Biology subjects

Claverol, S.

Publications and source records attributed to Claverol, S..

5 recordsLinked to original sources

bioRxiv2018

Cross-linking/Mass Spectrometry: A Community-Wide, Comparative Study Towards Establishing Best Practice Guidelines

The number of publications in the field of chemical cross-linking combined with mass spectrometry (XL-MS) to derive constraints for protein three-dimensional structure modeling and to probe protein-protein interactions has largely increased during the last years. As the technique is now becoming routine for in vitro and in vivo applications in proteomics and structural biology there is a pressing need to define protocols as well as data analysis and reporting formats that are generally accepted in the field and that have shown to lead to high-quality results. This first, community-based harmonization study on XL-MS is based on the results of 32 groups participating worldwide. The aim of this paper is to summarize the status quo of XL-MS and to compare and evaluate existing cross-linking strategies. From the results obtained, common protocols will be established. Our study serves as basis for establishing best practice guidelines in the field for conducting cross-linking experiments, performing data analysis, and reporting formats with the ultimate goal of assisting scientists to generate accurate and reproducible XL-MS results.

biochemistry↗

bioRxiv2018

Multiple C2 domains and Transmembrane region Proteins (MCTPs) tether membranes at plasmodesmata

AO_SCPLOWBSTRACTC_SCPLOWIn eukaryotes, membrane contact sites (MCS) allow direct communication between organelles. Plants have evolved unique MCS, the plasmodesmata intercellular pores, which combine endoplasmic reticulum (ER) - plasma membrane (PM) contacts with regulation of cell-to-cell signalling. The molecular mechanism and function of membrane tethering within plasmodesmata remains unknown.\n\nHere we show that the Multiple C2 domains and Transmembrane region Protein (MCTP) family, key regulators of cell-to-cell signalling in plants, act as ER - PM tethers specifically at plasmodesmata. We report that MCTPs are core plasmodesmata proteins that insert into the ER via their transmembrane region whilst their C2 domains dock to the PM through interaction with anionic phospholipids. A mctp3/4 loss-of-function mutant induces plant developmental defects while MCTP4 expression in a yeast {Delta}tether mutant partially restores ER-PM tethering. Our data suggest that MCTPs are unique membrane tethers controlling both ER-PM contacts and cell-cell signalling.

plant biology↗

bioRxiv2018

Functional recruitment of dynamin requires multimeric interactions for efficient endocytosis

During clathrin mediated endocytosis (CME), membrane scission is achieved by the concerted action of dynamin and its interacting partners. Essential interactions occur between the proline/arginine-rich domain of dynamin (dynPRD) and the Src-homology domain 3 (SH3) of various proteins including amphiphysins. Here we show that multiple SH3 domains must bind simultaneously to dynPRD through three adjacent motifs for dynamins efficient recruitment and function. First, we show in dynamin triple knock-out cells that mutant dynamins modified in a single motif, including the central amphiphysin SH3 (amphSH3) binding motif, are partially capable of rescuing CME. However, mutating two motifs largely prevents that ability. To support this observation, we designed divalent dynPRD-derived peptides. These ligands bind multimers of amphSH3 with >100-fold higher affinity than monovalent ones in vitro. Accordingly, dialyzing living cells with these divalent peptides through a patch-clamp pipette blocks CME 2 to 3 times more effectively than with monovalent ones. Finally, the frequency of endocytic events decreases with competing peptides or hypomorphic rescue mutants but the kinetics of dynamin recruitment is unaffected. This suggests that PRD-SH3 interactions act upstream of dynamin accumulation at the neck of nascent vesicles. We conclude from these data that dynamin drives vesicle scission via multivalent interactions in vivo.

cell biology↗

bioRxiv2018

The Integrative Conjugative Element (ICE) of Mycoplasma agalactiae: key elements involved in horizontal dissemination and influence of co-resident ICEs

