Analysis of Intermediate Filament Binding Proteins

中间丝结合蛋白的分析

基本信息

批准号：
13480241
负责人：
INAGAKI Masaki
金额：
$ 9.66万
依托单位：
Aichi Cancer Center
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2002
项目状态：
已结题

项目摘要

1. We identified Aurora-B as a cleavage furrow (CF) kinase. Aurora-B phosphorylates type III intermediate filaments (Ifs), vimentin, GFAP, and desmin, and the phosphorylation causes reduction of their filament forming ability. We next determined the phosphorylation sites of these Ifs by Aurora-B. IF mutants, in which phosphorylation sites by Aurora-B and/or Rho-kinase are changed to Ala or Gly, cause dramatic defect of filament separation between daughter cells in cytokinesis. These results indicate that Aurora-B coordinately regulates cytokinesis with Rho-kinase through the cleavage furrow-specific phosphorylation of type III Ifs.2. We identified human TNF receptor 1 (TNFR1)-associated death domain protein (TRADD) to be the keratin 18 (K18)-interacting protein. Endogenous TRADD coimmunoprecipitated with K18, and colocalized with K8/18 filaments in human mammary epithelial cells. Overexpression of the NH_2- terminus (aa 1-270) of K18 containing the TRADD-binding domain as well as overexpression of K8/18 in SW13 cells, which are devoid of keratins, rendered the cells more resistant to killing by TNF. These results suggest that K18 may sequester TRADD to attenuate interactions of TRADD with activated TNFR1 and moderate TNF-induced apoptosis in simple epithelial cells.3. We isolated δ-catenin/neural plakophilin-related armadillo-repeat protein (NPRAP) as a Densin- 180-interacting protein. Densin-180 is associated in vivo with δ-catenin/NPRAP and may be involved in organization of the synaptic cell-cell junction through interaction with theδ-catenin/NPRAP-N- cadherin complex.

1。我们将Aurora-B确定为裂解沟（CF）激酶。 Aurora-b磷酸化III型中间丝（IFS），波形蛋白，GFAP和Desmin，磷酸化导致其丝形成能力的降低。接下来，我们确定了Aurora-B的这些IFS的磷酸化位点。如果突变体（通过Aurora-B和/或Rho-kinase的磷酸化位点更改为ALA或GLY，则会导致细胞动力学中子细胞之间细丝分离的巨大缺陷。这些结果表明，Aurora-B协同通过Rho-kinase通过裂解III型的裂解沟特异性磷酸化来调节细胞因子。2。我们将人类TNF受体1（TNFR1）相关的死亡结构域蛋白（TRADD）确定为角蛋白18（K18）互动蛋白。内源性传统与K18共免疫沉淀，并与人类乳腺上皮细胞中的K8/18丝共定位。含有传统结合域的K18的NH_2-末端（AA 1-270）的过表达，以及没有角蛋白的SW13细胞中K8/18的过表达，这使细胞更能抵抗TNF杀死的细胞。这些结果表明，K18可能会隔离传统，以减轻传统与激活的TNFR1和中等TNF诱导的简单上皮细胞中凋亡的相互作用。3。我们将δ-catenin/神经plakophilin相关的腋窝重复蛋白（NPRAP）分离为densin-180相互作用蛋白。 densin-180与δ-catenin/nprap在体内相关联，并且可能通过与Δ-catenin/nprap-n-nprap-n-钙粘蛋白复合物相互作用而参与合成细胞 - 细胞连接的组织。