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bioRxiv2018

Identifying the Biosynthetic Gene Cluster for Triacsins with an N-hydroxytriazene Moiety

Triacsins are a family of natural products containing an N-hydroxytriazene moiety not found in any other known secondary metabolites. Though many studies have examined the biological activity of triacsins in lipid metabolism, the biosynthesis of triacsins has remained unknown. Here, we report the identification of the triacsin biosynthetic gene cluster in Streptomyces aureofaciens ATCC 31442. Bioinformatic analysis of the gene cluster led to the discovery of the tacrolimus producer Streptomyces tsukubaensis NRRL 18488 as a new triacsin producer. In addition to targeted gene disruption to identify necessary genes for triacsin production, stable isotope feeding was performed in vivo to advance the understanding of N-hydroxytriazene biosynthesis.

biochemistry↗

bioRxiv2018

Phase transition and amyloid formation by a viral protein as an additional molecular mechanism of virus-induced cell toxicity

Henipaviruses are severe human pathogens responsible for severe encephalitis. Their V protein is a key player in the evasion of the host innate immune response. We have previously reported a biophysical characterization of the Henipavirus V proteins and shown that they interact with DDB1, a cellular protein that is a component of the ubiquitin ligase E3 complex. Here, we serendipitously discovered that the Hendra virus V protein undergoes a liquidhydrogel phase transition. By combining experimental and bioinformatics approaches, we have identified the V region responsible for this phenomenon. This region (referred to as PNT3), which falls within the long intrinsically disordered region of V, was further investigated using a combination of biophysical and structural approaches. ThioflavinT and Congo red binding assays, together with negative-staining electron microscopy studies, show that this region forms amyloid-like, {beta}-enriched structures. Such structures are also formed in mammal cells transfected to express PNT3. Those cells also exhibit a reduced viability in the presence of a stress agent. Interestingly, mammal cells expressing a rationally designed, non-amyloidogenic PNT3 variant (PNT33A), appear to be much less sensitive to the stress agent, thus enabling the establishment of a link between fibril formation and cell toxicity. The present findings therefore pinpoint a so far never reported possible mechanism of virus-induced cell toxicity.

biophysics↗

bioRxiv2018

Id proteins promote a cancer stem cell phenotype in triple negative breast cancer via Robo1-dependent c-Myc activation

Breast cancers display phenotypic and functional heterogeneity and several lines of evidence support the existence of cancer stem cells (CSCs) in certain breast cancers, a minor population of cells capable of tumor initiation and metastatic dissemination. Identifying factors that regulate the CSC phenotype is therefore important for developing strategies to treat metastatic disease. The Inhibitor of Differentiation Protein 1 (Id1) and its closely related family member Inhibitor of Differentiation 3 (Id3) (collectively termed Id) are expressed by a diversity of stem cells and are required for metastatic dissemination in experimental models of breast cancer. In this study, we show that ID1 is expressed in rare neoplastic cells within ER-negative breast cancers. To address the function of Id1 expressing cells within tumors, we developed two independent murine models of Triple Negative Breast Cancer (TNBC) in which a genetic reporter permitted the prospective isolation of Id1+ cells. Id1+ cells are enriched for self-renewal in tumorsphere assays in vitro and for tumor initiation in vivo. Conversely, depletion of Id1 and Id3 in the 4T1 murine model of TNBC demonstrates that Id1/3 are required for cell proliferation and self-renewal in vitro, as well as primary tumor growth and metastatic colonization of the lung in vivo. Using combined bioinformatic analysis, we have defined a novel mechanism of Id protein function via negative regulation of the Roundabout Axon Guidance Receptor Homolog 1 (Robo1) leading to activation of a Myc transcriptional programme.

cancer biology↗

bioRxiv2018

Detection of a novel mutation G511T in the 530 loop in 16S rRNA in multi drugs resistant Mycobacterium tuberculosis isolated from Sudanese patients

