Wittmann Torsten, Bokoch Gary M, Waterman-Storer Clare M
Department of Cell Biology and Institute for Childhood and Neglected Diseases, La Jolla, CA 92037, USA.
J Cell Biol. 2003 Jun 9;161(5):845-51. doi: 10.1083/jcb.200303082.
Actin in migrating cells is regulated by Rho GTPases. However, Rho proteins might also affect microtubules (MTs). Here, we used time-lapse microscopy of PtK1 cells to examine MT regulation downstream of Rac1. In these cells, "pioneer" MTs growing into leading-edge protrusions exhibited a decreased catastrophe frequency and an increased time in growth as compared with MTs further from the leading edge. Constitutively active Rac1(Q61L) promoted pioneer behavior in most MTs, whereas dominant-negative Rac1(T17N) eliminated pioneer MTs, indicating that Rac1 is a regulator of MT dynamics in vivo. Rac1(Q61L) also enhanced MT turnover through stimulation of MT retrograde flow and breakage. Inhibition of p21-activated kinases (Paks), downstream effectors of Rac1, inhibited Rac1(Q61L)-induced MT growth and retrograde flow. In addition, Rac1(Q61L) promoted lamellipodial actin polymerization and Pak-dependent retrograde flow. Together, these results indicate coordinated regulation of the two cytoskeletal systems in the leading edge of migrating cells.
迁移细胞中的肌动蛋白受Rho GTP酶调节。然而，Rho蛋白也可能影响微管（MTs）。在这里，我们使用PtK1细胞的延时显微镜来检查Rac1下游的微管调节。在这些细胞中，与远离前沿的微管相比，生长到前沿突起中的“先锋”微管表现出较低的灾难频率和较长的生长时间。组成型活性Rac1（Q61L）在大多数微管中促进先锋行为，而显性负性Rac1（T17N）消除先锋微管，表明Rac1是体内微管动力学的调节因子。Rac1（Q61L）还通过刺激微管逆行流动和断裂来增强微管周转。抑制Rac1的下游效应器p21激活激酶（Paks）可抑制Rac1（Q61L）诱导的微管生长和逆行流动。此外，Rac1（Q61L）促进片状伪足肌动蛋白聚合和依赖Pak的逆行流动。总之，这些结果表明迁移细胞前沿的两个细胞骨架系统存在协同调节。
