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Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

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bioRxiv2019

Methylation content sensitive enzyme ddRAD (MCSeEd): a reference-free, whole genome profiling system to address cytosine/ adenine methylation changes

Methods for investigating DNA methylation nowadays either require a reference genome and high coverage, or investigate only CG methylation. Moreover, no large-scale analysis can be performed for N6-methyladenosine (6mA). Here we describe the methylation content sensitive enzyme double-digest restriction-site-associated DNA (ddRAD) technique (MCSeEd), a reduced-representation, reference-free, cost-effective approach for characterizing whole genome methylation patterns across different methylation contexts (e.g., CG, CHG, CHH, 6mA). MCSeEd can also detect genetic variations among hundreds of samples. MCSeEd is based on parallel restrictions carried out by combinations of methylation insensitive and sensitive endonucleases, followed by next-generation sequencing. Moreover, we present a robust bioinformatic pipeline (available at https://bitbucket.org/capemaster/mcseed/src/master/) for differential methylation analysis combined with single nucleotide polymorphism calling without or with a reference genome.

genomics↗

bioRxiv2019

Small RNA Mcr11 regulates genes involved in the central metabolism of Mycobacterium tuberculosis and requires 3 sequence along with the transcription factor AbmR for stable expression

Mycobacterium tuberculosis (Mtb), the etiologic agent of tuberculosis, must adapt to host-associated environments during infection by modulating gene expression. Small regulatory RNAs (sRNAs) are key regulators of bacterial gene expression, but their roles in Mtb are not well understood. Here, we address the expression and function of the Mtb sRNA Mcr11, which is associated with slow bacterial growth and latent infections in mice. We found, by using biochemical and genetic approaches, that the AbmR transcription factor and an extended region of native sequence 3 to the mcr11 gene enhance production of mature Mcr11. Additionally, we found that expression of Mcr11 was unstable in the saprophyte Mycobacterium smegmatis, which lacks an mcr11 orthologue. Bioinformatic analyses used to predict regulatory targets of Mcr11 identified 9-11 nucleotide regions immediately upstream of Rv3282 and lipB with potential for direct base-pairing with Mcr11. mcr11-dependent regulation of Rv3282, lipB, Rv2216 and pknA was demonstrated using qRT-PCR in wild type versus mcr11-deleted Mtb and found to be responsive to the presence of fatty acids. These studies establish that Mcr11 has roles in regulating growth and central metabolism in Mtb that warrant further investigation. In addition, our finding that multiple factors are required for production of stable, mature Mcr11 emphasizes the need to study mechanisms of sRNA expression and stability in TB complex mycobacteria to understand their roles in TB pathogenesis.\n\nAuthor SummaryBacterial pathogens must continuously modulate their gene expression in response to changing conditions to successfully infect and survive within their hosts. Transcription factors are well known regulators of gene expression, but there is growing recognition that small RNAs (sRNAs) also have critically important roles in bacterial gene regulation. Many sRNAs have been identified in M. tuberculosis (Mtb), but little is known about their expression, regulatory targets or roles in Mtb biology. In this study, we found that the Mtb sRNA Mcr11, which is expressed at high levels in slowly replicating Mtb and during mouse infection, regulates expression of several target genes involved in central metabolism. Importantly, we also discovered that mcr11 has unexpected requirements for stable expression in mycobacteria. In particular, we identified RNA sequence elements immediately downstream of mcr11 that enhance transcription termination and production of mature Mcr11 RNA in TB-complex mycobacteria. Meanwhile, ectopic expression of Mcr11 was unstable in a non-pathogenic strain of mycobacteria, suggesting that factors specific to pathogenic mycobacteria are required for the stable production of Mcr11. These studies identify sRNA stability as a new frontier for understanding gene expression in Mtb.

microbiology↗

bioRxiv2019

Polyglutamine-expanded androgen receptor disrupts muscle triad, calcium dynamics and the excitation-contraction coupling gene expression program

