INVESTIGADORES
MORENO Silvia Margarita
congresos y reuniones científicas
Título:
CHARACTERIZATION OF THE INITIATION TRANSLATION FACTOR eIF4G1 AS A PKA SUBSTRATE IN Saccharomyces cerevisiae
Autor/es:
SOLARI, CLARA; CAMARANO ADRIANA; VALACCO MP; MORENO SILVIA; PORTELA PAULA
Lugar:
Buenos Aires
Reunión:
Congreso; Reunion conjunta de biociencias y 53 Reunion Anual SAIB; 2017
Institución organizadora:
SAIB y 9 sociedades de biociencias mas
Resumen:
PKA has a central role in the control of the metabolism, stress resistanceand proliferation in Saccharomyces cerevisiae. In responseto nutrient consumption, S. cerevisiae arrests cell cycle, decreasesglobal protein synthesis and initiates translation through internalribosome entry sites (IRES). We have recently demonstratedthat PKA connects the nutritional state of the cell with translationat different levels: through interaction with translation factors, by localizationin mRNA granules and affecting the protein abundanceof translation initiation factor eIF4G1 by a post-translational mechanism.Phosphoproteomic assays have reported that eIF4G1 is aphosphoprotein. The aim of this work was to characterize eIF4G1 asa PKA substrate and to identify the phosphorylation target residues.For this purpose, the full length and different fragments of recombinantproteins derived from GST-eIF4G1 were expressed and purifiedusing a pGEX system. Using these proteins, we assayed in vitrophosphorylation with Tpk1 and Ca as the catalytic subunit from S.cerevisiae and mammals respectively. The GST-eIF4G1 phosphorylationwas studied by western blot using an anti-P-Ser antibodyand autoradiography for the incorporation of γ-32P [ATP]. The phosphorylatedresidues were identified by nano-LC MS/MS applying aniron trap chromatography. Our results demonstrate that eIF4G1 is aPKA substrate in vitro and identify Thr934 as a novel phosphorylationtarget residue in eIF4G1. Since, Thr934 residue localizes on theRNA-binding domain RNA3 at the extreme C-terminus, we will focuson studying the role of this phosphorylation site over eIF4G1 proteinabundance and its recruitment to the 43S preinitiation complex onthe mRNA.