Biology subjects

Guyeux, C.

Publications and source records attributed to Guyeux, C..

6 recordsLinked to original sources

bioRxiv2026

A conserved cysteine-histidine-glutamate metal site identifies DUF501 (Rv1025), an essential uncharacterised protein family of Mycobacterium tuberculosis, as a candidate metalloenzyme and drug target

A substantial fraction of the Mycobacterium tuberculosis proteome remains functionally uncharacterised. Rv1025, a 155-residue protein carrying the domain of unknown function DUF501 (Pfam PF04417), is essential by transposon mutagenesis and vulnerable by CRISPR interference, an attractive but neglected drug target, yet has never been functionally described. The family (4,370 proteins, no Gene Ontology term, no solved structure) is uncharacterised across all organisms and essential in three Actinobacterial genera. A Foldseek search of the AlphaFold model against complete structural databases finds no significant homolog, indicating a novel fold. The operon eno-divIC-Rv1025-ppx2 is conserved across the Actinobacteria phylum, yet AlphaFold-Multimer finds no direct complex between Rv1025 and its neighbour DivIC. Instead, conservation across 8,700 homologous sequences reveals a near-invariant Cys113-His115-Glu59 cluster forming a pocket. Holo AlphaFold3 predictions with Zn, Fe and Mn confidently place a divalent metal on this triad at 2.25-2.47 A; mutating the triad relocates the metal, and an independent backbone-geometry predictor recovers the same site, confirming specificity. The triad is universal across the family: present in all 1,472 near-complete bacterial sequences of the Pfam alignment, with no non-conservative substitution among the 2,228 sequences examined, a defining feature of bacterial DUF501 rather than a mycobacterial peculiarity. We propose that DUF501 is a metal-binding protein and candidate metalloenzyme, the first functional hypothesis for this family, whose conserved, essential metal pocket is a promising drug target. As the predictions build on a conservation-defined site within a fully computational study, they are supportive rather than proof of metal occupancy and warrant experimental validation.

microbiology↗

bioRxiv2026

Rv0810c: a genus-conserved, structurally ordered small protein of unknown function carrying DUF3073 in Mycobacterium tuberculosis

Small annotated open reading frames are the most neglected part of the functionally uncharacterised M. tuberculosis genome. We revisit Rv0810c, a 60-residue protein carrying the unknown-function domain DUF3073 (Pfam PF11273), flagged as an Actinobacteria-signature protein in 2006 but never studied since. Rv0810c is genuinely translated (detected in 11 of 16 M. tuberculosis proteomic datasets), with no significant human homologue, no neighbouring-gene overlap, and no CRISPR-interference polar effect on either flank. Residue-resolved confidence reveals a bipartite architecture: a rigid 33-residue module (pLDDT 91.9) followed by an extended, acidic, intrinsically disordered tail (radius of gyration 22.8 A against 11-12 A expected for a globular protein). The gene is under strong purifying selection (non-synonymous/synonymous ratio 0.86 against 1.93 among 74 size-matched controls, p=5.8x10-8), and its two commonest missense variants are each confined to one sub-lineage, indicating clonal expansion rather than relaxed constraint. DUF3073 is present without a single confirmed loss across 260 well-supported Actinomycetia genera. Despite this conservation, eight independent computational strategies, spanning sequence, structure, electrostatic-patch, embedding-similarity and homo-oligomerisation searches, converge on the same negative: no assignable fold, binding site, or functional neighbour in curated or uncurated sequence space. A phosphosite (Thr24), reproducibly reported by three laboratories, cannot be attributed to a kinase by chemical-genetic or sequence-motif evidence. The contradiction between predicted cytoplasmic topology and macrophage-secretory-fraction detection is narrowed, not resolved: ESX secretion, an immunodominant-epitope confound and host-induced transcription are excluded. Rv0810c exemplifies a class of genuinely uncharacterisable small proteins for which negative reporting, not a manufactured function, is the honest outcome.

microbiology↗

bioRxiv2023

TB-annotator: a scalable web application that allows in-depth analysis of very large sets of publicly available Mycobacterium tuberculosis complex genomes

Tuberculosis continues to be one of the most threatening bacterial diseases in the world. However, we currently have more than 160,000 Short Read Archives (SRAs) of Mycobacterium tuberculosis complex. Such a large amount of data should help to the understanding and the fight against this bacterium. To accomplish this, it would be necessary to thoroughly and comprehensively examine this significant mass of data. This is what TB-Annotator proposes to do, combining a database containing all the diversity of these 160,000 SRAs (at least, SRAs with a reasonable read size and quality), and a fully featured analysis platform to explore and query such a large amount of data. The objective of this article is to present this platform centered on the key notion of exclusivity, to show its numerous capacities (detection of single nucleotide variants, insertion sequences, deletion regions, spoligotyping, etc.) and its general functioning. We will compare TB-Annotator to existing tools for the study of tuberculosis, and show that its objectives are original and have no equivalent at present. The database on which it is based will be presented, with the numerous advanced search queries and screening capacities it offers, and the interest and originality of its phylogenetic tree navigation interface will be detailed. We will end this article with examples of the achievements made possible by the TB-Annotator, followed by avenues for future improvement.

microbiology↗

bioRxiv2022

An updated evolutionary history and taxonomy of Mycobacterium tuberculosis lineage 5, also called M. africanum