Spinal and bulbar muscular atrophy (SBMA) is caused by polyglutamine (polyQ) expansions in the androgen receptor (AR) gene. Although clinical and experimental evidence highlight a primary role for skeletal muscle in the onset, progression, and outcome of disease, the pathophysiological and molecular processes underlying SBMA muscle atrophy are poorly understood. Here we show that polyQ-expanded AR alters intrinsic muscle force generation before denervation. Reduced muscle force was associated with a switch in fiber-type composition, disrupted muscle striation, altered calcium (Ca++) dynamics in response to muscle contraction, and aberrant expression of excitation-contraction coupling (ECC) machinery genes in transgenic, knock-in and inducible SBMA mice and patients. Importantly, treatment to suppress polyQ-expanded AR toxicity restored ECC gene expression back to normal. Suppression of AR activation by surgical castration elicited similar ECC gene expression changes in normal mice, suggesting that AR regulates the expression of these genes in physiological conditions. Bioinformatic analysis revealed the presence of androgen-responsive elements on several genes involved in muscle function and homeostasis, and experimental evidence showed AR-dependent regulation of expression and promoter occupancy of the most up-regulated gene from transcriptomic analysis in SBMA muscle, i.e. sarcolipin, a key ECC gene. These observations reveal an unpredicted role for AR in the regulation of expression of genes involved in muscle contraction and Ca++ dynamics, a level of muscle function regulation that is disrupted in SBMA muscle, yet restored by pharmacologic treatment.

neuroscience↗

bioRxiv2019

Origin of a novel CYP20A1 lncRNA through 23 Alu exaptations in the human lineage creates a potential multi-miRNA sponge

BackgroundPrimate-specific Alus contribute to transcriptional novelties in conserved gene regulatory networks. Alu RNAs are present at elevated levels in stress conditions and consequently leads to transcript isoform specific functional role modulating the physiological outcome. One of the possible mechanisms could be Alu nucleated mRNA-miRNA interplay. ResultUsing combination of bioinformatics and experiments, we report a transcript isoform of an orphan gene, CYP20A1 (CYP20A1_Alu-LT) through exaptation of 23 Alus in its 9kb 3UTR. CYP20A1_Alu-LT, confirmed by 3RACE, is an outlier in length and expressed in multiple cell lines. We demonstrate its presence in single nucleus RNA-seq of [~]16000 human cortical neurons (including rosehip neurons). Its expression is restricted to the higher primates. Most strikingly, miRanda predicts [~]4700 miRNA recognition elements (MREs; with threshold< -25kcal/mol) for [~]1000 miRNAs, which have majorly originated within the 3UTR-Alus post exaptation. We hypothesized that differential expression of this transcript could modulate mRNA-miRNA networks and tested it in primary human neurons where CYP20A1_Alu-LT is downregulated during heat shock response and upregulated upon HIV1-Tat treatment. CYP20A1_Alu-LT could possibly function as a miRNA sponge as it exhibits features of a sponge RNA such as cytosolic localization and [≥]10 MREs for 140 miRNAs. Small RNA-seq revealed expression of nine miRNAs that can potentially be sponged by CYP20A1_Alu-LT in neurons. Additionally, CYP20A1_Alu-LT expression was positively correlated (low in heat shock and high in Tat) with 380 differentially expressed genes that contain cognate MREs for these nine miRNAs. This set is enriched in genes involved in neuronal development and hemostasis pathways. ConclusionWe demonstrate a potential role for CYP20A1_Alu-LT as miRNA sponge through preferential presence of MREs within Alus in a transcript isoform specific manner. This highlights a novel component of Alu-miRNA mediated transcriptional modulation leading to physiological homeostasis.

evolutionary biology↗

bioRxiv2019

The diversity of the immunogenic components of the melanoma immunopeptidome.

