynamism of the interactome during infection at three separate time points with two different bacterial species,Neisseria meningitidisandStaphylococcus aureus. During meningococcal infection, 13 unique proximal proteins associated with CD9 were significantly enriched across four hours compared to uninfected cells. However, upon staphylococcal infection far fewer enriched proximal proteins were identified demonstrating that different bacteria require different host factors during CD9-mediated bacterial adherence. Transient knockdown of CD44 and CD147, candidate receptor proteins identified in our screen, significantly reduced staphylococcal and meningococcal adherence respectively. This effect was ablated in the absence of CD9 or if epithelial cells were treated with a CD9-derived peptide demonstrating the association of these proteins during staphylococcal and meningococcal adherence. We demonstrate for the first time the CD9 interactome of epithelial cells and that bacteria hijack these interactions to efficiently adhere to epithelial cells. This process is bacterial species specific, recruiting several different proteins during infection but a host-derived peptide is able to interfere with this process. We have therefore developed a tool that can measure changes within the CD9 interactome after cellular challenge, established a mechanism in which CD9 is used as a universal organiser of bacterial adhesion platforms and demonstrated that this process can be stopped using a CD9-derived peptide.
Typhoid toxin ofSalmonellaTyphi elicits host antimicrobial response during acute typhoid fever
Authors: Srour Salma , ElGhazaly Mohamed , O’Connor Daniel , Gibani Malick M , Darton Thomas C , Pollard Andrew J , Collins Mark O , Humphreys Daniel 
Published: December 2024

SummarySalmonellaTyphi secretes typhoid toxin that activates cellular DNA damage responses (DDR) during acute typhoid fever. Human infection challenge studies revealed that the toxin suppresses bacteraemia via unknown mechanisms. Using quantitative proteomic analysis on the plasma of bacteraemic participants, we demonstrate that wild-type toxigenicSalmonellainduced secretion of lysozyme (LYZ) and apolipoprotein C3 (APOC3). Recombinant typhoid toxin or infection with toxigenicSalmonellarecapitulate LYZ and APOC3 secretion in cultured cells, which involved ATM/ATR-dependent DDRs and confirmed observations in typhoid fever. LYZ alone inhibited secretion of virulence effector proteins SipB and SopE. LYZ caused loss ofSalmonellamorphology characterised by spheroplast formation. Spheroplast formation was mediated by LYZ and enhanced by lactoferrin, which was identified by proteomics in participants with typhoid fever. Our findings may indicate that toxin-induced DDRs elicit antimicrobial responses, which suppressSalmonellabacteraemia during typhoid fever.
Redox-dependent binding and conformational equilibria govern the fluorescence decay of NAD(P)H in living cells
Authors: Blacker Thomas S. , Mistry Nimit , Plotegher Nicoletta , Westbrook Elizabeth R. , Sewell Michael D. E. , Carroll John , Szabadkai Gyorgy , Bain Angus J. , Duchen Michael R. 
Published: December 2024

AbstractWhen probed using fluorescence lifetime imaging microscopy (FLIM), the emission from reduced nicotinamide adenine dinucleotide (NADH) and its phosphorylated form NADPH have shown promise as sensitive intrinsic reporters of metabolism in living systems. However, an incomplete understanding of the biochemical processes controlling their fluorescence decay makes it difficult to draw unambiguous conclusions from NAD(P)H FLIM data. Here we utilised time-resolved fluorescence anisotropy imaging to identify multiple enzyme binding configurations in live cells associated with lifetimes both longer and shorter than unbound NAD(P)H. FLIM, combined with mathematical and computational modelling, revealed that the redox states of the NAD and NADP pools control the steady-state equilibrium of binding configurations, which in-turn determines the observed fluorescence decay. This knowledge will be foundational to developing the accurate interpretation of NAD(P)H FLIM.
Sp140L Is a Novel Herpesvirus Restriction Factor
Authors: Cable Jana M. , Wongwiwat Wiyada , Grabowski Jenna C. , White Robert E. , Luftig Micah A. 
Published: December 2024

