SEPTIN 2在爪蟾卵母细胞发育过程中的作用研究

Cytokinesis is one of the key steps in the late cell division process. Septin has been indicated to play conservative and key roles in cytokinesis in a number of species except for vegetal cells. The molecular characteristic of septin indicates its roles in cytokinesis: molecular scaffold makes cytokinesis related proteins ,also with their regulating ones, to locate and gather in the cell division furrow; And the molecular barrier function ensures the formation of the contractile ring and contraction as well as finally complete cytoplasmic division. Cytokinesis is affected by many kinds of signal regulations, Cdc42 / PAK is one of the main regulating pathways during cytokinesis. Inhibition of Cdc42 or PAK causes cytokinesis failure. Researches showed that septin is closely related to Cdc42 / PAK signal pathway in the process of cytokinesis, but the regulating mechanism of Cdc42 / PAK/septin in cytokinesis is not clear. The characteristics of in vitro reconstruction of Xenopus septin2 protein suggests that its potential role in cytokinesis. This research uses Xenopus oocyte polarbody formation process as a model, exploring laser confocal microscope and Time-lapes living observation as the main technology, combined with microscopic injection, to study the roles of septin2 in cytokinesis of Xenopus oocyte. The potential function of molecular barrier septin2 exerts is the main part and the regulating relationship of Cdc42 / PAK/septin will be elucided in this study.

胞质分裂是细胞分裂末期的关键步骤之一。Septin蛋白在细胞胞质分裂中有保守而关键作用。Septin分子特性赋予其在细胞胞质分裂中的作用：分子绞架作用使参与胞质分裂及相关调控蛋白定位聚集于细胞分裂沟；分子壁垒作用及力学性质确保细胞收缩环形成、收缩和胞质分裂完成。Cdc42/PAK通路是调控胞质分裂主要途径之一，抑制Cdc42或PAK均导致细胞胞质分裂失败。Septin在胞质分裂中的作用与Cdc42/PAK信号通路密切相关，但Cdc42/PAK/septin在细胞胞质分裂中的调控机制尚不清楚。重组爪蟾septin2蛋白特点提示其在胞质分裂中有潜在作用。本课题以爪蟾卵母细胞极体形成过程为模型，Time-lapes活体观察为主要技术，探讨爪蟾septin2在爪蟾卵母细胞胞质分裂中作用，重点研究septin2分子壁垒作用及其与Cdc42、PAK2激酶在爪蟾卵母细胞的胞质分裂中可能存在的调控机制。

结题摘要.细胞分裂包括核分裂和胞质分裂（cytokinesis）。胞质分裂受多种信号途径在时间和空间上精密调控。研究表明septin在参与胞质分裂上有相对保守的的作用。septin 定位与绞架作用、septin与细胞收缩环的主要骨架蛋白相互作用、septin分子壁垒作用是胞质分裂的关键。卵母细胞是研究细胞骨架和细胞周期调控机制的理想模型，其极体（polar body）形成是细胞极化的过程，伴随细胞收缩环的形成和胞质分裂的发生。爪蟾卵母细胞通过极体的形成来完成减数分裂，以确保遗传稳定性。而胞质分裂是的正常发生是极体形成的关键。这一减数分裂过程经历生发泡期(GV)期卵母细胞在外界条件刺激下进入胚泡破裂期（GVBD）,之后进入核分裂及胞质分裂，最后形成并释放极体。本研究将初步探讨Septin2是否在GVBD及之后的极体形成率及形成时间上具有调节作用，为进一步研究其在胞质分裂中时空上的的调控机制奠定基础。结果：实验组（septin2-siRNA）爪蟾卵母细胞GVBD形成率（10/187，5.3%）与对照组（H2O）卵母细胞GVBD形成率（8/190，4.2%）无显著性差异（P=0.622）；实验组（septin2-siRNA）爪蟾卵母细胞GVBD形成时间分布与对照组（H2O）卵母细胞GVBD形成时间分布无显著性差异（P=0.969）；实验组（septin2-siRNA）爪蟾卵母细胞极体形成率（14/98，14.2%）与对照组（H2O）卵母细胞极体形成率（104/116，89.7%）有显著性差异（P<0.001）。实验组（septin2-siRNA）与对照组（H2O）卵母细胞极体形成时间分布有显著性差异（P<0.01）。.结论：septin2-siRNA对爪蟾卵母细胞GVBD形成率及形成时间无明显影响；septin2-siRNA干涉导致爪蟾卵母细胞极体形成率下降，极体形成时间滞后。