Antigen-recognition by CD8+ T cells is governed by the pool of peptide antigens presented on the cell surface in the context of HLA class I complexes. Recent studies have shown not only a high degree of plasticity in the immunopeptidome, but also that a considerable fraction of all presented peptides is generated through proteasome-mediated splicing of non-contiguous regions of proteins to form novel peptide antigens. Here we used high-resolution mass-spectrometry combined with new bioinformatic approaches to characterize the immunopeptidome of melanoma cells in the presence or absence of interferon-{gamma}. In total, we identified more than 60,000 peptides from a single patient derived cell line (LM-MEL-44) and demonstrated that interferon-{gamma} induced marked changes in the peptidome with an overlap of only [~]50% between basal and treated cells. Around 6-8% of the peptides were identified as cis-spliced peptides, and 2213 peptides (1827 linear, 386 cis-spliced peptides) were derived from known melanoma-associated antigens. These peptide antigens were equally distributed between the constitutive and interferon-{gamma} induced peptidome. We next examined additional HLA-matched patient derived cell lines to investigate how frequently these peptides were identified and found that a high proportion of both linear and spliced peptides were conserved between individual patient tumors, drawing on data amassing to over 100,000 peptide sequences from these extended data sets. Moreover, several of these peptides showed in vitro immunogenicity across multiple melanoma patients. These observations highlight the breadth and complexity of the repertoire of immunogenic peptides that can be exploited therapeutically and suggest that spliced peptides are a major new class of tumor antigens.

immunology↗

bioRxiv2019

Computational and experimental analysis of the glycophosphatidylinositol-anchored proteome of the human parasitic nematode Brugia malayi

Further characterization of essential systems in the parasitic filarial nematode Brugia malayi is needed to better understand its biology, its interaction with its hosts, and to identify critical components that can be exploited to develop novel treatments. The production of glycophosphatidylinositol-anchored proteins (GPI-APs) is essential in humans, yeast, and the nematode Caenorhabditis elegans. In addition, GPI-APs perform many important roles for cells. In this study, we characterized the B. malayi GPI-anchored proteome using both computational and experimental approaches. We used bioinformatic strategies to show the presence or absence of B. malayi GPI-AP biosynthetic pathway genes and to compile a putative B. malayi GPI-AP proteome using available prediction programs. We verified these in silico analyses using proteomics to identify GPI-AP candidates prepared from the surface of intact worms and from membrane enriched extracts. Our study represents the first description of the GPI-anchored proteome in B. malayi and lays the groundwork for further exploration of this essential protein modification as a target for novel anthelmintic therapeutic strategies.

molecular biology↗

bioRxiv2019

A Novel Chromatin-Opening Element for Stable Long-term Transgene Expression

Long-term stable expression of transgenes in mammalian cells is a challenge in gene therapy, recombinant protein production, and mammalian synthetic biology due to epigenetic silencing and position effect variegation. While multiple classes of regulatory elements have been discovered and proposed to help stabilize expression, the most efficacious has been the Ubiquitous Chromatin Opening Element (UCOE), and in particular, the prototypical A2UCOE from the HNRPA2B1-CBX3 locus. We developed a feature-driven bioinformatics algorithm to discover other putative UCOEs from the human genome, and identified a novel UCOE (SRF-UCOE) that can resist transgene silencing in the methylation-prone P19 cell line. We demonstrate that a 767 bp core sequence of SRF-UCOE is modular to four common mammalian promoters. Notably, SRF-UCOE stabilizes gene expression in transduced P19 cells up to 2.4-fold better over 26 days than the existing A2UCOE by resisting constructs susceptibility to DNA methylation and histone deacetylation. Unlike existing UCOEs, SRF-UCOE lacks inherent transcriptional initiation activity, which can bolster its safe and predictable use in gene therapy constructs. We expect that expanding the set of UCOEs available will expand their utility to novel applications in gene therapy, synthetic biology, and biomanufacturing, as well as contribute to understanding their molecular mechanism.

synthetic biology↗

bioRxiv2019

Strategies for building computing skills to support microbiome analysis: a five-year perspective from the EDAMAME workshop

Here, we report our educational approach and learner evaluations of the first five years of the Explorations in Data Analysis for Metagenomic Advances in Microbial Ecology (EDAMAME) workshop, held annually at Michigan State Universitys Kellogg Biological Station from 2014-2018. We hope this information will be useful for others who want to organize computing-intensive workshops and encourage quantitative skill development among microbiologists.\n\nImportanceHigh-throughput sequencing and related statistical and bioinformatic analyses have become routine in microbiology in the past decade, but there are few formal training opportunities to develop these skills. A week-long workshop can offer sufficient time for novices to become introduced to best computing practices and common workflows in sequence analysis. We report our experiences in executing such a workshop targeted to professional learners (graduate students, post-doctoral scientists, faculty, and research staff).

