Biology subjects

Descorps-Declere, S.

Publications and source records attributed to Descorps-Declere, S..

8 recordsLinked to original sources

bioRxiv2025

The E. coli CNF1 toxin induces fetal reprogramming of intestinal stem cells and a serrated colorectal cancer transcriptional signature in organoids

Mechanistic insights are essential to understand how gut microbiota pathobionts contribute to tumorigenesis, given the mounting evidence linking them to colorectal cancer. We report a higher prevalence of the Rho GTPases-targeting cnf1 toxin gene from Escherichia coli in the microbiota of early-stage, proximal colorectal cancer. Comparative gene set enrichment analysis reveals a concordant serrated pathway signature between colorectal cancer tissue of patients colonized with cnf1+ bacteria and CNF1-treated mouse intestinal organoids. RNA sequencing of organoids shows that CNF1 induces a fetal-like transcriptional reprogramming. Using integrated approaches, we demonstrate that CNF1 reprograms Lgr5 stem cells into a Ly6a/Sca-1 fetal-like state, that exhibits enhanced stemness potential. This reprogramming is preceded by a Yap/Taz-driven early transcriptional program and nuclear translocation of Yap. Functional analyses identify a RhoA/Rock-Yap/Taz-dependent transition to Ly6a/Sca-1 stem cells, highlighting a mechanistic link between bacterial effectors and stem cell plasticity in colorectal tumorigenesis.

cancer biology↗

bioRxiv2023

TALEN-induced contraction of CTG trinucleotide repeats in myotonic dystrophy type 1 cells

Trinucleotide repeat expansions are the cause of two dozen neurodegenerative and developmental disorders. One of these, myotonic dystrophy type 1 (Steinert disease, or DM1) is due to the expansion of a CTG triplet in the 3 UTR of the DMPK gene. We used highly specific DNA endonucleases to induce a double-strand break in the repeat tract to contract it below pathological length. Expression of a TALE Nuclease (TALEN) in human DM1 cells induced moderate CTG repeat contractions in 27% of the clones analyzed. These clones exhibited large internal deletions within the TALEN, occurring by homologous recombination between internal TALE repeats, inactivating the nuclease, and explaining its reduced efficacy. Taking advantage of the degeneracy of the genetic code, we recoded the TALEN sequence, to decrease internal redundancy and optimize codon usage. The new recoded TALEN showed increased efficacy in DM1 cells, with 68% of clones exhibiting a moderate to large contraction of the CTG repeat tract. In contrast, Staphylococcus aureus Cas9 (SaCas9) was unable to contract the CTG repeat tract. In parallel, we completely sequenced to very high coverage the DM1 genome using the PacBio technology. Several clones in which the TALEN was induced were also totally sequenced. In some of them, length changes of other long CTG repeats were detected, possibly corresponding to off-target effects, all of them in introns or intergenic regions. Repeat contractions were never associated with recombination of flanking markers, suggesting that contractions most probably occur by an intra-allelic mechanism such as single-strand annealing. TALENs should now be considered as a promising gene therapy approach, not only for DM1 but also for many other microsatellite expansion disorders.

genomics↗

bioRxiv2022

Functional variability in adhesion and flocculation of yeast megasatellite genes

Megasatellites are large tandem repeats found in all fungal genomes but especially abundant in the opportunistic pathogen Candida glabrata. They are encoded in genes involved in cell-cell interactions, either between yeasts or between yeast and human cells. In the present work, we have been using an iterative genetic system to delete several C. glabrata megasatellite-containing genes and found that two of them were positively involved in adhesion to epithelial cells, whereas three genes controlled negatively adhesion. Two of the latter, CAGL0B05061g or CAGL0A04851g, are also negative regulators of yeast-to-yeast adhesion, making them central players in controlling C. glabrata adherence properties. Using a series of synthetic Saccharomyces cerevisiae strains in which the FLO1 megasatellite was replaced by other tandem repeats of similar length but different sequences, we showed that the capacity of a strain to flocculate in liquid culture was unrelated to its capacity to adhere to epithelial cells or to invade agar. Finally, in order to understand how megasatellites were initially created and subsequently expanded, an experimental evolution system was set up, in which modified yeast strains containing different megasatellite seeds were grown in bioreactors for more than 200 generations and selected for their ability to sediment at the bottom of the culture tube. Several flocculation-positive mutants were isolated. Functionally relevant mutations included general transcription factors as well as a 230 kb segmental duplication.

genetics↗

bioRxiv2021

Megasatellite formation and evolution in vertebrates

Since the formation of the first proto-eukaryotes, more than 1.5 billion years ago, eukaryotic gene repertoire as well as genome complexity has significantly increased. Among genetic elements that are responsible for this increase in genome coding capacity and plasticity are tandem repeats such as microsatellites, minisatellites and their bigger brothers, megasatellites. Although microsatellites have been thoroughly studied in many organisms for the last 20 years, little is known about the distribution and evolution of mini- and megasatellites. Here, we describe the first genome-wide analysis of megasatellites in 58 vertebrate genomes, belonging to 12 monophyletic groups. We show that two bursts of megasatellite formation occurred, one after the radiation between agnatha et gnathostomata fishes and the second one later, in therian mammals. Megasatellites are frequently encoded in genes involved in transcription regulation (zinc-finger proteins) and intracellular trafficking, but also in cell membrane metabolism, reminiscent of what was observed in fungi genomes. The presence of many introns within young megasatellites suggests a model in which an exon-intron DNA segment is first duplicated and amplified before the accumulation of mutations in intronic parts partially erase the tandem repeat in such a way that it becomes detectable only in exonic regions. In addition, evidence for the genetic transfer of megasatellites between unrelated genes suggests that megasatellite formation and evolution is a very dynamic and still ongoing process in vertebrate genomes.

evolutionary biology↗

bioRxiv2019

Differential efficacies of Cas nucleases on microsatellites involved in human disorders and associated off-target mutations

Microsatellite expansions are the cause of more than 20 neurological or developmental human disorders. Shortening expanded repeats using specific DNA endonucleases may be envisioned as a gene editing approach. Here, a new assay was developed to test several CRISPR-Cas nucleases on microsatellites involved in human diseases, by measuring at the same time double-strand break rates, DNA end resection and homologous recombination efficacy. Broad variations in nuclease performances were detected on all repeat tracts. Streptococcus pyogenes Cas9 was the most efficient of all. All repeat tracts did inhibit double-strand break resection. We demonstrate that secondary structure formation on the guide RNA was a major determinant of nuclease efficacy. Using deep sequencing, off-target mutations were assessed genomewide. Out of 221 CAG/CTG or GAA/TTC trinucleotide repeats of the yeast genome, three were identified as carrying statistically significant low frequency mutations, corresponding to off-target effects.

genetics↗

bioRxiv2019

Resection and repair of a Cas9 double-strand break at CTG trinucleotide repeats induces local and extensive chromosomal rearrangements

Microsatellites are short tandem repeats, ubiquitous in all eukaryotes and represent [~]2% of the human genome. Among them, trinucleotide repeats are responsible for more than two dozen neurological and developmental disorders. Targeting microsatellites with dedicated DNA endonucleases could become a viable option for patients affected with dramatic neurodegenerative disorders. Here, we used the Streptococcus pyogenes Cas9 to induce a double-strand break within the expanded CTG repeat involved in myotonic dystrophy type 1, integrated in a yeast chromosome. Repair of this double-strand break generated unexpected large chromosomal rearrangements around the repeat tract. These rearrangements depended on RAD52, DNL4 and SAE2, and both non-homologous end-joining and single-strand annealing pathways were involved. Resection and repair of the double-strand break (DSB) were totally abolished in a rad50{Delta} strain, whereas they were impaired in a sae2{Delta} mutant, only on the DSB end containing most of the repeat tract. This proved that Sae2 plays significant different roles in resecting a DSB end containing a repeated and structured sequence as compared to a non-repeated DSB end.\n\nIn addition, we also discovered that gene conversion was less efficient when the DSB could be repaired using a homologous template, suggesting that the trinucleotide repeat may interfer with gene conversion too. Altogether, these data show that SpCas9 is probably not a good choice when inducing a double-strand break at or near a microsatellite, especially in mammalian genomes that contain many more dispersed repeated elements than the yeast genome.

