ゼブラフィッシュ卵母細胞におけるサイクリンB1 mRNAの翻訳と局在を制御する機構

[Chapter I]In immature zebrafish oocytes, dormant cyclin B1mRNAslocalizetothe animalpolar cytoplasmas aggregates.After hormonal stimulation, cyclin B1mRNAs are dispersed and translationally activated, which are necessary and sufficient for the induction ofzebrafish oocyte maturation.Besides CPE and CPEB, Pum1 anda cis-acting element in the coding region of cyclin B1mRNAare important for the subcellular localization and timing of translational activationof the mRNA.However, mechanisms underlying the spatio-temporal control of cyclin B1mRNA translation during oocyte maturation are not fully understood. Ireport thatIMP3, which was initially described as a protein bound tovg1mRNAlocalizedto the vegetal pole of Xenopus oocytes,bindstoand co-localizes with cyclin B1mRNAin immature zebrafish oocytes. Although protein content and phosphorylation state of IMP3 remain unchanged during oocyte maturation, IMP3 dissociates fromcyclin B1mRNA after hormonal stimulation.In accordance with this,themRNA-dependent interactionsof IMP3 with Pum1 and CPEB in immature oocyteswere disruptedin mature oocytes. The overexpressed IMP3 proteins were accumulated at the animal polar cytoplasmand caused delay in the polyadenylation of cyclin B1mRNA, synthesis of Cyclin B1 proteinand progression of oocyte maturation. On the basis of these results,Ipropose that IMP3 repressesthe translationof cyclin B1mRNA in immature zebrafish oocytesand that its release from the mRNA triggers the translational activation. [Chapter II] Precise localization of maternal mRNAs in oocytes is critical for oocyte maturation and embryonic development. In full-grown zebrafish oocytes, dormant cyclin B1 mRNAs are localized to the animal polar cytoplasm and translationally activated during oocyte maturation. To elucidate the molecular basis of cyclin B1 mRNA localization, I analyzed zebrafish Staufen1, a protein known to play a central role in mRNA localization in various cells. Zebrafish Staufen1 interacts with cyclin B1 mRNA throughout oocyte growth and maturation. Both cyclin B1 mRNA and Staufen1 protein are evenly distributed in the cytoplasm of young oocytes but are co-localized to the animal polar cytoplasm in later stages. In mature oocytes, cyclin B1 mRNA is dispersed throughout the cytoplasm, while a subpopulation of Staufen1 remains at the animal pole. Transport of cyclin B1 reporter mRNA to the animal pole was inhibited by disruption of microtubules and injection of antibodies against Staufen1 or Kinesin1, a plus-end-directed microtubule motor that binds to Staufen1, indicating that the transport depends on movement along microtubules toward the plus ends. The plus ends of microtubules are enriched in the animal polar cytoplasm of zebrafish oocytes, contrary to Xenopus oocytes, in which they are enriched in the vegetal polar cytoplasm. Reporter mRNAs with an element required for the vegetal localization of vg1 mRNA in Xenopus oocytes were localized to the animal polar cytoplasm in zebrafish oocytes. These findings indicate that a common mRNA transport machinery functions in zebrafish and Xenopus oocytes but that its transport direction is opposite probably due to different organizations of microtubules in oocytes