scientific communication and education↗

bioRxiv2019

Discovery of interaction-related sRNAs and their targets in the Brachypodium distachyon and Magnaporthe oryzae pathosystem

Microbial pathogens secrete small RNA (sRNA) effectors into plant hosts to aid infection by silencing transcripts of immunity and signaling-related genes through RNA interference (RNAi). Similarly, sRNAs from plant hosts have been shown to contribute to plant defense against microbial pathogens by targeting transcripts involved in virulence. This phenomenon is called bidirectional RNA communication or cross kingdom RNAi (ckRNAi). How far this RNAi-mediated mechanism is evolutionarily conserved is the subject of controversial discussions. We examined the bidirectional accumulation of sRNAs in the interaction of the hemibiotrophic rice blast fungus Magnaporthe oryzae (Mo) with the grass model plant Brachypodium distachyon (Bd). By comparative deep sequencing of sRNAs and mRNAs from axenic fungal cultures and infected leaves and roots, we found a wide range of fungal sRNAs that accumulated exclusively in infected tissues. Amongst those, 20-21 nt candidate sRNA effectors were predicted in silico by selecting those Mo reads that had complementary mRNA targets in Bd. Many of those mRNAs predicted to be targeted by Mo sRNAs were differentially expressed, particularly in the necrotrophic infection phase, including gene transcripts involved in plant defense responses and signaling. Vice versa, by applying the same strategy to identify Bd sRNA effectors, we found that Bd produced sRNAs targeting a variety of fungal transcripts, encoding fungal cell wall components, virulence genes and transcription factors. Consistent with function as effectors of these Bd sRNAs, their predicted fungal targets were significantly down-regulated in the infected tissues compared to axenic cultures, and deletion mutants for some of these target genes showed heavily impaired virulence phenotypes. Overall, this study provides the first experimentally-based evidence for bidirectional ckRNAi in a grass-fungal pathosystem, paving the way for further validation of identified sRNA-target duplexes and contributing to the emerging research on naturally occurring cross-kingdom communication and its implications for agriculture on staple crops.\n\nAuthor SummaryIn the present work, we provide first experimental evidence for bidirectional RNA communication in a grass-fungal pathosystem. We deployed the monocotyledonous plant Brachypodium distachyon, which is a genetic model for the staple crops wheat and rice, to investigate the interaction-related sRNAs for their role in RNA communication. By applying a previously published bioinformatics pipeline for the detection of sRNA effectors we identified potential plant targets for fungal sRNAs and vice versa, fungal targets for plant sRNAs. Inspection of the respective targets confirmed their downregulation in infected relative to uninfected tissues and fungal axenic cultures, respectively. By focusing on potential fungal targets, we identified several genes encoding fungal cell wall components, virulence proteins and transcription factors. The deletion of those fungal targets has already been shown to produce disordered virulence phenotypes. Our findings establish the basis for further validation of identified sRNA-mRNA target duplexes and contribute to the emerging research on naturally occurring cross-kingdom communication and its implications for agriculture.

plant biology↗

bioRxiv2019

Regeneration Rosetta: An interactive web application to explore regeneration-associated gene expression and chromatin accessibility

Time-course high-throughput assays of gene expression and enhancer usage in zebrafish provide a valuable characterization of the dynamic mechanisms governing gene regulatory programs during CNS axon regeneration. To facilitate the exploration and functional interpretation of a set of fully-processed data on regeneration-associated temporal transcription networks, we have created an interactive web application called Regeneration Rosetta. Using either built-in or user-provided lists of genes in one of dozens of supported organisms, our web application facilitates the (1) visualization of clustered temporal expression trends; (2) identification of proximal and distal regions of accessible chromatin to expedite downstream motif analysis; and (3) description of enriched functional gene ontology categories. By enabling a straightforward interrogation of these rich data without extensive bioinformatic expertise, Regeneration Rosetta is broadly useful for both a deep investigation of time-dependent regulation during regeneration in zebrafish and hypothesis generation in other organisms.

