RESUMO
The variety of mechanisms providing viability of organisms bearing nonsense-mutations in the essential genes is unknown at present. In yeast Saccharomyces cerevisiae nonsense-mutants containing premature stop-codon in mRNA of the essential SUP45 gene were obtained. These strains are viable in the absence of mutant suppressor tRNA, therefore it is supposed that there are alternative mechanisms providing nonsense-suppression and mutants viability. Analysis of transformants obtained by transformation of strain bearing nonsense-mutant allele of SUP45 gene with multicopy yeast genomic library revealed three genes encoding wild type tRNA(Tyr) and four genes encoding wild type tRNA(Gln) that improve nonsense-mutants viability. Moreover, overexpression of these genes leads to the increase in the amount of full-length eRF1 protein in cell and compensates nonsense-mutants sensitivity to high temperature. Probable mechanisms of tRNA(Tyr) and tRNA(Gln) influence on the increase of viability of nonsense-mutants in SUP45 gene are discussed in this work.
Assuntos
Códon sem Sentido , Regulação Fúngica da Expressão Gênica/fisiologia , Fatores de Terminação de Peptídeos/metabolismo , RNA Fúngico/biossíntese , RNA de Transferência de Glutamina/biossíntese , RNA de Transferência de Tirosina/biossíntese , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Fatores de Terminação de Peptídeos/genética , RNA Fúngico/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA de Transferência de Glutamina/genética , RNA de Transferência de Tirosina/genética , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genéticaRESUMO
Nonlethal nonsense mutations obtained earlier in the essential gene SUP45 encoding the translation termination eRFI factor in the yeast Saccharomyces cerevisiae were further characterized. Strains carrying these mutations retain the viability, since the full-length eRF1 protein is present in these strains, although in decreased amounts as compared to wild-type cells, together with a truncated eRF1. All nonsense mutations are likely to be located in a weak termination context, because a change in the stop codon UGAA (in the case of mutation sup45-107) to UAGA (sup45-107.2) led to the alteration of the local context from a weak to strong and to the lethality of the strain carrying sup45-107.2. All nonsense mutations studied are characterized by thermosensitivity expressed as cell mortality after cultivation at 37 degrees C. When grown under nonpermissive conditions (37 degrees C), cells of nonsense mutants sup45-104, sup45-105. and sup45-107 display a decrease in the amount of the truncated eRF1 protein without reduction in the amount of the full-length eRF1 protein. The results of this study suggest that the N-terminal eRF1 fragment is indispensable for cell viability of nonsense mutants due to the involvement in termination of translation.