SummaryHerpesviruses, including the oncogenic Epstein-Barr Virus (EBV), must bypass host DNA sensing mechanisms to establish infection. The first viral latency protein expressed, EBNA-LP, is essential for transformation of naïve B cells, yet its role in evading host defenses remains unclear. Using single-cell RNA sequencing of EBNA-LP-Knockout (LPKO)-infected B cells, we reveal an antiviral response landscape implicating the ‘speckled proteins’ as key restriction factors countered by EBNA-LP. Specifically, loss ofSP100or the primate-specificSP140Lreverses the restriction of LPKO, suppresses a subset of canonically interferon-stimulated genes, and restores viral gene transcription and cellular proliferation. Notably, we also identify Sp140L as a restriction target of the herpesvirus saimiri ORF3 protein, implying a role in immunity to other DNA viruses. This study reveals Sp140L as a restriction factor that we propose links sensing and transcriptional suppression of viral DNA to an IFN-independent innate immune response, likely relevant to all nuclear DNA viruses.
Engineering prolyl hydroxylase-dependent proteolysis enables the orthogonal control of hypoxia responses in plants
Authors: Shukla Vinay , Iacopino Sergio , Dalle Carbonare Laura , He Yuming , Chiaro Alessia Del , Papachristodoulou Antonis , Giuntoli Beatrice , Licausi Francesco 
Published: December 2024

AbstractVascular plants and metazoans use selective proteolysis of transcription factors to control the adaptive responses to hypoxia, although through distinct biochemical mechanisms. The reason for this divergence is puzzling, especially when considering that the molecular components necessary to establish both strategies are conserved across the two kingdoms. To explore an alternative evolutionary scenario where plants sense hypoxia as animals do, we engineered a three-components system aimed to target proteins for degradation in an oxygen dependent mannerin Arabidopsis thaliana. Applying the synthetic biology framework, we produced a hypoxia-responsive switch independent of endogenous pathways. When applied to control transcription, the synthetic system partially restored hypoxia responsiveness in oxygen-insensitive mutants. Additionally, we demonstrated its potential to regulate growth under flood-induced hypoxia. Our work highlights the use of synthetic biology to reprogram signalling pathways in plants, providing insights into the evolution of oxygen sensing and ofering tools for crop improvement under stress conditions.
YibN, a bona fide interactor of YidC with implications in membrane protein insertion and membrane lipid production
Authors: Zhao Zhiyu , Yamamoto Nachi , Young John W. , Solis Nestor , Fong Amos , Al-Seragi Mohammed , Kim Sungyoung , Aoki Hiroyuki , Phanse Sadhna , Le Hai-Tuong , Overall Christopher M. , Nishikawa Hanako , Babu Mohan , Nishiyama Ken-ichi , Duong van Hoa Franck 
Published: December 2024

AbstractYidC, a prominent member of the Oxa1 superfamily, is essential for the biogenesis of the bacterial inner membrane, significantly influencing its protein composition and lipid organization. It interacts with the Sec translocon, aiding the proper folding of multi-pass membrane proteins. It also functions independently, serving as an insertase and lipid scramblase, augmenting the insertion of smaller membrane proteins while contributing to the organization of the bilayer. Despite the wealth of structural and biochemical data available, how YidC operates remains unclear. To investigate this, we employed proximity-dependent biotin labeling (BioID), leading to the identification of YibN as a crucial component within the YidC protein environment. We then demonstrated the association between YidC and YibN by affinity purification-mass spectrometry assays conducted on native membranes, with further confirmation using on-gel binding assays with purified proteins. Co-expression studies andin vitroassays indicated that YibN enhances the production and membrane insertion of YidC substrates, such as M13 and Pf3 phage coat proteins, ATP synthase subunit c, and various small membrane proteins like SecG. Additionally, the overproduction of YibN was found to stimulate membrane lipid production and promote inner membrane proliferation, perhaps by interfering with YidC lipid scramblase activity. Consequently, YibN emerges as a significant physical and functional interactor of YidC, influencing membrane protein insertion and lipid organization.
The CarSR Two-Component System Directly ControlsradDExpression as a Global Regulator that Senses Bacterial Coaggregation inFusobacterium nucleatum
Authors: Bibek G C , Wu Chenggang 
Published: December 2024