genomics↗

bioRxiv2019

Peptides derived of kunitz-type serine protease inhibitor as potential vaccine against experimental schistosomiasis

Schistosomiasis is a significant public health problem in sub-Saharan Africa, China, South-East Asia and regions of South and central America affecting about 189 million people. Kunitz-type serine protease inhibitors have been identified as important players in the interaction of other flatworm parasites with their mammalian hosts. Here, we evaluate the protective efficacy of chemically synthesized of T- and B-cell peptide epitopes derived from a kunitz protein from Schistosoma mansoni. Putative kunitz-type protease inhibitor proteins were identified in the S. mansoni genome and their expression analyzed by RNA-seq. Gene expression analyses showed that the kunitz protein Smp_147730 (Syn. Smp_311670) was dramatically and significantly up-regulated in schistosomula and adult worms when compared to the invading cercariae. T- and B-cell epitopes were predicted using bioinformatics tools, chemically synthesized and formulated in the Adjuvant Adaptation (ADAD) vaccination system. BALB/c mice were vaccinated and challenged with S. mansoni cercariae. Kunitz peptides were highly protective in vaccinated BALB/c mice showing significant reductions in recovery of adult females (89-91%), and in the numbers of eggs trapped in the livers (77-81%) and guts (57-77%) of mice. Moreover, liver lesions were significantly reduced in vaccinated mice (64-65%) compared to infected control mice. The vaccination regime was well tolerated with both peptides. We propose the use of these peptides, alone or in combination, as reliable candidates for vaccination against schistosomiasis.

immunology↗

bioRxiv2019

Quantitative proteomics revealed a tight correlation between mitotic phosphorylation/dephosphorylation and structural properties of substrate domains

Protein phosphorylation plays a critical role in the regulation and progression of mitosis. More than 10,000 phosphorylated residues and the associated kinases have been identified to date via proteomic analyses. Although some of these phosphosites are associated with regulation of either protein-protein interactions or the catalytic activity of the substrate protein, the roles of most mitotic phosphosites remain unclear. In this study, we examined structural properties of mitotic phosphosites and neighboring residues to understand the role of heavy phosphorylation in non-structured domains. Quantitative mass spectrometry analysis of mitosis-arrested and non-arrested HeLa cells revealed >4,100 and >2,200 residues either significantly phosphorylated or dephosphorylated, respectively, at mitotic entry. The calculated disorder scores of amino acid sequences of neighboring individual phosphosites revealed that >70% of dephosphorylated phosphosites exist in disordered regions, whereas 50% of phosphorylated sites exist in non-structured domains. A clear inverse correlation was observed between probability of phosphorylation in non-structured domain and increment of phosphorylation in mitosis. These results indicate that at entry to mitosis, a significant number of phosphate groups are removed from non-structured domains and transferred to more-structured domains. Gene ontology term analysis revealed that mitosis-related proteins are heavily phosphorylated, whereas RNA-related proteins are both dephosphorylated and phosphorylated, suggesting that heavy phosphorylation/dephosphorylation in non-structured domains of RNA-binding proteins plays a role in dynamic rearrangement of RNA-containing organelles, as well as other intracellular environments.\n\nSignificance StatementProgression of mitosis is tightly regulated by protein phosphorylation/dephosphorylation. Although proteomic studies have identified tens of thousands of phosphosites in mitotic cells, the roles of them remain to be answered. We approached this question from the viewpoint of the higher-order structure of phosphosites. Quantitative proteomics and bioinformatic analyses revealed that more than 70% of mitotic dephosphorylation events occurred in non-structured regions. Non-structured regions of cellular proteins are attracting considerable attention in terms of their involvement in dynamic rearrangements of intracellular membrane-less organelles and protein assembly/disassembly processes. Our results suggest the possibility that a vast amount of mitosis-associated dephosphorylation/phosphorylation at non-structured regions plays a role in regulating the dynamic assembly/disassembly of intracellular architectures and organelles such as chromosomes and nucleolus.

