RESUMO
One important task of eukaryotic cells is to translate only mRNAs that were correctly processed to prevent the production of truncated proteins, found in neurodegenerative diseases and cancer. Nuclear quality control of splicing requires the SR-like proteins Gbp2 and Hrb1 in S. cerevisiae, where they promote the degradation of faulty pre-mRNAs. Here we show that Gbp2 and Hrb1 also function in nonsense mediated decay (NMD) of spliced premature termination codon (PTC)-containing mRNAs. Our data support a model in which they are in a complex with the Upf-proteins and help to transmit the Upf1-mediated PTC recognition to the transcripts ends. Most importantly they appear to promote translation repression of spliced transcripts that contain a PTC and to finally facilitate degradation of the RNA, presumably by supporting the recruitment of the degradation factors. Therefore, they seem to control mRNA quality beyond the nuclear border and may thus be global surveillance factors. Identification of SR-proteins as general cellular surveillance factors in yeast will help to understand the complex human system in which many diseases with defects in SR-proteins or NMD are known, but the proteins were not yet recognized as general RNA surveillance factors.
Assuntos
Proteínas de Transporte Nucleocitoplasmático/metabolismo , Proteínas de Ligação a Poli(A)/metabolismo , RNA Helicases/metabolismo , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/genética , Códon sem Sentido , Citoplasma/genética , Regulação Fúngica da Expressão Gênica , Degradação do RNAm Mediada por Códon sem Sentido , RNA Fúngico/genética , Saccharomyces cerevisiae/metabolismoRESUMO
Translation termination requires eRF1 and eRF3 for polypeptide- and tRNA-release on stop codons. Additionally, Dbp5/DDX19 and Rli1/ABCE1 are required; however, their function in this process is currently unknown. Using a combination of in vivo and in vitro experiments, we show that they regulate a stepwise assembly of the termination complex. Rli1 and eRF3-GDP associate with the ribosome first. Subsequently, Dbp5-ATP delivers eRF1 to the stop codon and in this way prevents a premature access of eRF3. Dbp5 dissociates upon placing eRF1 through ATP-hydrolysis. This in turn enables eRF1 to contact eRF3, as the binding of Dbp5 and eRF3 to eRF1 is mutually exclusive. Defects in the Dbp5-guided eRF1 delivery lead to premature contact and premature dissociation of eRF1 and eRF3 from the ribosome and to subsequent stop codon readthrough. Thus, the stepwise Dbp5-controlled termination complex assembly is essential for regular translation termination events. Our data furthermore suggest a possible role of Dbp5/DDX19 in alternative translation termination events, such as during stress response or in developmental processes, which classifies the helicase as a potential drug target for nonsense suppression therapy to treat cancer and neurodegenerative diseases.