ABSTRACTTwo-component systems (TCS) enable bacteria to sense and respond to environmental signals, facilitating rapid adaptation.Fusobacterium nucleatum, a key oral pathobiont, employs the CarSR TCS to modulate coaggregation with various Gram-positive partners by regulating the expression ofradD, encoding a surface adhesion protein, as revealed by RNA-Seq analysis. However, the direct regulation of theradD-containing operon (radABCD) by the response regulator CarR, the broader CarR regulon, and the signals sensed by this system remain unclear. In this study, chromatin immunoprecipitation followed by high-throughput DNA sequencing (ChIP-seq) identified approximately 161 CarR-enriched loci across the genome and a 17-bp consensus motif that likely serves as the CarR binding site. Notably, one such binding motif was found in the promoter region of theradABCDoperon. The interaction of CarR with this binding motif was further validated using electrophoretic mobility shift assays (EMSA), mutagenesis, and DNase I footprinting analyses. Beyond regulatingradABCD, CarR directly controls genes involved in fructose and amino acid (cysteine, glutamate, lysine) utilization, underscoring its role as a global regulator inF. nucleatum. Lastly, we discovered that RadD-mediated coaggregation enhancesradDexpression, and deletion ofcarSabolished this enhancement, suggesting that coaggregation itself serves as a signal sensed by this TCS. These findings provide new insights into the CarR regulon and the regulation of RadD, elucidating the ecological and pathogenic roles ofF. nucleatumin dental plaque formation and disease processes.IMPORTANCEFusobacterium nucleatumis an essential member of oral biofilms, acting as a bridging organism that connects early and late colonizers, thus driving dental plaque formation. Its remarkable ability to aggregate with diverse bacterial partners is central to its ecological success, yet the mechanisms it senses and responds to these interactions remain poorly understood. This study identifies the CarSR two-component system as a direct regulator of RadD, the primary adhesin mediating coaggregation, and reveals its role in sensing coaggregation as a signal. These findings uncover a novel mechanism by whichF. nucleatumdynamically adapts to polymicrobial environments, offering new perspectives on biofilm formation and bacterial communication in complex oral microbial ecosystems.
A mutualistic model bacterium is lethal to non-symbiotic hosts via the type VI secretion system
Authors: Gaddy Keegan E. , Septer Alecia N. , Mruk Karen , Milton Morgan E. 
Published: December 2024

AbstractWhat makes a bacterium pathogenic? Since the early days of germ theory, researchers have categorized bacteria as pathogens or non-pathogens, those that cause harm and those that do not, but this binary view is not always accurate.Vibrio fischeriis an exclusive mutualistic symbiont found within the light organs of Hawaiian bobtail squid. This symbiotic interaction requiresV. fischerito utilize a range of behaviors and produce molecules that are often associated with pathogenicity. This juxtaposition of employing “pathogenic” behaviors for a symbiotic relationship led the field to focus on howV. fischeriestablishes a beneficial association with its host. In this study, we observe thatV. fischeriinduces mortality in zebrafish embryos andArtemianauplii. Non-lethal doses ofV. fischerileads to zebrafish growth delays and phenotypes indicative of disease. Our data also provide evidence that the conserved type VI secretion system on chromosome I (T6SS1) plays a role in theV. fischeri-induced mortality of zebrafish embryos andArtemianauplii. These results support the hypothesis that theV. fischeriT6SS1 is involved in eukaryotic cell interactions. Despite its traditional view as a beneficial symbiont, we provide evidence thatV. fischeriis capable of harming aquatic organisms, indicating its potential to be pathogenic toward non-symbiotic hosts.
MMETHANE: interpretable AI for predicting host status from microbial composition and metabolomics data
Authors: Dawkins Jennifer J. , Gerber Georg K. 
Published: December 2024