BackgroundTuberculosis (TB) is a bacterial disease considered as a global public health emergency by the World Health Organization (WHO) since 1993. In Sudan, MDR-TB represents a growing threat and one of the most important challenges that faced national tuberculosis program to establish a comprehensive multidrug-resistant tuberculosis management system.\n\nObjectiveTo characterize the diversity and frequency of mutations in Sudanese MDR-TB strains isolated from Wad Madani, Al-Gadarif and Khartoum using 16S rRNA and phylogeny approach.\n\nMaterial and MethodsA total of 60 MDR-TB isolates from Wad-Madani, Al-Gadarif and Khartoum were tested with molecular LPA (Genotype MTBDR plus) and GeneXpert MTB/RIF assay and Spoligotyping to confirm their resistance to RIF and INH. Sequencing and phylogenetic analysis was carried out using in silico tools.\n\nResultThis study revealed the circulation of different Sudanese MDR-TB strains isolated from Wad Madani and Al-Gadarif belonging to two distinct common ancestors. Two isolates from Wad Madani (isolate3 and isolate11) found in one main group which characterized by a novel mutation G511T in the 530 loop.\n\nConclusionThe recurrence of C217A mutation in Wad Madani (isolate11) indicates the spread of this mutation in Sudanese MDR-TB strains and the diversity of this inheritance leading to generate new G511T novel mutation. So, understanding the molecular characterization of resistance mechanisms in MD-TB can facilitate the early detection of resistance, the choice of appropriate treatment and ultimately the management of MD-TB transmission. Bioinformatics approaches provide helpful tools for analyzing molecular mechanisms of resistance in pathogens.

microbiology↗

bioRxiv2018

Accessory genome contributes to the virulence and resistance of the ocular isolate of Pseudomonas aeruginosa: A complete genome analysis

Bacteria can acquire an accessory genome through the horizontal transfer of genetic elements from non-parental lineages. This leads to rapid genetic evolution allowing traits such as antibiotic resistance and virulence to spread through bacterial communities. The study of complete genomes of bacterial strains helps to understand the genomic traits associated with virulence and antibiotic resistance. We aimed to investigate the complete accessory genome of an ocular isolate of P. aeruginosa. We obtained the complete genome of the ocular isolate strain PA34 of P. aeruginosa utilising genome sequence reads from Illumina and Oxford Nanopore Technology followed by PCR to close any identified gaps. In-depth genomic analysis was performed using various bioinformatics tools. The phenotypic properties of susceptibility to heavy metals and cytotoxicity were determined to confirm expression of certain traits. The complete genome of PA34 includes a chromosome of 6.8 Mbp and two plasmids of 95.4 Kbp (pMKPA34-1) and 26.8 Kbp (pMKPA34-2). PA34 had a large accessory genome of 1,213 genes and had 543 unique genes not present in other strains. These exclusive genes encoded features related to metal and antibiotic resistance, phage integrase and transposons. At least 24 GIs were predicated in the complete chromosome, of which two were integrated into novel sites. Eleven GIs carried virulence factors or replaced pathogenic genes. A bacteriophage carried the aminoglycoside resistance gene (aac(3)-IId). The two plasmids carried other six antibiotic resistance genes. The large accessory genome of this ocular isolate plays a large role in shaping its virulence and antibiotic resistance.

microbiology↗

bioRxiv2018

Transcriptomic responses to thermal stress and varied phosphorus conditions in Symbiodinium kawagutii