Contrarily to other lineages such as L2 and L4, there are still scarce whole-genome-sequence data on L5-L6 MTBC clinical isolates in public genomes repositories. Recent results suggest a high complexity of L5 history in Africa. It is of importance for an adequate assessment of TB infection in Africa, that is still related to the presence of L5-L6 MTBC strains. This study reports a significant improvement of our knowledge of L5 diversity, phylogeographical history, and global population structure of Mycobacterium africanum L5. To achieve this aim, we sequenced new clinical isolates from Northern Nigeria and from proprietary collections, and used a new powerful bioinformatical pipeline, TB-Annotator that explores not only the shared SNPs but also shared missing genes, identical IS6110 insertion sites and shared regions of deletion. This study using both newly sequenced genomes and available public genomes allows to describe new L5 sublineages. We report that the MTBC L5 tree is made-up of at least 12 sublineages from which 6 are new descriptions. We confront our new classification to the most recent published one and suggest new naming for the discovered sublineages. Finally, we discuss the phylogeographical specificity of sublineages 5.1 and sublineage 5.2 and suggest a new hypothesis of L5-L6 emergence in Africa. Impact statementRecent studies on Mycobacterium africanum (L5-L6-L9 of MTBC) genomic diversity and its evolution in Africa discovered three new lineages of the Mycobacterium tuberculosis complex (MTBC) in the last ten years (L7, L8, L9). These discoveries are symptomatic of the delay in characterizing the diversity of the MTBC on the African continent. Another understudied part of MTBC diversity is the intra-lineage diversity of L5 and L6. This study unravels an hidden diversity of L5 in Africa and provides a more exhaustive description of specific genetic features of each sublineage by using a proprietary "TB-Annotator" pipeline. Furthermore, we identify different phylogeographical localization trends between L5.1 and L5.2, suggesting different histories. Our results suggest that a better understanding of the spatiotemporal dynamics of MTBC in Africa absolutely requires a large sampling effort and powerful tools to dig into the retrieved diversity. Data summary[A section describing all supporting external data, software or code, including the DOI(s) and/or accession numbers(s), and the associated URL. If no data was generated or reused in the research, please state this.] The search was done in the TB-Annotator 15901 genomes version which is available at: http://(to be added). The new sequenced genomes are available via NCBI Bioproject accession number: (to be added). The authors confirm all supporting data, code and protocols have been provided within the article or through supplementary data files.

genomics↗

bioRxiv2019

Unexpected diversity of CRISPR unveils some evolutionary patterns of repeated sequences in Mycobacterium tuberculosis

Diversity of the CRISPR locus of Mycobacterium tuberculosis complex has been studied since 1997 for molecular epidemiology purposes. By targeting solely the 43 spacers present in the two first sequenced genomes (H37Rv and BCG), it gave a biased idea of CRISPR diversity and ignored diversity in the neighbouring cas-genes. We set up tailored pipelines to explore the diversity of CRISPR-cas locus in Short Reads. We analyzed data from a representative set of 198 clinical isolates as evidenced by well-characterized SNPs. We found a relatively low diversity in terms of spacers: we recovered only the 68 spacers that had been described in 2000. We found no partial or global inversions in the sequences, letting always the Direct Variant Repeats (DVR) in the same order. In contrast, we found an unexpected diversity in the form of: SNPs in spacers and in Direct Repeats, duplications of various length, and insertions at various locations of the IS6110 insertion sequence, as well as blocks of DVR deletions. The diversity was in part specific to lineages. When reconstructing evolutionary steps of the locus, we found no evidence for SNP reversal. DVR deletions were linked to recombination between IS6110 insertions or between Direct Repeats. This work definitively shows that CRISPR locus of M. tuberculosis did not evolve by classical CRISPR adaptation (incorporation of new spacers) since the last most recent common ancestor of virulent lineages. The evolutionary mechanisms that we discovered could be involved in bacterial adaptation but in a way that remains to be identified.

microbiology↗

bioRxiv2019

Exhaustive reconstruction of the CRISPR locus in Mycobacterium tuberculosis complex using short reads

Spoligotyping, a graphical partial display of the CRISPR locus that can be produced in vitro or in silico, is an important tool for analyzing the diversity of given Mycobacterium tuberculosis complex (MTC) isolates. As other CRISPR loci, this locus is made up of an alternation between direct repeats and spacers, and flanked by cas genes. Unveiling the genetic mechanisms of its evolution requires to have a fairly large amount of fully reconstructed loci among all MTC lineages. In this article, we point out and resolve the problem of CRISPR reconstruction based on short read sequences. We first show that more than 1/3 of the currently assembled genomes available for this complex contain a CRISPR locus erroneously reconstructed, and errors can be very significant. Second, we present a new computational method allowing this locus to be reconstructed extensively and reliably in silico using short read sequencing runs. Third, using this method, we describe new structural characteristics of CRISPR locus by lineages. We show how both the classical experimental in vitro approach and the basic in silico spoligotyping provided by existing analytic tools miss a whole diversity of this locus in MTC, by not capturing duplications, spacer and direct repeats variants, and IS6110 insertion locations. This description is extended in a second article that presents general rules for the evolution of the CRISPR locus in MTC. This work opens new perspectives for a larger exploration of CRISPR loci diversity and of mechanisms involved in its evolution and its functionality.

genomics↗