Villin function in the organization of the actin cytoskeleton -: Correlation of in vivo effects to its biochemical activities in vitro
Villin function in the organization of the actin cytoskeleton -: Correlation of in vivo effects to its biochemical activities in vitro
复制标题
DOI：
10.1074/jbc.274.38.26751
复制
发表时间：
1999-09-17
影响因子：
4.8
通讯作者：
Vandekerckhove, J
中科院分区：
文献类型：
--
作者：
Friederich, E;Vancompernolle, K;Vandekerckhove, J
绒毛蛋白是肠刷状缘的肌动蛋白结合蛋白，其在体外以Ca 2+依赖的方式使肌动蛋白成束、成核、帽化和切断。绒毛蛋白诱导转染细胞中微绒毛的生长，这一活性需要羧基末端定位的KKEK基序。通过结合细胞转染和生化检测，我们表明，绒毛蛋白诱导细胞中微绒毛生长的能力与其在体外捆绑F-肌动蛋白的能力相关，但与其成核活性无关。与其在细胞中对微丝成束的重要性一致，羧基末端F-肌动蛋白结合位点的KKEK基序对于体外成束是至关重要的。此外，位于绒毛蛋白氨基末端部分的第二个位点中的碱性残基的取代损害了其在细胞中的活性，并且在缺乏Ca 2+的情况下降低了其与F-肌动蛋白的结合以及其体外成束和切断活性。总之，这些研究结果表明，绒毛参与组织和稳定的刷状缘核心束，但不启动其组装成核的肌动蛋白丝。
Villin is an actin-binding protein of the intestinal brush border that bundles, nucleates, caps, and severs actin in a Ca2+-dependent manner in vitro, Villin induces the growth of microvilli in transfected cells, an activity that requires a carboxyl-terminally located KKEK motif. By combining cell transfection and biochemical assays, we show that the capacity of villin to induce growth of microvilli in cells correlates with its ability to bundle F-actin in vitro but not with its nucleating activity. In agreement with its importance for microfilament bundling in cells, the KKEK motif of the carboxyl-terminal F-actin-binding site is crucial for bundling in vitro, In addition, substitutions of basic residues in a second site, located in the amino-terminal portion of villin, impaired its activity in cells and reduced its binding to F-actin in the absence of Ca2+ as well as its bundling and severing activities in vitro. Altogether, these findings suggest that villin participates in the organization and stabilization of the brush border core bundle but does not initiate its assembly by nucleation of actin filaments.