Biology subjects

Egorov, A. A.

Publications and source records attributed to Egorov, A. A..

6 recordsLinked to original sources

bioRxiv2024

Mechanism of phage sensing and abortion by toxin-antitoxin-chaperone systems

Toxin-antitoxins (TAs) are prokaryotic two-gene systems comprised of a toxin neutralised by an antitoxin. Toxin-antitoxin-chaperone (TAC) systems additionally include a SecB-like chaperone that stabilises the antitoxin by recognising its chaperone addiction (ChAD) element. TACs have been shown to mediate antiphage defence, but the mechanisms of viral sensing and restriction are unexplored. We identify and characterise two Escherichia coli antiphage TAC systems containing HigBA and CmdTA TA units, HigBAC and CmdTAC. The HigBAC is triggered through recognition of the gpV major tail protein of phage {lambda}. Both the ChAD and gpV are recognised by the HigC chaperone through analogous aromatic molecular patterns, explaining the mechanism of activation. We show that the CmdT ADP-ribosyltransferase toxin modifies mRNA to shut down protein synthesis. We establish the modularity of TACs by creating a hybrid broad-spectrum antiphage system combining the CmdTA TA warhead with the HigC chaperone phage sensor. HighlightsE. coli HigBAC and CmdTAC are translation-targeting phage immunity TAC systems HigC chaperone recognises phage {lambda} major tail protein to trigger HigBAC toxicity CmdT ADP-ribosyltransferase toxin abrogates translation through modification of mRNA HigC combined with CmdTA yields hybrid broad-spectrum antiphage defence system

microbiology↗

bioRxiv2024

A Comprehensive Resource for Exploring Antiphage Defense: DefenseFinder Webservice, Wiki and Databases.

In recent years, a vast number of novel antiphage defense mechanisms were uncovered. To facilitate the exploration of mechanistic, ecological, and evolutionary aspects related to antiphage defense systems, we released DefenseFinder in 2021 (Tesson et al., 2022). DefenseFinder is a bioinformatic program designed for the systematic identification of known antiphage defense mechanisms. The initial release of DefenseFinder v1.0.0 included 60 systems. Over the past three years, the number of antiphage systems incorporated into DefenseFinder has grown to 152. The increasing number of known systems makes it a challenge to enter the field and makes the interpretation of detections of antiphage systems difficult. Moreover, the rapid development of sequence-based predictions of structures offers novel possibilities of analysis and should be easily available. To overcome these challenges, we present a hub of resources on defense systems, including: 1) an updated version of DefenseFinder with a web-service search function, 2) a community-curated repository of knowledge on the systems, and 3) precomputed databases, which include annotations done on RefSeq genomes and structure predictions generated by AlphaFold. These pages can be freely accessed for users as a starting point on their journey to better understand a given system. We anticipate that these resources will foster the use of bioinformatics in the study of antiphage systems and will serve the community of researchers who study antiphage systems. This resource is available at: https://defensefinder.mdmlab.fr.

genomics↗

bioRxiv2023

Human tissues exhibit diverse composition of translation machinery

While protein synthesis is vital for the majority of cell types of the human body, diversely differentiated cells require specific translation regulation. This suggests specialization of translation machinery across tissues and organs. Using transcriptomic data from GTEx, FANTOM, and Gene Atlas we systematically explored the abundance of transcripts encoding translation factors and aminoacyl-tRNA synthetases (ARSases) in human tissues. We revised a few known and identified several novel translation-related genes exhibiting strict tissue-specific expression. The proteins they encode include eEF1A1, eEF1A2, PABPC1L, PABPC3, eIF1B, eIF4E1B, eIF4ENIF1, and eIF5AL1. Furthermore, our analysis revealed a pervasive tissue-specific relative abundance of translation machinery components (e.g. PABP and eRF3 paralogs, eIF2B subunits, eIF5MPs, and some ARSases), suggesting presumptive variance in the composition of translation initiation, elongation, and termination complexes. These conclusions were largely confirmed by the analysis of proteomic data. Finally, we paid attention to sexual dimorphism in the repertoire of translation factors encoded in sex chromosomes (eIF1A, eIF2{gamma}, and DDX3), and identified testis and brain as organs with the most diverged expression of translation-associated genes.