SUMMARYMetabolite production, consumption, and exchange are intimately involved with host health and disease, as well as being key drivers of host-microbiome interactions. Despite the increasing prevalence of datasets that jointly measure microbiome composition and metabolites, computational tools for linking these data to the status of the host remain limited. To address these limitations, we developed MMETHANE, an open-source software package that implements a purpose-built deep learning model for predicting host status from paired microbial sequencing and metabolomic data. MMETHANE incorporates prior biological knowledge, including phylogenetic and chemical relationships, and is intrinsically interpretable, outputting an English-language set of rules that explains its decisions. Using a compendium of six datasets with paired microbial composition and metabolomics measurements, we showed that MMETHANE always performed at least on par with existing methods, including blackbox machine learning techniques, and outperformed other methods on >80% of the datasets evaluated. We additionally demonstrated through two cases studies analyzing inflammatory bowel disease gut microbiome datasets that MMETHANE uncovers biologically meaningful links between microbes, metabolites, and disease status.
MaAsLin 3: Refining and extending generalized multivariable linear models for meta-omic association discovery
Authors: Nickols William A. , Kuntz Thomas , Shen Jiaxian , Maharjan Sagun , Mallick Himel , Franzosa Eric A. , Thompson Kelsey N. , Nearing Jacob T. , Huttenhower Curtis 
Published: December 2024

AbstractA key question in microbial community analysis is determining which microbial features are associated with community properties such as environmental or health phenotypes. This statistical task is impeded by characteristics of typical microbial community profiling technologies, including sparsity (which can be either technical or biological) and the compositionality imposed by most nucleotide sequencing approaches. Many models have been proposed that focus on how the relative abundance of a feature (e.g. taxon or pathway) relates to one or more covariates. Few of these, however, simultaneously control false discovery rates, achieve reasonable power, incorporate complex modeling terms such as random effects, and also permit assessment of prevalence (presence/absence) associations and absolute abundance associations (when appropriate measurements are available, e.g. qPCR or spike-ins). Here, we introduce MaAsLin 3 (Microbiome Multivariable Associations with Linear Models), a modeling framework that simultaneously identifies both abundance and prevalence relationships in microbiome studies with modern, potentially complex designs. MaAsLin 3 also newly accounts for compositionality with experimental (spike-ins and total microbial load estimation) or computational techniques, and it expands the space of biological hypotheses that can be tested with inference for new covariate types. On a variety of synthetic and real datasets, MaAsLin 3 outperformed current state-of-the-art differential abundance methods in testing and inferring associations from compositional data. When applied to the Inflammatory Bowel Disease Multi-omics Database, MaAsLin 3 corroborated many previously reported microbial associations with the inflammatory bowel diseases, but notably 77% of associations were with feature prevalence rather than abundance. In summary, MaAsLin 3 enables researchers to identify microbiome associations with higher accuracy and more specific association types, especially in complex datasets with multiple covariates and repeated measures.
Structural basis of transcription-coupled H3K36 trimethylation by Set2 and RNAPII elongation complex in the nucleosome
Authors: Kujirai Tomoya , Ehara Haruhiko , Ito Tomoko , Henmi Masami , Sekine Shun-ichi , Kurumizaka Hitoshi 
Published: December 2024

AbstractTrimethylation of the histone H3K36 residue (H3K36me3) plays an indispensable role in ensuring transcription fidelity by suppressing undesired cryptic transcription in chromatin. The H3K36me3 modification is accomplished by Set2/SETD2 during transcription elongation by the RNA polymerase II elongation complex (EC). Here we found that the Set2-mediated H3K36me3 deposition occurs primarily on the nucleosome reassembling behind the EC. Cryo-electron microscopy structures of the transcribing EC complexed with Set2 and the reassembled nucleosome revealed that Set2 is anchored by the Spt6 subunit of the EC and captures an H3 N-terminal tail of the nucleosome. Abrogation of the Set2-Spt6 interaction leads to defective transcription-coupled H3K36me3 deposition. These insights elucidate the structure-based mechanism of transcription-coupled H3K36me3 deposition in chromatin.One-Sentence SummaryCryo-EM structures of the EC-Set2-nucleosome complex reveal the mechanism of H3 Lys36 trimethylation in chromatin.
Rapid Sterilization of Clinical Apheresis Blood Products using Ultra-High Dose Rate Radiation
Authors: Melemenidis Stavros , Nguyen Khoa D. , Baraceros-Pineda Rosella , Barclay Cherie K. , Bautista Joanne , Lau Hubert , Ashraf M. Ramish , Manjappa Rakesh , Dutt Suparna , Soto Luis Armando , Katila Nikita , Lau Brianna , Visvanathan Vignesh , Yu Amy S. , Surucu Murat , Skinner Lawrie B. , Engleman Edgar G. , Loo Billy W. , Pham Tho D. 
Published: December 2024