Symbiodinium species are essential symbionts of tropical reef-building corals and the disruption of their symbiosis with corals as a consequence of seawater warming and other stress conditions leads to the globally widespread coral bleaching. As coral reefs live in the oligotrophic environment, Symbiodinium photosynthesis can also face nutrient stress. How metabolic pathways in Symbiodinium respond to thermal stress and phosphate depletion is poorly understood and underexplored for many species. Here we conducted RNA-seq analysis to investigate transcriptomic responses to thermal stress, phosphate deprivation and glycerol-3-phosphate (Gro3P) replacement in S. kawagutii. RNA-seq and bioinformatic analysis were conducted for the above-mentioned three treatments and a control. We identified 221 (2.04%) genes showing no significant differential expression among all conditions, and defined them as \"core\" genes of S. kawagutii, which mostly were in the Gene Ontology terms of catalytic activity and binding. Using algorithms edgeR and NOIseq in combination, we identified a set of differentially expressed genes (DEGs) for each treatment relative to the control. Under heat stress 357 (4.42%) DEGs were found, with predicted roles in active molecular (protein-protein/RNA/DNA) interaction, cell wall modulation and transport (including nutrients, iron, and oxygen). About as many DEGs (396, 4.73%) were identified under P deprivation while nearly double of that (671, 8.05%) were detected under Gro3P utilization; in both cases most of the DEGs were up-regulated and predicted to function in photosystem and defensome. Further KEGG pathway comparison revealed different molecular responses between phosphate deprivation and Gro3P utilization. Catalytic activity and binding seem to be two important core functions in S. kawagutii. The most significant transcriptional response in S. kawagutii to heat stress was regulation of molecular interaction, cell wall modulation, and transport of iron, oxygen, and major nutrients, suggesting that this species uses a unique mechanism to cope with heat stress, possibly conferring thermal tolerance. The greatest transcriptomic impact of phosphate deprivation and Gro3P replacement were the up-regulation of photosystem and defense. This study provides new clues about molecular mechanisms underpinning responses in Symbiodinium to temperature and nutrient stresses, which will generate new hypotheses and set a new framework for future investigations.

microbiology↗

bioRxiv2018

Functional evaluation of transposable elements as transcriptional enhancers in mouse embryonic and trophoblast stem cells

The recurrent invasion and expansion of transposable elements (TEs) throughout evolution brought with it a vast array of coding and non-coding sequences that can serve as substrates for natural selection. Namely, TEs are thought to have contributed to the establishment of gene regulatory networks via their cis-acting elements. Both the embryonic and extraembryonic lineages of the early mouse embryo are thought to have benefited from the co-option of TEs as distal enhancer elements. However, there is little to no evidence that these particular TEs play significant roles in the regulation of gene expression. Here we tested for roles of TEs as enhancers in mouse embryonic and trophoblast stem cells by combining bioinformatic analyses with genetic and epigenetic editing experiments. Epigenomic and transcriptomic data from wildtype cells suggested that a large number of TEs played a role in the establishment of highly tissue-specific gene expression programmes. Through genetic editing of individual TEs we confirmed a subset of these regulatory relationships. However, a wider survey via CRISPR interference of RLTR13D6 elements in embryonic stem cells revealed that only a minority play significant roles in gene regulation. Our results suggest that a small proportion of TEs contribute to the mouse pluripotency regulatory network, and highlight the importance of functional experiments when evaluating the role of TEs in gene regulation.

genetics↗

bioRxiv2018

Profiling gene expression of the host response to a Plasmodium vivax irradiated sporozoite immunization and infectious challenge

The development of vaccines that provide sterile protection against human malaria is a major global public health priority requiring a better understanding of the mechanisms involved in natural and vaccine-induced sterile immunity. RNAseq was used to profile gene expression of peripheral blood samples from 12 Duffy positive (Fy+) (Plasmodium vivax susceptible) volunteers enrolled in a phase 2 trial who were vaccinated with radiation attenuated P. vivax sporozoites (RAS; 5 were protected, 7 not) and from 5 Fy- (P. vivax resistant) volunteers exposed to mosquitoes harboring live non-attenuated sporozoites. Blood samples were obtained before and after immunization as well as after controlled infection with live P vivax sporozoites. The most profound changes in gene expression were observed between baseline and post-challenge, with 97 distinct signatures differentiating protected and not protected Fy+ individuals. Differentiation was also observed between Fy- and Fy+ protected individuals, notably with downregulation of multiple inflammatory responses as well as extracellular matrix-related gene activity. Analysis of transcriptional modules shows that both B-cell and T-cell signaling are reduced while cell cycle regulation, interferon response, and other informative signatures are elevated in individuals who are not protected against malaria. An asymptomatic individual had an intermediate profile indicative of differential transcription associated with pathology and symptomology. Systems biology thus provides insight into how whole malaria-attenuated sporozoites prime the immune system to protect against malaria, as well as the transcription responses that are associated with sterile protection.\n\nAUTHOR SUMMARYTranscriptomic analysis has shed considerable light on the molecular mechanisms of vaccine efficacy against a variety of viral pathogens, but has not yet been used in relation to parasite vaccinology. Here we describe whole peripheral blood gene expression profiling to contrast the contributions of myeloid and lymphoid immune responses to vaccine-induced and natural protection against Plasmodium vivax- mediated malaria. Two modes of modular bioinformatic analyses are applied to RNAseq data from pre- and post-vaccination samples from 19 volunteer participants in a clinical trial investigating the efficacy of an irradiated sporozoite vaccine. During active infection, both B- and T-cell activity are reduced in individuals who are not protected by vaccination relative to those who are protected. Lack of the Fy antigen prevents entry of parasite into red blood cells, thereby providing natural clinical immunity, but we also document differences in T-cell and neutrophil activity in these individuals relative to those protected by irradiation of the parasite. One individual who was asymptomatic for malaria but had blood parasite despite vaccination showed an intermediate response indicating how systems biology approaches can distinguish gene expression associated with pathology and infection.

