Dissection of formin pathways coordinating polarity with cell migration downstream of Ras signalling

福尔明通路的剖析，协调极性与 Ras 信号下游细胞迁移

• 批准号: 169554986
• 负责人: Professor Dr. Jan Faix
• 金额: --
• 依托单位: Institut für Biophysikalische Chemie
• 依托单位国家: 德国
• 项目类别: Priority Programmes
• 财政年份: 2010
• 资助国家: 德国
• 起止时间: 2009-12-31 至 2016-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/169554986?language=en
• 关键词: Dissection; formin; pathways; coordinating; polarity

Diaphanous-related formins constitute a conserved family of ubiquitous multidomain proteins that catalyze the de novo nucleation of linear actin filaments and act as downstream effectors of Rho GTPase signaling pathways to control numerous cellular functions including the establishment of cell shape or cell migration. Our ongoing analysis of the 10 Dictyostelium formins unexpectedly revealed that the elimination of the regulated formin dDia1 strongly accelerated cell motility, whereas migration of dDia1-overexpressing cells was significantly suppressed. As assessed by imaging of an active dDia1 variant, the formin accumulated in a thin layer around the plasma membrane and was most strongly enriched in vesicular structures at the cell cortex. The aim of this project is to characterize these highly dynamic structures by epifluorescence and correlative electron microscopy and find out how exactly dDia1 brings about it unexpected negative effect on cell migration. In vivo formins seem to strictly depend on the recruitment of profilin-actin complexes by the FH1-domain for further assembly by the FH2 2 domain. Therefore, we also intend to thoroughly study actin polymerization by Dictyostelium ForC, because it is the only known formin lacking the FH1-region. As ForC is also devoid of other canonical G-actin binding sites, the second project aims to identify and characterize possible mechanisms by which this formin accomplishes to recruit actin subunits for subsequent filament assembly. We expect that both projects will greatly add to our knowledge of formin-mediated actin assembly.

透明相关的formin构成了一个保守的家族普遍存在的多结构域蛋白，催化线性肌动蛋白丝的从头成核，并作为Rho GT3信号通路的下游效应物，以控制许多细胞功能，包括建立细胞形状或细胞迁移。我们正在进行的10个Dictyosteoblasts formins的分析出乎意料地发现，消除调节的dDia 1强烈加速细胞的运动，而迁移dDia 1过表达的细胞被显着抑制。通过对活性dDia 1变体进行成像评估，发现dDia 1在质膜周围的薄层中积累，并且在细胞皮层的囊泡结构中最强烈地富集。该项目的目的是通过落射荧光和相关电子显微镜来表征这些高度动态的结构，并找出dDia 1究竟如何对细胞迁移产生意想不到的负面影响。在体内formin似乎严格依赖于招募的profilin-actin复合物的FH 1-域进一步组装的FH 2 - 2域。因此，我们也打算彻底研究肌动蛋白聚合的网骨藻ForC，因为它是唯一已知的缺乏FH 1-区域的酶。由于ForC也缺乏其他典型的G-肌动蛋白结合位点，第二个项目的目的是确定和表征可能的机制，通过该机制，该酶完成招募肌动蛋白亚基用于随后的细丝组装。我们希望这两个项目将大大增加我们的知识，福尔明介导的肌动蛋白组装。