Biology subjects

GRAILLE, M.

Publications and source records attributed to GRAILLE, M..

4 recordsLinked to original sources

bioRxiv2025

Mapping interactions within the NMD decapping complex reveals how Upf1 recruits Dcp2 onto mRNA targets

Upf1 RNA helicase is a pivotal factor in the conserved nonsense-mediated mRNA decay (NMD) process. Upf1 is responsible for coordinating the recognition of premature termination codons (PTC) in a translation-dependent manner and subsequently triggering mRNA degradation. Multiple factors assist Upf1 during these two consecutive steps. In Saccharomyces cerevisiae, Upf2 and Upf3 associated to Upf1 (Upf1-2/3) contribute to PTC recognition but are absent from the Upf1-decapping complex that includes Nmd4, Ebs1, Dcp1, and Dcp2. Despite their importance for NMD, the organization and dynamics of these Upf1-containing complexes remains unclear. Here, we show how distinct domains of Upf1 make direct contacts with Dcp1/Dcp2, Nmd4 and Ebs1 using recombinant proteins. These proteins also bind to each other forming an extended network of interactions within the Upf1-decapping complex. Dcp2 and Upf2 are shown to compete for the same binding site in the Upf1 N-terminal CH domain. This finding accounts for the isolation of two mutually exclusive Upf1-containing complexes from cell lysates. Furthermore, we show how Nmd4-assisted recruitment of Upf1 promotes the anchoring of the decapping protein to the mRNA.

biochemistry↗

bioRxiv2022

The SARS-CoV-2 protein NSP2 impairs the microRNA-induced silencing capacity of human cells

The coronavirus SARS-CoV-2 is the cause of the ongoing pandemic of COVID-19. Given the absence of effective treatments against SARS-CoV-2, there is an urgent need for a molecular understanding of how the virus influences the machineries of the host cell. The SARS-CoV-2 generates 16 Non-Structural Proteins (NSPs) through proteolytic cleavage of a large precursor protein. In the present study, we focused our attention on the SARS-CoV-2 protein NSP2, whose role in the viral pathogenicity is poorly understood. Recent proteomic studies shed light on the capacity of NSP2 to bind the 4EHP-GIGYF2 complex, a key factor involved in microRNA-mediated silencing of gene expression in human cells. In order to gain a better understanding of the function of NSP2, we attempted to identify the molecular basis of its interaction with 4EHP-GIGYF2. Our data demonstrate that NSP2 physically associates with the endogenous 4EHP-GIGYF2 complex in the cytoplasm. Using co-immunoprecipitation and in vitro interaction assays, we identified both 4EHP and a central segment in GIGYF2 as binding sites for NSP2. We also provide functional evidence that NSP2 impairs the function of GIGYF2 in mediating mRNA silencing using reporter-based assays, thus leading to a reduced activity of microRNAs. Altogether, these data reveal the profound impact of NSP2 on the post-transcriptional silencing of gene expression in human cells, pointing out 4EHP-GIGYF2 targeting as a possible strategy of SARS-CoV-2 to take over the silencing machinery and to suppress host defenses.

cell biology↗

bioRxiv2021

Artificial intelligence to solve the X-ray crystallography phase problem: a case study report.

The determination of three dimensional structures of macromolecules is one of the actual challenge in biology with the ultimate objective of understanding their function. So far, X-ray crystallography is the most popular method to solve structure, but this technique relies on the generation of diffracting crystals. Once a correct data set has been obtained, the calculation of electron density maps requires to solve the so-called < > using different approaches. The most frequently used technique is molecular replacement, which relies on the availability of the structure of a protein sharing strong structural similarity with the studied protein. Its success rate is directly correlated with the quality of the models used for the molecular replacement trials. The availability of models as accurate as possible is then definitely critical. Very recently, a breakthrough step has been made in the field of protein structure prediction thanks to the use of machine learning approaches as implemented in the AlphaFold or RoseTTAFold structure prediction programs. Here, we describe how these recent improvements helped us to solve the crystal structure of a protein involved in the nonsense-mediated mRNA decay pathway (NMD), an mRNA quality control pathway dedicated to the elimination of eukaryotic mRNAs harboring premature stop codons.

biochemistry↗

bioRxiv2019

Formation of S. pombe Erh1 homodimer mediates gametogenic gene silencing and meiosis progression

Timely and accurate expression of the genetic information relies on the integration of environmental cues and the activation of regulatory networks involving transcriptional and post-transcriptional mechanisms. In fission yeast, meiosis-specific transcripts are selectively targeted for degradation during mitosis by the EMC complex, composed of Erh1, the ortholog of human ERH, and the YTH family RNA-binding protein Mmi1. Here, we present the crystal structure of Erh1 and show that it assembles as a homodimer. Mutations of amino acid residues to disrupt Erh1 homodimer formation result in loss-of-function phenotypes, similar to erh1{triangleup} cells: expression of meiotic genes is derepressed in mitotic cells and meiosis progression is severely compromised. Interestingly, formation of Erh1 homodimer is dispensable for interaction with Mmi1, suggesting that only fully assembled EMC complexes consisting of two Mmi1 molecules bridged by an Erh1 dimer are functionally competent. We also show that Erh1 does not contribute to Mmi1-dependent down-regulation of the meiosis regulator Mei2, supporting the notion that Mmi1 performs additional functions beyond EMC. Overall, our results provide a structural basis for the assembly of the EMC complex and highlight its biological relevance in gametogenic gene silencing and meiosis progression.

biochemistry↗