genomics↗

bioRxiv2018

Low-Density Lipoprotein Receptor-Mediated Lipidome-Transcriptome Reprogramming Impulses to Cisplatin Insensitivity

Platinum-based therapy remains the cornerstone for cancer patient management; however, its efficacy varies. Theis study demonstrated the differential expressions of low-density lipoprotein receptor (LDLR) in subtypes of epithelial ovarian carcinoma (EOC) determines cisplatin sensitivity. Its sensitive in serous EOCs (low LDLR), where insensitive in endometrioid and clear cell EOCs (high LDLR). Meanwhile, knocked-down or overexpressed LDLR in EOC could reversed the chemosensitivity pattern both in vitro and in vivo. Mechanistic dissection with transcriptome vs. lipidome trans-omics analyses elucidated the LDLR[->]LPC (Lyso-PhosphotidylCholine)[->]FAM83B (phospholipase-related)[->]FGFRs (cisplatin sensitivity and phospholipase-related) regulatory axis in cisplatin insensitivity. Implementing LPC-liposome encapsulated cisplatin could facilitate DNA-adduct formation via lipid droplets (LDs) delivery. Furthermore, Bioinformatics analyses found that the LDL/R[->]LD homeostasis alteration is critical for therapeutic prognosis. Lastly, using LPC-liposome-cisplatin improved cisplatin sensitivities in gastric cancer, renal cell carcinoma, hepatocellular carcinoma, cholangiocarcinoma, and pancreatic adenocarcinoma cells. In conclusion, this report discovered a LDL/R-reprogrammed transcriptome-lipidome network, by which impulses platinum insensitivity and disease outcome. The drug specific lipidome for liposome manufacture might be an efficienct pharmaceutics strategy for chemoagents.\n\nSignificanceLDLR reprograms cellular lipidome and transcriptome profiles to determines chemotherapy therapeutic efficacy. The LDLR-reduced LPC abundance disturbs phospholipids homeostasis of Lands cycle in LD, by which attenuates intracellular platinum transportation for DNA-adduct formation. Targeting LDLR-LD-lipidome with LPC-liposome-platinum could boost therapeutic efficacy for insensitivity.

cancer biology↗

bioRxiv2018

VIKNGS: A C++ VARIANT INTEGRATION KIT FOR NEXTGENERATION SEQUENCING ASSOCIATION ANALYSIS

MotivationIntegration of next generation sequencing data (NGS) across different research studies can improve the power of genetic association testing by increasing sample size and can obviate the need for sequencing controls. Unfortunately, if differential genotype uncertainty across studies is not accounted for, combining data sets can also produce spurious association results. The robust variance score statistic (RVS) for genetic association of rare and common variants has been shown to effectively adjust for bias caused by the differences in read depth in case-control genetic association studies when the two groups were sequenced using different experimental designs. To enable consortium research, the aggregation of several data sets for genetic association analysis of quantitative and binary traits with covariate adjustment is required, and we developed the Variant Integration Kit for NGS (VikNGS) that expands the functionality of RVS (vRVS) for this purpose.\n\nResultsVikNGS is a fast and computationally efficient cross-platform software package that provides an implementation for vRVS, as well as conventional rare and common variant genotype-based association analysis approaches. The package includes a graphical user interface that contains power simulation functionality and data visualization tools.\n\nAvailability and ImplementationThe VikNGS package can be downloaded at http://www.tcag.ca/tools/index.html\n\nDocumentation can be found at https://VikNGSdocs.readthedocs.io/en/latest/\n\nContactlisa.strug@sickkids.ca\n\nSupplementary informationSupplementary data are available at Bioinformatics online.