cell biology↗

bioRxiv2019

Discovery, Expression, Cellular Localization, and Molecular Properties of a Novel, Alternative Spliced HP1g Isoform, Lacking the Chromoshadow Domain

By reading the H3K9Me3 mark through their N-terminal chromodomain (CD), HP1 proteins play a significant role in cancer-associated processes, including cell proliferation, differentiation, chromosomal stability, and DNA repair. Here, we used a combination of bioinformatics-based methodologies, as well as experimentally-derived datasets, that reveal the existence of a novel short HP1{gamma} (CBX3) isoform, named here sHP1{gamma}, generated by alternative splicing of the CBX3 locus. The sHP1{gamma} mRNA encodes a protein composed of 101 residues and lacks the C-terminal chromoshadow domain (CSD) that is required for dimerization and heterodimerization in the previously described 183 a. a HP1{gamma} protein. Fold recognition, order-to-disorder calculations, threading, homology-based molecular modeling, docking, and molecular dynamic simulations show that the sHP1{gamma} is comprised of a CD flanked by intrinsically disordered regions (IDRs) with an IDR-CD-IDR domain organization and likely retains the ability to bind to the H3K9Me3. Both qPCR analyses and mRNA-seq data derived from large-scale studies confirmed that sHP1{gamma} mRNA is expressed in the majority of human tissues at approximately constant ratios with the chromoshadow domain containing isoform. However, sHP1{gamma} mRNA levels appear to be dysregulated in different cancer types. Thus, our data supports the notion that, due to the existence of functionally different isoforms, the regulation of HP1{gamma}-mediated functions is more complex than previously anticipated.

molecular biology↗

bioRxiv2019

Simple rules govern the diversity of bacterial nicotianamine-like metallophores

In metal-scarce environments, some pathogenic bacteria produce opine-type metallophores mainly to face the hosts nutritional immunity. This is the case of staphylopine, pseudopaline and yersinopine, identified in Staphylococcus aureus, Pseudomonas aeruginosa and Yersinia pestis respectively. These metallophores are synthesized by two (CntLM) or three enzymes (CntKLM), CntM catalyzing the last step of biosynthesis using diverse substrates (pyruvate or -ketoglutarate), pathway intermediates (xNA or yNA) and cofactors (NADH or NADPH), depending on the species. Here, we explored substrate specificity of CntM by combining bioinformatics and structural analysis with chemical synthesis and enzymatic studies. We found that NAD(P)H selectivity was mainly due to the amino acid at position 33 (S. aureus numbering) which ensures a preferential binding to NADPH when it is an arginine. Moreover, whereas CntM from P. aeruginosa preferentially uses yNA over xNA, the staphylococcal enzyme is not stereospecific. Most importantly, selectivity towards -ketoacids is largely governed by a single residue at position 150 of CntM (S. aureus numbering): an aspartate at this position ensures selectivity towards pyruvate whereas an alanine leads to the consumption of both pyruvate and -ketoglutarate. Modifying this residue in P. aeruginosa led to a complete reversal of selectivity. Thus, opine-type metallophore diversity is mainly mediated by the absence/presence of a cntK gene encoding a histidine racemase, and the presence of an aspartate/alanine at position 150 of CntM. These two simple rules predict the production of a fourth metallophore by Paenibacillus mucilaginosus, which was confirmed in vitro and called bacillopaline.

biochemistry↗

bioRxiv2019

Knowledge-guided analysis of ‘omics’ data using the KnowEnG cloud platform

We present KnowEnG, a free-to-use computational system for analysis of genomics data sets, designed to accelerate biomedical discovery. It includes tools for popular bioinformatics tasks such as gene prioritization, sample clustering, gene set analysis and expression signature analysis. The system offers knowledge-guided data-mining and machine learning algorithms, where user-provided data are analyzed in light of prior information about genes, aggregated from numerous knowledge-bases and encoded in a massive Knowledge Network. KnowEnG adheres to FAIR principles: its tools are easily portable to diverse computing environments, run on the cloud for scalable and cost-effective execution of compute-intensive and data-intensive algorithms, and are interoperable with other computing platforms. They are made available through multiple access modes including a web-portal, and include specialized visualization modules. We present use cases and re-analysis of published cancer data sets using KnowEnG tools and demonstrate its potential value in democratization of advanced tools for the modern genomics era.