molecular biology↗

bioRxiv2022

Genome-encoded ABCF factors implicated in intrinsic antibiotic resistance in Gram-positive bacteria: VmlR2, Ard1 and CplR

Genome-encoded antibiotic resistance (ARE) ATP-binding cassette (ABC) proteins of the F subfamily (ARE-ABCFs) mediate intrinsic resistance in diverse Gram-positive bacteria. The diversity of chromosomally-encoded ARE-ABCFs is far from being fully experimentally explored. Here we characterise phylogenetically diverse genome-encoded ABCFs from Actinomycetia (Ard1 from Streptomyces capreolus, producer of the nucleoside antibiotic A201A), Bacilli (VmlR2 from soil bacterium Neobacillus vireti) and Clostridia (CplR from Clostridium perfringens, Clostridium sporogenes and Clostridioides difficile). We demonstrate that Ard1 is a narrow spectrum ARE-ABCF that specifically mediates self-resistance against nucleoside antibiotics. The single-particle cryo-EM structure of a VmlR2-ribosome complex allows us to rationalise the resistance spectrum of this ARE-ABCF that is equipped with an unusually long antibiotic resistance determinant (ARD) subdomain. We show that CplR contributes to intrinsic pleuromutilin, lincosamide and streptogramin A resistance in Clostridioides, and demonstrate that C. difficile CplR (CDIF630_02847) synergises with the transposon-encoded 23S ribosomal RNA methyltransferase Erm to grant high levels of antibiotic resistance to the C. difficile 630 clinical isolate. Finally, assisted by our novel tool for detection of upstream open reading frames, we dissect the translational attenuation mechanism that controls the induction of cplR expression upon an antibiotic challenge.

microbiology↗

bioRxiv2022

uORF4u: a tool for annotation of conserved upstream open reading frames

SummaryUpstream open reading frames (uORFs, encoding so-called leader peptides) can regulate translation and transcription of downstream main ORFs (mORFs) in prokaryotes and eukaryotes. However, annotation of novel functional uORFs is challenging due their short size of usually less than 100 codons. While transcription- and translation-level next generation sequencing (NGS) methods can be used for genome-wide uORF identification, this data is not available for the vast majority of species with sequenced genomes. At the same time, the exponentially increasing amount of genome assemblies gives us the opportunity to take advantage of evolutionary conservation in our predictions of ORFs. Here we present a tool for conserved uORF annotation in 5' upstream sequences of a user-defined protein of interest or a set of protein homologues. It can also be used to find small ORFs within a set of nucleotide sequences. The output includes publication-quality figures with multiple sequence alignments, sequence logos and locus annotation of the predicted uORFs in graphical vector format. Availability and ImplementationuORF4u is written in Python3 and runs on Linux and MacOS. The command-line interface covers most practical use cases, while the provided Python API allows usage within a Python program and additional customisation. Source code is available from the GitHub page: https://github.com/art-egorov/uorf4u. Detailed documentation that includes an example-driven guide available at the software home page: https://art-egorov.github.io/uorf4u.

bioinformatics↗

bioRxiv2021

A standard knockout procedure alters expression of adjacent loci at the translational level

The S. cerevisiae gene deletion collection is widely used for functional gene annotation and genetic interaction analyses. However, the standard G418-resistance cassette used to produce knockout mutants delivers strong regulatory elements into the target genetic loci. To date, its side effects on the expression of neighboring genes have never been systematically assessed. Here, using ribosome profiling data, RT-qPCR, and reporter expression, we investigated perturbations induced by the KanMX module. Our analysis revealed significant alterations in the transcription efficiency of neighboring genes and, more importantly, severe impairment of their mRNA translation, leading to changes in protein abundance. In the "head-to-head" orientation of the neighbor and the deleted gene, knockout often led to a shift of the transcription start site of the neighboring gene, introducing new uAUG codon(s) into the expanded 5 untranslated region (5 UTR). In the "tail-to-tail" arrangement, knockout led to activation of alternative polyadenylation signals in the neighboring gene, thus altering its 3 UTR. These events may explain the so-called neighboring gene effect (NGE), i.e. false genetic interactions of the deleted genes. We estimate that in as much as [~]1/5 of knockout strains the expression of neighboring genes may be substantially (>2-fold) deregulated at the level of translation.

molecular biology↗