The discovery of integrative conjugative elements (ICEs) in wall-less mycoplasmas and the demonstration of their role in massive gene flows within and across species has shed new light on the evolution of these minimal bacteria. Of these, ICEA of the ruminant pathogen Mycoplasma agalactiae represents a prototype and belongs to a new clade of the Mutatorlike superfamily that has no preferential insertion site and often occurs as multiple chromosomal copies. Here, functional genomics and mating experiments were combined to address ICEA functions and define the minimal ICEA chassis conferring conjugative properties to M. agalactiae. Data further indicated a complex interaction among co-resident ICEAs, since the minimal ICEA structure was influenced by the occurrence of additional ICEA copies that can trans-complement conjugative-deficient ICEAs. However, this cooperative behavior was limited to the CDS14 surface lipoprotein, which is constitutively expressed by co-resident ICEAs, and did not extend to other ICEA proteins including the cis-acting DDE recombinase and components of the mating channel whose expression was only sporadically detected. Remarkably, conjugative-deficient mutants containing a single ICEA copy knocked-out in cdsl4 can be complemented by neighboring cells expressing CDS14. This result, together with the conservation of CDS14 functions in closely related species, may suggest a way for mycoplasma ICEs to extend their interaction outside of their chromosomal environment. Overall, this study provides a first model of conjugative transfer in mycoplasmas and offers valuable insights towards the understanding of horizontal gene transfer in this highly adaptive and diverse group of minimal bacteria.\n\nIMPRTANCEIntegrative conjugative elements (ICEs) are self-transmissible mobile genetic elements that are key mediators of horizontal gene flow in bacteria. Recently, a new category of ICEs has been identified that confer conjugative properties to mycoplasmas, a highly adaptive and diverse group of wall-less bacteria with reduced genomes. Unlike classical ICEs, these mobile elements have no preferential insertion specificity and multiple mycoplasma ICE copies can be found randomly integrated into the host chromosome. Here, the prototype ICE of Mycoplasma agalactiae was used to define the minimal conjugative machinery and propose the first model of ICE transfer in mycoplasmas. This model unveils the complex interactions taking place among co-resident ICEs and suggests a way for these elements to extend their influence outside of their chromosomal environment. These data pave the way for future studies aiming at deciphering chromosomal transfer, an unconventional mechanism of DNA swapping that has been recently associated with mycoplasma ICEs.

microbiology↗

bioRxiv2017

H2S mediates interbacterial communication through the air reverting intrinsic antibiotic resistance

Hydrogen sulfide, a gas classically considered as a by-product of cellular metabolism, is today recognized as a crucial gasotransmitter in Eukaryotes. Moreover, most bacteria harbor the eukaryotic orthologous genes for H2S synthesis, and these genes have been linked to different metabolic pathways.\n\nSome bacteria, however, produce high amounts of H2S in their extracellular space, a characteristic classically used for identification purposes. This is the case of Salmonella Typhimurium, which produces H2S by its phsABC operon. Here we show that extracellular release of H2S by S. Typhimurium is solely dependent on its phsABC operon. Furthermore, we show that S. Typhimurium and other H2S-producing bacteria can interact with physically distant bacteria through H2S production. We demonstrate how H2S can revert intrinsic cephalosporin resistance of Enterococccus faecalis and Enterococcus faecium to complete susceptibility. This study constitutes a significant step in the study of bacterial interplay and niche competition. Furthermore, as H2S releasing drugs have already been designed, our results open the way to future therapeutic alternatives for the treatment of infections caused by enterococci, multiresistant pathogens for which no treatments are clinically available.\n\nAuthor SummaryIt has been known for decades that bacteria can communicate with each other through the diffusion of metabolites in the media. However, the capacity of a bacterium to interact with other physically distant cell is a recent discovery of the 21st century. In this work we show how some well-studied bacteria, as it is Salmonella spp., interacts with other bacteria thanks to the compound hydrogen sulfide (H2S) that they produce and release to the environment.\n\nIn our study we have designed novel techniques that allow us to study the interaction between two bacteria, and we have seen that Salmonella is able to affect other species that is even 1 cm away, i.e., a distance corresponding to 10.0000 times its own size.\n\nWhat is more astonishing is that Enterococcus, when exposed to the H2S, is dramatically becomes susceptible to many antibiotics, to which it is supposed to be naturally resistant. Enterococcus spp. are responsible for life-threatening infections in hospitals worldwide. Thus, our observations reveal that bacteria can communicate through the air with H2S, and that this molecule can make bacteria that are highly resistant to antibiotics susceptible to antibiotics, making untreatable infections treatable with current antibiotics.

microbiology↗