genetics↗

bioRxiv2018

The Salivary Microbiome: Analysis of by Pyrosequencing and the Relationship with Helicobacter pylori Infection

AbstractsO_ST_ABSBackgroundsC_ST_ABSThere have been reports of Helicobacter pylori (H. pylori) in the oral cavity and it has been suggested that the oral cavity may be a reservoir for H. pylori reflux from the stomach.\n\nObjectivesHigh-throughput pyrosequencing was used to assess the structure and composition of oral microbiota communities in individuals with or without confirmed H. pylori infection.\n\nMethodsSaliva samples were obtained from 34 H. pylori infected and 24 H. pylori uninfected subjects. Bacterial genomic DNA was extracted and examined by pyrosequencing by amplification of the 16S rDNA V3-V4 hypervariable regions followed by bioinformatics analysis. Saliva sampling was repeated from 22 of the 34 H. pylori infected subjects 2 months after H. pylori eradication.\n\nResultsHigh-quality sequences (2,812,659) clustered into 95,812 operational taxonomic units (OTUs; 97% identity), representing 440 independent species belonging to 138 genera, 68 families, 36 orders, 21 classes, and 11 phyla. Species richness (alpha diversity) of H. pylori infected subjects was similar to that of uninfected subjects. Eradication treatment decreased saliva bacterial diversity. Beta diversity analysis showed that the salivary microbial community structure differed between H. pylori infected and uninfected subjects both before and after H. pylori eradication.\n\nConclusionsSalivary microbiota diversity was similar in H. pylori infected and uninfected individuals. Antibiotic therapy was associated with a decline in salivary bacterial diversity. Both H. pylori infection and its eradication caused the oral microbiota alterations in community and structure. The present of H. pylori in oral cavity was not related with its infection status in stomach.\n\nTrial registrationClinicalTrials.gov, Identifier: NCT03730766\n\nImportanceThe oral cavity plays a vital role in Helicobacter pylori transmission among human. High-throughput pyrosequencing of the 16S rDNA V3-V4 hypervariable regions was used to assess the structure and composition of oral microbiota communities in individuals with or without confirmed Helicobacter pylori infection. We show that both Helicobacter pylori infection and eradication cause microbiota alterations in the oral microbiota. Prior studies report detection of Helicobacter pylori in the oral cavity by polymerase chain reaction. We show that the presence of Helicobacter pylori in the oral cavity is unrelated with its infection status in the stomach.

microbiology↗

bioRxiv2018

The Genomic Landscape of Centromeres in Various Cancers

Centromere genomics remain poorly characterized in cancer, due to technologic limitations in sequencing and bioinformatics methodologies that make high-resolution delineation of centromeric loci difficult to achieve. We here leverage a highly specific and targeted rapid PCR methodology to quantitatively assess the genomic landscape of centromeres in cancer cell lines and primary tissue. PCR-based profiling of centromeres revealed widespread heterogeneity of centromeric and pericentromeric sequences in cancer cells and tissue as compared to healthy counterparts. Quantitative reductions in select centromeric core and pericentromeric markers were observed in neoplastic samples as compared to healthy counterparts. Subsequent phylogenetic analysis of a pericentromeric endogenous retrovirus amplified by PCR revealed possible gene conversion events occurring at numerous pericentromeric loci in the setting of malignancy. Our findings collectively represent the first look into centromere genetics in the setting of malignancy, providing valuable insight into the evolution and reshuffling of centromeric sequences in cancer development and progression.

genomics↗

bioRxiv2019

Bacillus clarus sp. nov. is a new Bacillus cereus group species isolated from soil

