RESUMEN
Understanding mechanisms of post-transcriptional control of gene expression has come under much scrutiny in recent years. A key question in this field is how the translation of specific mRNAs is activated or repressed both spatially and temporally in a given cell. In oocytes of the frog Xenopus laevis a number of mRNAs are localized early in oogenesis and subsequently translated at later stages. We have developed a highly active cell-free translation system from oocytes in the early stages of oogenesis that is applicable to the study of translation and translational control of both endogenous and exogenous mRNAs.
Asunto(s)
Oocitos/citología , Xenopus laevis/metabolismo , Animales , Sistema Libre de Células , Técnicas Citológicas , Genes Reporteros , Luciferasas/genética , Luciferasas/metabolismo , Modelos Biológicos , Modelos Genéticos , Oocitos/metabolismo , Biosíntesis de Proteínas , ARN Mensajero/metabolismo , Factores de TiempoRESUMEN
mRNA localization is a crucial mechanism for post-transcriptional control of gene expression used in numerous cellular contexts to generate asymmetric enrichment of an encoded protein. This process has emerged as a fundamental regulatory mechanism that operates in a wide range of organisms to control an array of cellular processes. Recently, significant advances have been made in our understanding of the mechanisms that regulate several steps in the mRNA localization pathway. Here we discuss the progress made in understanding localization element recognition, paying particular attention to the role of RNA structure. We also consider the function of mRNP granules in mRNA transport, as well as new results pointing to roles for the endocytic pathway in mRNA localization.
Asunto(s)
Transporte de ARN , ARN Mensajero/metabolismo , Animales , Endocitosis , Células Eucariotas/metabolismo , Humanos , Conformación de Ácido Nucleico , ARN Mensajero/química , ARN Mensajero/genética , Ribonucleoproteínas/metabolismoRESUMEN
RNA localization is a widely conserved mechanism for generating cellular asymmetry. In Xenopus oocytes, microtubule-dependent transport of RNAs to the vegetal cortex underlies germ layer patterning. Although kinesin motors have been implicated in this process, the apparent polarity of the microtubule cytoskeleton has pointed instead to roles for minus-end-directed motors. To resolve this issue, we have analyzed participation of kinesin motors in vegetal RNA transport and identified a direct role for Xenopus kinesin-1. Moreover, in vivo interference and biochemical experiments reveal a key function for multiple motors, specifically kinesin-1 and kinesin-2, and suggest that these motors may interact during transport. Critically, we have discovered a subpopulation of microtubules with plus ends at the vegetal cortex, supporting roles for these kinesin motors in vegetal RNA transport. These results provide a new mechanistic basis for understanding directed RNA transport within the cytoplasm.
Asunto(s)
Cinesinas/metabolismo , Microtúbulos/metabolismo , Oocitos/metabolismo , Transporte de ARN/fisiología , ARN/metabolismo , Animales , Polaridad Celular , Cinesinas/genética , Microtúbulos/ultraestructura , Modelos Biológicos , Datos de Secuencia Molecular , Mutación , Oocitos/citología , Fenotipo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Secuencias Reguladoras de Ácido Ribonucleico , Xenopus laevis/genética , Xenopus laevis/metabolismoRESUMEN
The effect of a number of antibiotics on stop codon readthrough during protein synthesis in Escherichia coli was examined. Inhibitors which bind close to the entrance of the peptide exit tunnel on the 50S ribosomal subunit promote substantial levels of readthrough, presumably by disrupting the mechanism of peptide release.