Biology subjects

Fichant, G.

Publications and source records attributed to Fichant, G..

4 recordsLinked to original sources

bioRxiv2025

rnaends: an R package targeted to study the exact RNA ends at the nucleotide resolution

AO_SCPLOWBSTRACTC_SCPLOW5 and 3 RNA-end sequencing protocols have unlocked new opportunities to study aspects of RNA metabolism such as synthesis, maturation and degradation, by enabling the quantification of exact ends of RNA molecules in vivo. From RNA-Seq data that have been generated with one of the specialized protocols, it is possible to identify transcription start sites (TSS) and/or endoribonucleolytic cleavage sites, and even, in some cases, co-translational 5 to 3 degradation dynamics. Furthermore, post-transcriptional addition of ribonucleotides at the 3 end of RNA can be studied at the nucleotide resolution. While different RNA-end sequencing library protocols exist that have been adapted to a specific organism (prokaryote or eukaryote) or specific biological question, the generated RNA-Seq data are very similar and share common processing steps. Most importantly, the major aspect of RNA-end sequencing is that only the 5 or 3 end mapped location is of interest, contrary to conventional RNA sequencing that considers genomic ranges for gene expression analysis. This translates to a simple representation of the quantitative data as a count matrix of RNA-end location on the reference sequences. This representation seems under-exploited and is, to our knowledge, not available in a generic package focused on the analyses on the exact transcriptome ends. Here, we present the rnaends R package which is dedicated to RNA-end sequencing analysis. It offers functions for raw read pre-processing, RNA-end mapping and quantification, RNA-end count matrix post-processing, and further downstream count matrix analyses such as TSS identification, fast Fourier transform for signal periodic pattern analysis, or differential proportion of RNA-end analysis. The use of rnaends is illustrated here with applications in RNA metabolism studies through selected rnaends workflows on published RNA-end datasets: (i) TSS identification, (ii) ribosome translation speed and co-translational degradation, (iii) post-transcriptional modification analysis and differential proportion analysis.

bioinformatics↗

bioRxiv2019

RNA processing machineries in Archaea: the 5’-3’ exoribonuclease aRNase J of the β-CASP family is engaged specifically with the helicase ASH-Ski2 and the 3’-5’ exoribonucleolytic RNA exosome machinery

A network of RNA helicases, endoribonucleases, and exoribonucleases regulates the quantity and quality of cellular RNAs. To date, mechanistic studies focused on bacterial and eukaryal systems due to the challenge of identifying the main drivers of RNA decay and processing in Archaea. Here, our data support that aRNase J, a 5-3 exoribonuclease of the {beta}-CASP family conserved in Euryarchaea, engages specifically with a Ski2-like helicase and the RNA exosome to potentially exert control over RNA surveillance, and that this occurs in the vicinity of the ribosome. Proteomic landscapes and direct protein-protein interaction analyses demonstrated that aRNase J interplay with ASH-Ski2 and the Csl4 cap exosome subunit. These in vitro data are strengthened by our phylogenomic studies showing a taxonomic co-distribution of aRNase J and ASH-Ski2 among the archaeal phylogeny. Finally, our T. barophilus whole-cell extract fractionation experiments provide evidences that an aRNase J/ASH-Ski2 complex might exist in vivo and hint at an association of aRNase J with the ribosome or polysomes that is stressed in absence of ASH-Ski2. While aRNase J homologues are found among bacteria, the RNA exosome and the Ski2-like RNA helicase have eukaryotic homologues, underlining the mosaic aspect of archaeal RNA machines. Altogether, these results suggest, for the first time, a fundamental role of {beta}-CASP RNase/helicase complex in archaeal RNA metabolism. Finally, our results position aRNase J at the junction of RNA surveillance and translation processes, thus opening new perspectives and evolutionary scenario on RNA processing players in Archaea.

molecular biology↗

bioRxiv2018

Dynamic modeling of Streptococcus pneumoniae competence provides regulatory mechanistic insights

In the human pathogen Streptococcus pneumoniae, the gene regulatory circuit leading to the transient state of competence for natural transformation is based on production of an auto-inducer that activates a positive feedback loop. About one hundred genes are activated in two successive waves linked by a central alternative sigma factor ComX. This mechanism appears to be fundamental to the biological fitness of S. pneumoniae. We have developed a knowledge-based model of the competence cycle that describes average cell behavior. It reveals that the expression rates of the two competence operon, comAB and comCDE, involved in the positive feedback loop must be coordinated to elicit spontaneous competence. Simulations revealed the requirement for an unknown late com gene product that shuts of competence by impairing ComX activity. Further simulations led to the predictions that the membrane protein ComD bound to CSP reacts directly to pH change of the medium and that blindness to CSP during the post-competence phase is controlled by late DprA protein. Both predictions were confirmed experimentally.

systems biology↗

bioRxiv2018

Single-strand DNA processing: phylogenomics and sequence diversity of a superfamily of potential prokaryotic HuH endonucleases

BackgroundSome mobile genetic elements target the lagging strand template during DNA replication. Bacterial examples are insertion sequences IS608 and ISDra2 (IS200/IS605 family members). They use obligatory single-stranded circular DNA intermediates for excision and insertion and encode a transposase, TnpAIS200, which recognizes subterminal secondary structures at the insertion sequence ends. Similar secondary structures, Repeated Extragenic Palindromes (REP), are present in many bacterial genomes. TnpAIS200-related proteins, TnpAREP, have been identified and could be responsible for REP sequence proliferation. These proteins share a conserved HuH/Tyrosine core domain responsible for catalysis and are involved in processes of ssDNA cleavage and ligation. Our goal is to characterize the diversity of these proteins collectively referred as the TnpAY1 family.\n\nResultsA genome-wide analysis of sequences similar to TnpAIS200 and TnpAREP in prokaryotes revealed a large number of family members with a wide taxonomic distribution. These can be arranged into three distinct classes and 12 subclasses based on sequence similarity. One subclass includes sequences similar to TnpAIS200. Proteins from other subclasses are not associated with typical insertion sequence features. These are characterized by specific additional domains possibly involved in protein/DNA or protein/protein interactions. Their genes are found in more than 25% of species analyzed. They exhibit a patchy taxonomic distribution consistent with dissemination by horizontal gene transfers followed by loss. The tnpAREP genes of five subclasses are flanked by typical REP sequences in a REPtron-like arrangement. Four distinct REP types were characterized with a subclass specific distribution. Other subclasses are not associated with REP sequences but have a large conserved domain located in C-terminal end of their sequence. This unexpected diversity suggests that, while most likely involved in processing single-strand DNA, proteins from different subfamilies may play a number of different roles.\n\nConclusionsWe established a detailed classification of TnpAY1 proteins, consolidated by the analysis of the conserved core domains and the characterization of additional domains. The data obtained illustrate the unexpected diversity of the TnpAY1 family and provide a strong framework for future evolutionary and functional studies. By their potential function in ssDNA edition, they may confer adaptive responses to host cell physiology and metabolism.

genomics↗