Functional interaction between Rho effector proteins

定向细胞迁移需要细胞极化和粘附转换，其中肌动蛋白细胞骨架和微管起关键作用。Rho gtpase诱导特定类型的肌动蛋白细胞骨架并调节微管动力学。在迁移细胞中，Cdc42调节细胞极性，Rac参与细胞膜突起。然而，Rho在迁移中的作用却鲜为人知。Rho作用于两个主要的效应器，ROCK和mDia1，其中mDia1产生直肌动蛋白丝并排列微管。我们发现Rho-mDia1通路通过排列微管和肌动蛋白丝并将Apc/Cdc42和c-Src传递到大鼠C6胶质瘤细胞各自的作用位点来调节极化和粘附转换。媒质蛋白属于双胍蛋白家族，催化肌动蛋白成核和聚合。虽然formin家族蛋白在果蝇等非哺乳动物的细胞质分裂中是必不可少的，但mDia蛋白是否以及如何在细胞质分裂中起作用尚不清楚。为了解决这个问题，我们通过RNA干扰去除NIH 3T3细胞中的三种培养基亚型。mDia2的缺失选择性地增加了双核细胞的数量。mDia2在分裂后期到末期在分裂沟中积累，在细胞质分裂末期集中在中间。mDia2的缺失在分裂细胞的异常部位诱导收缩，在那里收缩环成分如RhoA、肌球蛋白、苯胺和磷酸化ERM积累。blebbistatin抑制了异常收缩，纠正了上述成分的定位，并发现mDia2 RNAi显著降低了后期/末期赤道区F-actin的量。这些结果表明，mDia2在哺乳动物细胞分裂中是必不可少的，并且mDia2诱导的f -肌动蛋白为收缩环形成支架，并保持其在分裂细胞中间的位置。此外，我们还分析了另一种Rho效应分子ROCK在胚胎发育过程中血管发生中的作用。少

Directed cell migration requires cell polarization and adhesion turnover, in which the actin cytoskeleton and microtubules work critically. The Rho GTPases induce specific types of actin cytoskeleton and regulate microtubule dynamics. In migrating cells, Cdc42 regulates cell polarity and Rac works in membrane protrusion. However, the role of Rho in migration is little known. Rho acts on two major effectors, ROCK and mDia1, among which mDia1 produces straight actin filaments and aligns microtubules. We found that the Rho-mDia1 pathway regulates polarization and adhesion turnover by aligning microtubules and actin filaments and delivering Apc/Cdc42 and c-Src to their respective sites of action in rat C6 glioma cells.mDia proteins belong to the formin family proteins that catalyze actin nucleation and polymerization. Although formin family proteins of nonmammalian species such as Drosophila diaphanous are essential in cytokinesis, whether and how mDia proteins function in cytokinesis rema … More in unknown. To address this issue, we depleted each of the three mDia isoforms in NIH 3T3 cells by RNA interference. Depletion of mDia2 selectively increased the number of binucleate cells. mDia2 accumulates in the cleavage furrow during anaphase to telophase, and concentrates in the midbody at the end of cytokinesis. Depletion of mDia2 induced contraction at aberrant sites of dividing cells, where contractile ring components such as RhoA, myosin, anillin, and phosphorylated ERM accumulated. Treatment with blebbistatin suppressed abnormal contraction, corrected localization of the above components, and revealed that the amount of F-actin at the equatorial region during anaphase/telophase was significantly decreased with mDia2 RNAi. These results demonstrate that mDia2 is essential in mammalian cell cytokinesis and that mDia2-induced F-actin forms a scaffold for the contractile ring and maintains its position in the middle of a dividing cell.Furthermore, we are also analyzing the role of another Rho effector molecule, ROCK, in vasculogenesis during embryonic development. Less