Bacillus cereus group or B. cereus sensu lato (s.l.), is comprised of Gram-positive spore-forming, rod-like bacteria that are widespread in natural environments. Although the species in this group are known to be highly related in terms of phenotypic characteristics, they display different levels of pathogenicity. Biochemical assays are therefore considered to be insufficient for accurate taxonomic classification of B. cereus group species. To facilitate accurate taxonomic classification and associated prediction of pathogenic potential, we have conducted comparative genomic analyses of publicly available genome assemblies of B. cereus group isolates. Through that, we found that an isolate previously known as B. mycoides ATCC 21929 was sufficiently distant from valid and effective type strains to be considered a putative new species. We have conducted biochemical and bioinformatic characterization of strain ATCC 21929 that had been isolated from soil in Papua New Guinea. Strain ATCC 21929 most closely resembles B. paramycoides NH24A2T, producing ANIb and DDH values of 86.70% and 34.1%, respectively. Phenotypically, isolate ATCC 21929 does not possess cytochrome c oxidase activity, and is able to grow at a range of temperatures 15{degrees}C - 43{degrees}C and at a range of pH 6 - 9. With regards to fatty acid composition, this isolate has iso-C17:0 in highest abundance. We propose the strain ATCC 21929T (=PS00077AT = PS00077BT = PSU-0922T = BHPT) as a new species named Bacillus clarus sp. nov. to facilitate accurate taxonomic classification of B. cereus group isolates.

microbiology↗

bioRxiv2019

Organism-wide single-cell transcriptomics of long-lived C. elegans daf-2 mutants reveals tissue-specific reprogramming of gene expression networks

A critical requirement for a systems-level understanding of complex biological processes such as aging is the ability to directly characterize interactions between cells and tissues within a multicellular organism. C. elegans nematodes harboring mutations in the insulin-like receptor daf-2 exhibit dramatically-increased lifespans. To identify tissue-specific biochemical mechanisms regulating aging plasticity, we single-cell sequenced 3-mRNA libraries generated from seven populations of whole day-one adult wild-type and daf-2-/- worms using the 10x ChromiumV1platform. The age-synchronized samples were bioinformatically merged into a single aligned dataset containing 40,000 age-synchronized wild-type and daf-2-/- cellular transcriptomes partitioned into 101 clusters, using unsupervised machine-learning algorithms to identify common cell types. Here we describe the basic features of the adult C. elegans single-cell transcriptome and summarize functional alterations observed in the gene expression profiles of long-lived daf-2-/- worms. Comprehensive methods and datasets are provided. This is the first study to directly quantify cell-specific differential gene expression between two age-synchronized, genetically-distinct populations of multicellular organisms. This novel approach answers fundamental questions regarding tissue-specific regulation of gene expression and helps to establish a foundation for a comprehensive C. elegans single-cell gene expression atlas.

cell biology↗

bioRxiv2019

Identifying small proteins by ribosome profiling with stalled initiation complexes

Small proteins consisting of 50 or fewer amino acids have been identified as regulators of larger proteins in bacteria and eukaryotes. Despite the importance of these molecules, the true prevalence of small proteins remains unknown because conventional annotation pipelines usually exclude small open reading frames (smORFs). We previously identified several dozen small proteins in the model organism Escherichia coli using theoretical bioinformatic approaches based on sequence conservation and matches to canonical ribosome binding sites. Here, we present an empirical approach for discovering new proteins, taking advantage of recent advances in ribosome profiling in which antibiotics are used to trap newly-initiated 70S ribosomes at start codons. This approach led to the identification of many novel initiation sites in intergenic regions in E. coli. We tagged 41 smORFs on the chromosome and detected protein synthesis for all but three. The corresponding genes are not only intergenic, but are also found antisense to other genes, in operons, and overlapping other open reading frames (ORFs), some impacting the translation of larger downstream genes. These results demonstrate the utility of this method for identifying new genes, regardless of their genomic context.\n\nIMPORTANCEProteins comprised of 50 or fewer amino acids have been shown to interact with and modulate the function of larger proteins in a range of organisms. Despite the possible importance of small proteins, the true prevalence and capabilities of these regulators remain unknown as the small size of the proteins places serious limitations on their identification, purification and characterization. Here, we present a ribosome profiling approach with stalled initiation complexes that led to the identification of 38 new small proteins.

microbiology↗

bioRxiv2019

Phage-encoded cationic antimicrobial peptide used for outer membrane disruption in lysis

