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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亚隔室的网状结构和束在板状伪足。肌动蛋白分子聚合成细丝产生了细胞运动的机械力。这些纤维的组织通过肌动蛋白丝结合蛋白来调节。肌成束蛋白、LIM和SH 3结构域蛋白1（lasp-1）和lasp-2在束中的定位表明它们参与了该组织;然而，它们的贡献仍不清楚。我们比较了这些蛋白质的营业额与肌动蛋白在束。光漂白后，EGFP-肌动蛋白恢复向内从束尖，与逆行流通过螺旋铣削一致。相反，EGFP-fascin、-lasp-1和-lasp-2的恢复发生在顺行方向。Lasp-2具有3个结构域：LIM结构域、Nebulin-repeat结构域和SH 3结构域，但与肌动蛋白结合的区域尚不清楚。我们已经在NG 108 -15和C2 C12细胞中表达了带有EGFP标签的lasp-2片段。我们发现，从LIM结构域的第一个星云蛋白重复模块的N-末端片段保留肌动蛋白结合活性和类似的亚细胞定位全长lasp-2，但LIM结构域片段没有。LIM结构域的部分截短导致肌动蛋白结合活性和亚细胞定位的损失。这些结果表明，lasp-2与F-肌动蛋白的相互作用是由LIM结构域和第一个星云蛋白重复模块在体外和体内的合作介导的。