Project description:Ribosome specialization is an emerging concept which challenges the common assumption that translation relies on a standardized molecular machinery. In this work, we demonstrate that Tma108, a yeast uncharacterized translation machinery-associated factor, defines a subpopulation of the cellular ribosomes specifically involved in the translation of less than 200 mRNAs encoding proteins with ATP or zinc binding domains. Ribonucleoparticle dissociation experiments support the fact that Tma108 directly interacts with the nascent protein chain. Comparative genomic analyses and molecular modeling point out Tma108 as an original M1 metallopeptidase with specific residues in the catalytic pocket which may explain its selectivity. The involvement of Tma108 in co-translational regulations is attested by the drastic perturbation of the subcellular localization of ATP2 mRNA, one of its targets, upon TMA108 inactivation. Tma108 is an unique example of a nascent chain-associated factor with high selectivity and illustrates the existence of specific translation-associated factors, besides RNA binding proteins.
Project description:This project aims to identify novel RNA binding proteins in the baker's yeast, Saccharomyces cerevisiae. Since interactions between RNAs and proteins may be transient, yeast cells were crosslinked with UV light at 254 nm which promotes the covalent link between proteins and RNAs. After this, polyadenylated mRNAs were purified via oligo(dT) coupled to magentic beads under stringet conditions. Finally, samples were subjected to mass spectrometry analysis. To rule out the possibility of RNA-independent binding we also analysed other samples: i) samples digested with RNase one; ii) samples where we performed competition assays with polyadenylic acid.
