Force production by actin polymerization motor at leading edge of locomoting cell

运动细胞前缘的肌动蛋白聚合马达产生力

基本信息

批准号：
18570154
负责人：
NAKAGAWA Hiroyuki
金额：
$ 2.47万
依托单位：
Fukuoka University (2007)Kyushu Institute of Technology (2006)
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2006
资助国家：
日本
起止时间：
2006 至 2007
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18570154/
关键词：
actin lasp-2 actin bundle fascin

项目摘要

Actin filaments are organized into sub-compartments of meshwork and bundles in lamellipodia. Polymerization of actin molecules into filament produces a mechanical force for cell movement. Organization of these filaments is through to be regulated by actin filament binding proteins. Localization of fascin, the LIM and SH3 domain protein 1 (lasp-1), and lasp-2 to the bundles suggest their involvement in that organization; however, their contributions remain unclear. We have compared the turnover of these proteins with actin at the bundle. After photobleaching, EGFP-actin recovered inwards from the bundle tip, consistent with the retrograde flow by treadmilling. In contrast, the recovery of EGFP-fascin, -lasp-1 and -lasp-2 occurred from the anterograde direction. These results suggest that these molecules would participate in the stabilization of bundles but not in initiation.Lasp-2 has been identified to have three domains : a LIM domain, nebulin-repeat domain and an SH3 domain in lasp-2 ; however, the region responsible for actin-binding is still unclear. We have expressed lasp-2 fragments tagged with EGFP in NG108-15 and C2C12 cells. We showed that the N-terminal fragment from the LIM domain to the first nebulin-repeat module retained actin-binding activity and a similar subcellular localization to full-length lasp-2, but the LIM domain fragment did not. Partial truncation of the LIM domain caused the loss of actin-binding activity and subcellular localization. These results suggest that lasp-2 interaction with F-actin is mediated by the cooperation of the LIM domain and first nebulin-repeat module both in vitro and in vivo.

肌动蛋白丝被组织成薄片中的网状和束子的子室。将肌动蛋白分子聚合到细丝中会产生细胞运动的机械力。这些细丝的组织将受肌动蛋白丝结合蛋白的调节。 Fascin，LIM和SH3域蛋白1（LASP-1）和LASP-2的定位表明它们参与了该组织。但是，他们的贡献尚不清楚。我们已经将这些蛋白质的营业额与丁基的肌动蛋白进行了比较。光漂白后，EGFP-肌动蛋白从束尖端向内恢复，与跑步机的逆行流程一致。相反，从顺行向发生EGFP-FASCIN，-LASP-1和-LASP-2的恢复。这些结果表明，这些分子将参与束的稳定，但不参与启动。LASP-2已确定为具有三个域：LIM结构域，Nebulin-Repeat结构域和LASP-2中的SH3域；但是，负责肌动蛋白结合的地区尚不清楚。我们已经表达了在NG108-15和C2C12细胞中用EGFP标记的LASP-2片段。我们表明，从LIM结构域到第一个Nebulin重复模块的N末端片段保留了肌动蛋白结合活性，并且与全长LASP-2相似的亚细胞定位，但LIM域片段没有。 LIM结构域的部分截断导致肌动蛋白结合活性和亚细胞定位的丧失。这些结果表明，LASP-2与F-肌动蛋白的相互作用是通过在体外和体内的LIM结构域和第一个Nebulin-Repeat模块的合作介导的。