genetics↗

bioRxiv2018

More than 16,000 effectors in the Legionella genus genome provide multiple, independent combinations for replication in human cells

SignificanceLegionella pneumophila is a bacterial pathogen causing outbreaks of a lethal pneumonia. The genus Legionella comprises 65 species for which aquatic amoebae are the natural reservoirs. Using functional and comparative genomics to deconstruct the entire bacterial genus we reveal the surprising parallel evolutionary trajectories that have led to the emergence of human pathogenic Legionella. An unexpectedly large and unique repository of secreted proteins (>16,000) containing eukaryotic-like proteins acquired from all domains of life (plant, animal, fungal, archaea) is contrasting with a highly conserved type 4 secretion system. This study reveals an unprecedented environmental reservoir of bacterial virulence factors, and provides a new understanding of how reshuffling and gene-acquisition from environmental eukaryotic hosts, may allow for the emergence of human pathogens.\n\nAbstractThe bacterial genus Legionella comprises 65 species among, which Legionella pneumophila is a human pathogen causing severe pneumonia. To understand the evolution of an environmental to an accidental human pathogen, we have functionally analyzed 80 Legionella genomes spanning 58 species. Uniquely, an immense repository of 16,000 secreted proteins encoding 137 different eukaryotic-like domains and more than 200 eukaryotic-like proteins is paired with a highly conserved T4SS. Specifically, we show that eukaryotic Rho and Rab GTPase domains are found nearly exclusively in eukaryotes and Legionella species. Translocation assays for selected Rab-GTPase proteins revealed that they are indeed T4SS secreted substrates. Furthermore, F/U-box and SET domains were present in >70% of all species suggesting that manipulation of host signal transduction, protein turnover and chromatin modification pathways, respectively are fundamental intracellular replication strategies for Legionellae. In contrast, the Sec-7 domain was restricted to L. pneumophila and seven other species, indicating effector repertoire tailoring within different amoebae. Functional screening of 47 species revealed 60% were competent for intracellular replication in THP-1 cells, but interestingly this phenotype was associated with diverse effector assemblages. These data, combined with evolutionary analysis indicate that the capacity to infect eukaryotic cells has been acquired independently many times within the genus and that a highly conserved yet versatile T4SS secretes an exceptional number of different proteins shaped by inter-domain gene transfer. Furthermore we revealed the surprising extent to which legionellae have co-opted genes and thus cellular functions from their eukaryotic hosts and provides a new understanding of how dynamic reshuffling and gene-acquisition has led to the emergence of major human pathogens.

genomics↗

bioRxiv2017

Redesigning chromosomes to optimize conformation capture (Hi-C) assays

In all chromosome conformation capture based experiments the accuracy with which contacts are detected varies considerably because of the uneven distribution of restriction sites along genomes. In addition, repeated sequences as well as homologous, large identical regions remain invisible to the assay because of the ambiguities they introduce during the alignment of the sequencing reads along the genome. As a result, the investigation of homologs during meiosis prophase through 3C studies has been limited. Here, we redesigned and reassembled in yeast a 145kb region with regularly spaced restriction sites for various enzymes. Thanks to this Syn-3C design, we enhanced the signal to noise ratio and improved the visibility of the entire region. We also improved our understanding of Hi-C data and definition of resolution. The redesigned sequence is now distinguishable from its native homologous counterpart in an isogenic diploid strain. As a proof of principle, we track the establishment of homolog pairing during meiotic prophase in a synchronized population. This provides new insights on the individualization and pairing of homologs, as well as on their internal restructuration into arrays of loops during meiosis prophase. Overall, we show the interest of redesigned genomic regions to explore complex biological questions otherwise difficult to address.

synthetic biology↗