Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 3 de 3
Filtrar
Más filtros

Bases de datos
Tipo de estudio
Tipo del documento
País de afiliación
Intervalo de año de publicación
1.
Genetics ; 181(1): 105-18, 2009 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-18984568

RESUMEN

There is significant evidence linking nucleocytoplasmic transport to cell cycle control. The budding yeast, Saccharomyces cerevisiae, serves as an ideal model system for studying transport events critical to cell cycle progression because the nuclear envelope remains intact throughout the cell cycle. Previous studies linked the classical nuclear localization signal (cNLS) receptor, importin-alpha/Srp1, to the G(2)/M transition of the cell cycle. Here, we utilize two engineered mutants of importin-alpha/Srp1 with specific molecular defects to explore how protein import affects cell cycle progression. One mutant, Srp1-E402Q, is defective in binding to cNLS cargoes that contain two clusters of basic residues termed a bipartite cNLS. The other mutant, Srp1-55, has defects in release of cNLS cargoes into the nucleus. Consistent with distinct in vivo functional consequences for each of the Srp1 mutants analyzed, we find that overexpression of different nuclear transport factors can suppress the temperature-sensitive growth defects of each mutant. Studies aimed at understanding how each of these mutants affects cell cycle progression reveal a profound defect at the G(1) to S phase transition in both srp1-E402Q and srp1-55 mutants as well as a modest G(1)/S defect in the temperature-sensitive srp1-31 mutant, which was previously implicated in G(2)/M. We take advantage of the characterized defects in the srp1-E402Q and srp1-55 mutants to predict candidate cargo proteins likely to be affected in these mutants and provide evidence that three of these cargoes, Cdc45, Yox1, and Mcm10, are not efficiently localized to the nucleus in importin-alpha mutants. These results reveal that the classical nuclear protein import pathway makes important contributions to the G(1)/S cell cycle transition.


Asunto(s)
Fase G1 , Carioferinas/metabolismo , Señales de Localización Nuclear/metabolismo , Fase S , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/citología , alfa Carioferinas/metabolismo , Transporte Activo de Núcleo Celular , Núcleo Celular/metabolismo , Genes Supresores , Proteínas Fluorescentes Verdes/metabolismo , Proteínas Mutantes/metabolismo , Mutación/genética , Plásmidos/genética , Saccharomyces cerevisiae/genética
2.
Nucleic Acids Res ; 36(13): 4317-26, 2008 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-18586821

RESUMEN

Like its retroviral relatives, the long terminal repeat retrotransposon Ty1 in the yeast Saccharomyces cerevisiae must traverse a permanently intact nuclear membrane for successful transposition and replication. For retrotransposition to occur, at least a subset of Ty1 proteins, including the Ty1 integrase, must enter the nucleus. Nuclear localization of integrase is dependent upon a C-terminal nuclear targeting sequence. However, the nuclear import machinery that recognizes this nuclear targeting signal has not been defined. We investigated the mechanism by which Ty1 integrase gains access to nuclear DNA as a model for how other retroelements, including retroviruses like HIV, may utilize cellular nuclear transport machinery to import their essential nuclear proteins. We show that Ty1 retrotransposition is significantly impaired in yeast mutants that alter the classical nuclear protein import pathway, including the Ran-GTPase, and the dimeric import receptor, importin-alpha/beta. Although Ty1 proteins are made and processed in these mutant cells, our studies reveal that an integrase reporter is not properly targeted to the nucleus in cells carrying mutations in the classical nuclear import machinery. Furthermore, we demonstrate that integrase coimmunoprecipitates with the importin-alpha transport receptor and directly binds to importin-alpha. Taken together, these data suggest Ty1 integrase can employ the classical nuclear protein transport machinery to enter the nucleus.


Asunto(s)
Núcleo Celular/metabolismo , Integrasas/metabolismo , Proteínas Nucleares/metabolismo , Retroelementos , Proteínas de Saccharomyces cerevisiae/metabolismo , Transporte Activo de Núcleo Celular , Núcleo Celular/química , Citoplasma/química , Proteínas Fluorescentes Verdes/análisis , Integrasas/química , Mutación , Señales de Localización Nuclear , Saccharomyces cerevisiae/enzimología , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , alfa Carioferinas/genética , alfa Carioferinas/metabolismo , Proteína de Unión al GTP ran/genética , Proteína de Unión al GTP ran/metabolismo
3.
J Biol Chem ; 281(33): 23545-56, 2006 Aug 18.
Artículo en Inglés | MEDLINE | ID: mdl-16785238

RESUMEN

Nuclear localization signals (NLSs) target proteins into the nucleus through mediating interactions with nuclear import receptors. Here, we perform a quantitative analysis of the correlation between NLS receptor affinity and the steady-state distribution of NLS-bearing cargo proteins between the cytoplasm and the nucleus of live yeast, which reflects the relative import rates of various NLS sequences. We find that there is a complicated, but monotonic quantitative relationship between the affinity of an NLS for the import receptor, importin alpha, and the steady-state accumulation of the cargo in the nucleus. This analysis takes into consideration the impact of protein size. In addition, the hypothetical upper limit to an NLS affinity for the receptors is explored through genetic approaches. Overall, our results indicate that there is a correlation between the binding affinity of an NLS cargo for the NLS receptor, importin alpha, and the import rate for this cargo. This correlation, however, is not maintained for cargoes that bind to the NLS receptor with very weak or very strong affinity.


Asunto(s)
Señales de Localización Nuclear/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , alfa Carioferinas/metabolismo , Transporte Activo de Núcleo Celular/fisiología , Secuencia de Aminoácidos , Datos de Secuencia Molecular , Señales de Exportación Nuclear/fisiología , Unión Proteica/fisiología , Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/química , alfa Carioferinas/química
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA