RESUMO
It is known that nonsense suppression efficiency in yeast is controlled both genetically and epigenetically. As many components of translation machinery are represented by phosphoproteins, it depends, in particular, on the activity of kinases and phosphatases. The Ppz1p/Hal3p complex is among them. In this complex, the Ppz1p phosphatase is a catalytic subunit and Hal3p negatively regulates its function. The aim of this work was to study mechanisms which relate the activity of Ppz1p/Hal3p complex to nonsense suppression efficiency. In this study we used a genetic approach consisting of the analysis of nonsense suppression phenotype of strains over-expressing HAL3 or PPZ1 genes and also bearing deletions or mutant alleles of genes which presumably could participate in the manifestation of these over-expressions. We have shown that Hal3p inhibits not only Ppz1p, but also the homologous phosphatase Ppz2p. Our data indicate that Ppz2p is also involved in the control of nonsense suppression efficiency. In the course of search for Ppz1p target protein, it was shown that Ppz1p dephosphorylates at least two proteins participating in translation. Moreover, Ppz1p affects nonsense suppression efficiency not only due to its phosphatase activity but also due to another mechanism triggered by its interaction with Hsp70 chaperones.
Assuntos
Proteínas de Ciclo Celular/metabolismo , Códon sem Sentido/genética , Fosfoproteínas Fosfatases/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/genética , Supressão Genética , Proteínas de Ciclo Celular/genética , Fator 1 de Elongação de Peptídeos/genética , Fator 1 de Elongação de Peptídeos/metabolismo , Fosfoproteínas Fosfatases/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genéticaRESUMO
The prion-like determinant [ISP+] manifests itself as antisuppressor of certain sup35 mutations. To establish that [ISP+] actually represents a new yeast prion, it is necessary to identify the gene encoding protein corresponding in its prion form to [ISP+]. Analysis of transformants obtained by transformation of [ISP+] strain with insertion gene library revealed three genes controlling the [ISP+] maintenance. These genes are UPF1, UPF2 and SFP1. The SFP1 encodes potenlially prionogenic protein, as it is enriched with asparagine and glutamine residues. Therefore it is the most likely candidate to the role of [ISP+] structural gene. The UPF1 and UPF2 products are components of nonsense-mediated mRNA decay. It was shown that [ISP+] elimination caused by UPF1 and UPF2 inactivation is reversible. It was shown also that Upf1 and Upf2 proteins are not related functionally to Ppzlp phosphatase, influencing [ISP+] manifestation. Possible mechanisms of UPF1 and UPF2 influence on [ISP+] maintenance are discussed.
Assuntos
Regulação Fúngica da Expressão Gênica/fisiologia , Biblioteca Gênica , Príons/metabolismo , Saccharomyces cerevisiae/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Mutação , Fatores de Terminação de Peptídeos/genética , Fatores de Terminação de Peptídeos/metabolismo , Fosfoproteínas Fosfatases/genética , Fosfoproteínas Fosfatases/metabolismo , Príons/genética , RNA Helicases/genética , RNA Helicases/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismoRESUMO
The phenomenon of nonsense suppression, which leads to the reading of stop codons as sense codons, may be related to disturbances in the operation of various components of the translation apparatus and the proteins interacting with them. The phosphatase Ppzlp is one of the factors affecting the nonsense suppression efficiency in the saccharomycete yeast. In this work, the impact of the overexpression of gene PPZ1 and its mutant allele PPZ1-R451L on the phenotypic expression of various mutant alleles of genes SUP35 and SUP45 or the yeast prion [PSI+] was analyzed. On the basis of the data obtained, a suggestion about the possible role of proteins Sup35p and Sup45p in the processes mediating the influence of gene PPZ1 overexpression on the efficiency of nonsense suppression is made.
