RhoA-binding kinase α translocation is facilitated by the collapse of the vimentin intermediate filament network
RhoA-binding kinase α translocation is facilitated by the collapse of the vimentin intermediate filament network
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DOI：
10.1128/mcb.18.11.6325
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发表时间：
1998-11-01
影响因子：
5.3
通讯作者：
Lim, L
中科院分区：
文献类型：
--
作者：
Sin, WC;Chen, XQ;Lim, L
真核细胞形态变化的调节是一个复杂的过程，涉及细胞骨架的主要组成部分，包括肌动蛋白微丝、微管和中间丝 (IF)。RhoA 的假定效应器，RhoA 结合激酶 α (ROK α)，是一种丝氨酸/苏氨酸激酶，与肌动蛋白丝的重组和肌球蛋白收缩性有关。在这里，我们表明 ROK α 也直接参与影响 IF 的结构完整性。活性 ROK α 的过度表达（如 RhoA 的过度表达）会导致丝状波形蛋白（III 型 IF）的崩溃。 RhoA 结合缺陷、激酶失活的 ROK α 可抑制 HeLa 细胞中 RhoA 诱导的波形蛋白 Ifs 的崩溃。在体外，ROK α 在其头杆结构域结合并磷酸化波形蛋白，从而抑制波形蛋白的组装，ROK α 主要与丝状波形蛋白网络共定位，该网络在血清饥饿的细胞中保持完整。用长春花碱（一种微管破坏剂）处理细胞也会导致丝状波形蛋白塌陷，并伴随 ROK α 易位至细胞外周。当长春花碱处理的细胞在 4℃ 下或存在显性失活 RhoAN19 突变体时，ROK α 易位不会发生。在热休克的细胞中也观察到 ROK α 的瞬时易位，这导致波形蛋白网络解体。因此，RhoAV14 过表达或生长因子处理后 ROK α 易位至细胞外周与波形蛋白 IF 的分解相关。这些结果表明已知作用于微丝的 Rho 效应子可能参与调节 IF 的组装。波形蛋白磷酸化后也表现出对非活性 ROK α 的亲和力降低。在激活的 RhoA 和 ROK α 的作用下，ROK α 从 Ifs 易位到细胞外周，表明 ROK α 可能启动其自身的级联激活。
The regulation of morphological changes in eukaryotic cells is a complex process involving major components of the cytoskeleton including actin microfilaments, microtubules, and intermediate filaments (IFs), The putative effector of RhoA, RhoA-binding kinase alpha (ROK alpha), is a serine/threonine kinase that has been implicated in the reorganization of actin filaments and in myosin contractility, Here, we show that ROK alpha also directly affects the structural integrity of IFs. Overexpression of active ROK alpha, like that of RhoA, caused the collapse of filamentous vimentin, a type III IF. A RhoA-binding-deficient, kinase-inactive ROK alpha inhibited the collapse of vimentin Ifs induced by RhoA in HeLa cells. In vitro, ROK alpha bound and phosphorylated vimentin at its head-rod domain, thereby inhibiting the assembly of vimentin, ROK alpha colocalized predominantly with the filamentous vimentin network, which remained intact in serum-starved cells. Treatment of cells with vinblastine, a microtubule-disrupting agent, also resulted in filamentous vimentin collapse and concomitant ROK alpha translocation to the cell periphery. ROK alpha translocation did not occur when the vimentin network remained intact in vinblastine-treated cells at 4 degrees C or in the presence of the dominant-negative RhoAN19 mutant, Transient translocation of ROK alpha was also observed in cells subjected to heat shock, which caused the disassembly of the vimentin network. Thus, the translocation of ROK alpha to the cell periphery upon overexpression of RhoAV14 or growth factor treatment is associated with disassembly of vimentin IFs. These results indicate that Rho effecters known to act on microfilaments may be involved in regulating the assembly of IFs. Vimentin when phosphorylated also exhibits reduced affinity for the inactive ROK alpha. The translocation of ROK alpha from Ifs to the cell periphery upon action by activated RhoA and ROK alpha suggests that ROK alpha may initiate its own cascade of activation.