Etude de la structure de l'hélicase Prp43p et l'interaction avec ses partenaires à domaine G-patch

Les ribosomes sont les machines moléculaires responsables de la production des protéines, constituants majeurs de la cellule. La synthèse des ribosomes est un processus très important et très coûteux en énergie pour la cellule. Comme tous les processus de la vie de la cellule, la biogenèse des ribosomes doit être régulée de manière efficace. Si cette régulation est défectueuse, elle peut être à l'origine de certaines pathologies. Les mécanismes de cette régulation restent mal compris malgré leur caractère essentiel. Néanmoins, on sait que plusieurs enzymes interviennent dans la synthèse des ribosomes mais leur mode d'action reste toutefois très peu élucidé. L'activité de ces enzymes peut être régulée via l'interaction avec des protéines ayant des structures spécifiques. Ce projet de recherche vise à étudier Prp43p, une des enzymes intervenant dans la synthèse des ribosomes et à comprendre comment elle agit en association avec ces protéines. Une meilleure compréhension de ces interactions est nécessaire pour élucider le mécanisme d'activation de Prp43p.Ribosomes are molecular machines responsible for the production of proteins, which are major components of the cell. The synthesis of ribosomes is a very important and energy-consuming process for the cell. Like most processes of the cell, ribosome biogenesis must be regulated in an effective and coordinated manner. If this regulation is defective, it may be the cause of certain diseases. The mechanisms of this regulation remain poorly understood despite their essential character. However, it is known that several enzymes are involved in the synthesis of ribosomes but their mode of action remains little understood. The activity of these enzymes can be regulated via the interaction with proteins having specific structures. This research project aims at studying Prp43p, one of the enzymes involved in the synthesis of ribosomes and understand how it acts in association with these proteins. A better understanding of these interactions is needed to elucidate the mechanism of activation of Prp43p

Prp43p contains a processive helicase structural architecture with a specific regulatory domain

The DEAH/RNA helicase A (RHA) helicase family comprises proteins involved in splicing, ribosome biogenesis and transcription regulation. We report the structure of yeast Prp43p, a DEAH/RHA helicase remarkable in that it functions in both splicing and ribosome biogenesis. Prp43p displays a novel structural architecture with an unforeseen homology with the Ski2-like Hel308 DNA helicase. Together with the presence of a β-hairpin in the second RecA-like domain, Prp43p contains all the structural elements of a processive helicase. Moreover, our structure reveals that the C-terminal domain contains an oligonucleotide/oligosaccharide-binding (OB)-fold placed at the entrance of the putative nucleic acid cavity. Deletion or mutations of this domain decrease the affinity of Prp43p for RNA and severely reduce Prp43p ATPase activity in the presence of RNA. We also show that this domain constitutes the binding site for the G-patch-containing domain of Pfa1p. We propose that the C-terminal domain, specific to DEAH/RHA helicases, is a central player in the regulation of helicase activity by binding both RNA and G-patch domain proteins

The ex planta signal activity of a Medicago ribosomal uL2 protein suggests a moonlighting role in controlling secondary rhizobial infection

We recently described a regulatory loop, which we termed autoregulation of infection (AOI), by which Sinorhizobium meliloti, a Medicago endosymbiont, downregulates the root susceptibility to secondary infection events via ethylene. AOI is initially triggered by so-far unidentified Medicago nodule signals named signal 1 and signal 1’ whose transduction in bacteroids requires the S. meliloti outer-membrane-associated NsrA receptor protein and the cognate inner-membrane-associated adenylate cyclases, CyaK and CyaD1/D2, respectively. Here, we report on advances in signal 1 identification. Signal 1 activity is widespread as we robustly detected it in Medicago nodule extracts as well as in yeast and bacteria cell extracts. Biochemical analyses indicated a peptidic nature for signal 1 and, together with proteomic analyses, a universally conserved Medicago ribosomal protein of the uL2 family was identified as a candidate signal 1. Specifically, MtRPuL2A (MtrunA17Chr7g0247311) displays a strong signal activity that requires S. meliloti NsrA and CyaK, as endogenous signal 1. We have shown that MtRPuL2A is active in signaling only in a non-ribosomal form. A Medicago truncatula mutant in the major symbiotic transcriptional regulator MtNF-YA1 lacked most signal 1 activity, suggesting that signal 1 is under developmental control. Altogether, our results point to the MtRPuL2A ribosomal protein as the candidate for signal 1. Based on the Mtnf-ya1 mutant, we suggest a link between root infectiveness and nodule development. We discuss our findings in the context of ribosomal protein moonlighting. Copyright: © 2020 Sorroche et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited