RESUMO
Given the prominence and the biological importance of the nucleus it is remarkable how little is still known about structure-forming proteins in the nuclear interior. The karyoskeletal protein NO145 has been identified as a major constituent of a filamentous network surrounding the amplified nucleoli of Xenopus laevis oocytes. We now show that an orthologous protein also occurs in female germ cells of a wide range of other vertebrates, where it forms dot-like structures. Using the Xenopus oocyte system we further report a specific regulatory mechanism responsible for (1) the rapid degradation of the NO145 protein during meiotic maturation, and (2) the cell-type-dependent translation of NO145 mRNA. Microinjection experiments have revealed that NO145 is a target of proteasomes and the use of the rapid amplification of cDNA ends-polyadenylation test (RACE-PAT) has disclosed the existence of NO145 mRNAs differing in their 3' UTRs. Reporter systems as well as polyribosome profiling experiments have revealed the regulatory importance of the 3' UTRs, which affect the translational efficiency as well as the stability of the encoded protein. The highly conserved cell-type specificity and the extremely tight temporal regulation of NO145 synthesis suggest an important role of this protein in female meiotic prophase.
Assuntos
Proteínas do Citoesqueleto/biossíntese , Oócitos/metabolismo , Proteínas de Xenopus/biossíntese , Regiões 3' não Traduzidas , Sequência de Aminoácidos , Animais , Sequência de Bases , Proteínas do Citoesqueleto/química , Proteínas do Citoesqueleto/genética , Primers do DNA , Genes Reporter , Dados de Sequência Molecular , Biossíntese de Proteínas , RNA Mensageiro/genética , Homologia de Sequência de Aminoácidos , Proteínas de Xenopus/química , Proteínas de Xenopus/genética , Xenopus laevisRESUMO
We have developed a method to produce live somatic clones in the rabbit, one of the mammalian species considered up to now as difficult to clone. To do so, we have modified current cloning protocols proven successful in other species by taking into account both the rapid kinetics of the cell cycle of rabbit embryos and the narrow window of time for their implantation after transfer into foster recipients. Although our method still has a low level of efficiency, it has produced several clones now proven to be fertile. Our work indicates that cloning can probably be carried out successfully in any mammalian species by taking into account physiological features of their oocytes and embryos. Our results will contribute to extending the use of rabbit models for biomedical research.