ABSTRACTBACKGROUND AND OBJECTIVESApheresis platelets products and plasma are essential for medical interventions, but both still have inherent risks associated with contamination and viral transmission. Platelet products are vulnerable to bacterial contamination due to storage conditions, while plasma requires extensive screening to minimize virus transmission risks. Here we investigate rapid irradiation to sterilizing doses for bacteria and viruses as an innovative pathogen reduction technology.MATERIALS AND METHODSWe configured a clinical linear accelerator to deliver ultra-high dose rate (6 kGy/min) irradiation to platelet and plasma blood components. Platelet aliquots spiked with 105CFU ofE.coliwere irradiated with 0.1-20 kGy, followed byE.coligrowth and platelet count assays. COVID Convalescent Plasma (CCP) aliquots were irradiated at a virus-sterilizing dose of 25 kGy and subsequently, RBD- specific antibody binding was assessed.RESULTS1 kGy irradiation of bacteria-spiked platelets reducedE.coligrowth by 2.7- log without significant change of platelet count, and 5 kGy or higher produced complete growth suppression. The estimated sterilization (6-log bacterial reduction) dose was 2.3 kGy, corresponding to 31% platelet count reduction. A 25 kGy virus sterilizing dose to CCP produced a 9.2% average drop of RBD-specific IgG binding.CONCLUSIONThis study shows proof-of-concept of a novel rapid blood sterilization technique using a clinical linear accelerator. Promising platelet counts and CCP antibody binding were maintained at bacteria and virus sterilizing doses, respectively. This represents a potential point-of-care blood product sterilization solution. If additional studies corroborate these findings, this may be a practical method for ensuring blood products safety.HIGHLIGHTS1 kGy irradiation of bacteria-spiked platelets reducedE.coligrowth by 2.7-log without significant change of platelet count, and 5 kGy or higher produced complete growth suppression.The estimated sterilization (6-log bacterial reduction) dose was 2.3 kGy, corresponding to 31% platelet count reduction.A 25 kGy virus sterilizing dose to CCP produced a 9.2% average drop of RBD- specific IgG binding.
Salmonellamultimutants enable efficient identification of SPI-2 effector protein function in gut inflammation and systemic colonization
Authors: Newson Joshua P. M. , Gürtler Flavia , Piffaretti Pietro , Meyer Annina , Sintsova Anna , Barthel Manja , Steiger Yves , McHugh Sarah C. , Enz Ursina , Alto Neal M. , Sunagawa Shinichi , Hardt Wolf-Dietrich 
Published: December 2024

AbstractSalmonellaenterica spp. rely on translocation of effector proteins through the SPI-2 encoded type III secretion system (T3SS) to achieve pathogenesis. More than 30 effectors contribute to manipulation of host cells through diverse mechanisms, but interdependency or redundancy between effectors complicates the discovery of effector phenotypes using single mutant strains. Here, we engineer six mutant strains to be deficient in cohorts of SPI-2 effector proteins, as defined by their reported function. Using various animal models of infection, we show that three principle phenotypes define the functional contribution of the SPI-2 T3SS to infection. Multimutant strains deficient for intracellular replication, for manipulation of host cell defences, or for expression of virulence plasmid effectors all showed strong attenuationin vivo,while mutants representing approximately half of the known effector complement showed phenotypes similar to the wild-type parent strain. By additionally removing the SPI-1 T3SS, we find cohorts of effector proteins that contribute to SPI-2 T3SS-driven enhancement of gut inflammation. Further, we provide an example of how iterative mutation can be used to find a minimal number of effector deletions required for attenuation, and thus establish that the SPI-2 effectors SopD2 and GtgE are critical for the promotion of gut inflammation and mucosal pathology. This strategy provides a powerful toolset for simultaneous parallel screening of all known SPI-2 effectors in a single experimental context, and further facilitates the identification of the responsible effectors, and thereby provides an efficient approach to study how individual effectors contribute to disease.