cancer biology↗

bioRxiv2019

Image-based consensus molecular subtype classification (imCMS) of colorectal cancer using deep learning

Image analysis is a cost-effective tool to associate complex features of tissue organisation with molecular and outcome data. Here we predict consensus molecular subtypes (CMS) of colorectal cancer (CRC) from standard H&E sections using deep learning. Domain adversarial training of a neural classification network was performed using 1,553 tissue sections with comprehensive multi- omic data from three independent datasets. Image-based consensus molecular subtyping (imCMS) accurately classified CRC whole-slide images and preoperative biopsies, spatially resolved intratumoural heterogeneity and provided accurate secondary calls with higher discriminatory power than bioinformatic prediction. In all three cohorts imCMS established sensible classification in CMS unclassified samples, reproduced expected correlations with (epi)genomic alterations and effectively stratified patients into prognostic subgroups. Leveraging artificial intelligence for the development of novel biomarkers extracted from histological slides with molecular and biological interpretability has remarkable potential for clinical translation.

cancer biology↗

bioRxiv2019

A putative microcin amplifies Shiga toxin 2a production of Escherichia coli O157:H7

Escherichia coli O157:H7 is a foodborne pathogen, implicated in various multi-state outbreaks. It encodes Shiga toxin on a prophage, and Shiga toxin production is linked to phage induction. An E. coli strain, designated 0.1229, was identified that amplified Stx2a production when co-cultured with E. coli O157:H7 strain PA2. Growth of PA2 in 0.1229 cell-free supernatants had a similar effect, even when supernatants were heated to 100{degrees}C for 10 min, but not after treatment with Proteinase K. The secreted molecule was shown to use TolC for export and the TonB system for import. The genes sufficient for production of this molecule were localized to a 5.2 kb region of a 12.8 kb plasmid. This region was annotated, identifying hypothetical proteins, a predicted ABC transporter, and a cupin superfamily protein. These genes were identified and shown to be functional in two other E. coli strains, and bioinformatic analyses identified related gene clusters in similar and distinct bacterial species. These data collectively suggest E. coli 0.1229 and other E. coli produce a microcin that induces the SOS response in target bacteria. Besides adding to the limited number of microcins known to be produced by E. coli, this study provides an additional mechanism by which stx2a expression is increased in response to the gut microflora.\n\nImportanceHow the gut microflora influences the progression of bacterial infections is only beginning to be understood. Antibiotics are counter-indicated for E. coli O157:H7 infections, and therefore treatment options are limited. An increased understanding of how the gut microflora directs O157:H7 virulence gene expression may lead to additional treatment options. This work identified E. coli that enhance the production of Shiga toxin by O157:H7, through the secretion of a proposed microcin. This work demonstrates another mechanism by which non-O157 E. coli strains may increase Shiga toxin production, and adds to our understanding of microcins, a group of antimicrobials that are less well understood than colicins.

microbiology↗

bioRxiv2019

Identifying gene function and module connections by the integration of multi-species expression compendia

The functions of many eukaryotic genes are still poorly understood. We developed and validated a new method, termed GeneBridge, which is based on two linked approaches to impute gene function and bridge genes with biological processes. First, Gene-Module Association Determination (G-MAD) allows the annotation of gene function. Second, Module-Module Association Determination (M-MAD) allows predicting connectivity among modules. We applied the GeneBridge tools to large-scale multi-species expression compendia--1,700 datasets with over 300,000 samples from human, mouse, rat, fly, worm, and yeast--collected in this study. Unlike most existing bioinformatics tools, GeneBridge exploits both positive and negative gene/module-module associations. We constructed association networks, such as those bridging mitochondria and proteasome, mitochondria and histone demethylation, as well as ribosomes and lipid biosynthesis. The GeneBridge tools together with the expression compendia are available at systems-genetics.org, to facilitate the identification of connections linking genes, modules, phenotypes, and diseases.

systems biology↗