Spanins are required for the last step in bacteriophage lysis: the disruption of the outer membrane. Bioinformatic analysis has shown that ~15% of phages lack a spanin gene, which suggests an alternate mechanism of outer membrane disruption. To address this, we selected virulent podophage {phi}KT as a spaninless exemplar and tested {phi}KT genes for outer membrane disruption during lysis. Hypothetical novel gene 28 causes outer membrane disruption when co-expressed with {phi}KT lysis genes and complements the lysis defect of a {lambda} spanin mutant. Gp28 is a 56 aa cationic peptide with predicted amphipathic helical structure and is associated with the particulate fraction after lysis. Urea and KCl washes did not release gp28 from the particulate, suggesting a strong hydrophobic interaction with the membrane. Super high-resolution microscopy supports a primarily outer membrane localization for the peptide. Additionally, holin function is not required for gp28-mediated lysis. Gp28 is similar in size, charge, predicted fold, and membrane association to the human cathelicidin antimicrobial peptide LL-37. In standard assays to measure bactericidal and inhibitory effects of antimicrobial peptides on bacterial cells, synthesized gp28 performed equivalently to LL-37. The studies presented here suggest that {phi}KT Gp28 disrupts bacterial outer membranes during lysis in a manner akin to antimicrobial peptides.\n\nSignificanceHere we provide evidence that {phi}KT produces an antimicrobial peptide for outer membrane disruption during lysis. The disruptin is a new paradigm for phage lysis, and has no similarities to other known lysis genes. Many mechanisms have been proposed for the function of antimicrobial peptides, however there is not a consensus on the molecular basis of membrane disruption. Additionally, there is no established genetic selection system to support such studies. Therefore, the {phi}KT disruptin may represent the first genetically tractable antimicrobial peptide.

microbiology↗

bioRxiv2019

Environmental and epigenetic regulation of Rider retrotransposons in tomato

Transposable elements in crop plants are the powerful drivers of phenotypic variation that has been selected during domestication and breeding programs. In tomato, transpositions of the LTR (long terminal repeat) retrotransposon family Rider have contributed to various phenotypes of agronomical interest, such as fruit shape and colour. However, the mechanisms regulating Rider activity are largely unknown. We have developed a bioinformatics pipeline for the functional annotation of retrotransposons containing LTRs and defined all full-length Rider elements in the tomato genome. Subsequently, we showed that accumulation of Rider transcripts and transposition intermediates in the form of extrachromosomal DNA is triggered by drought stress and relies on abscisic acid signalling. We provide evidence that residual activity of Rider is controlled by epigenetic mechanisms involving siRNAs and the RNA-dependent DNA methylation pathway. Finally, we demonstrate the broad distribution of Rider-like elements in other plant species, including crops. Thus our work identifies Rider as an environment-responsive element and a potential source of genetic and epigenetic variation in plants.

plant biology↗

bioRxiv2019

Phylogenomics reveals major diversification rate shifts in the evolution of silk moths and relatives

The silkmoths and their relatives (Bombycoidea) are an ecologically and taxonomically diverse superfamily that includes some of the most charismatic species of all the Lepidoptera. Despite displaying some of the most spectacular forms and ecological traits among insects, relatively little attention has been given to understanding their evolution and the drivers of their diversity. We heavily sampled (both in taxa and loci) all major lineages of the Bombycoidea, producing a well-supported phylogeny that identified important evolutionary patterns (e.g., morphology, biogeography, and differences in speciation and extinction). Importantly, analysis of diversification rates highlights the stark increases that exist within the Sphingidae (hawkmoths) and Saturniidae (wild silkmoths). We postulate that these rate shifts are due to differences in the intense selective pressures from insectivorous bats. The study also introduces a new Bombycoidea-specific Anchored Hybrid Enrichment (AHE) probe set, a modified DNA extraction protocol for Lepidoptera specimens from natural history collections, and additional information on the existing AHE bioinformatics pipeline. Our research highlights the flexibility of AHE to generate genomic data from a wide range of museum specimens, both age and preservation method, and will allow researchers to tap into the wealth of biological data residing in natural history collections around the globe.

evolutionary biology↗