Membrane ruffling requires coordination between type Iα phosphatidylinositol phosphate kinase and Rac signaling
Membrane ruffling requires coordination between type Iα phosphatidylinositol phosphate kinase and Rac signaling
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DOI：
10.1074/jbc.m211397200
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发表时间：
2003-06-20
影响因子：
4.8
通讯作者：
Anderson, RA
中科院分区：
文献类型：
--
作者：
Doughman, RL;Firestone, AJ;Anderson, RA
膜褶皱的形成需要皮质肌动蛋白细丝的重塑，这一过程依赖于小G蛋白RAC。生长因子通过整合调节肌动蛋白结合蛋白的信号通路来刺激肌动蛋白重塑和膜褶皱。磷脂酰肌醇4，5-二磷酸(PIP2)调节多种肌动蛋白结合蛋白的活性，由I型磷脂酰肌醇磷酸酶(PIPKI)产生。在这里，我们显示在MG-63细胞中，只有PIPKIpha亚型定位于血小板衍生生长因子(PDGF)诱导的膜褶皱。此外，作为显性负性突变体的激酶Dead PIPKIpha的表达阻止了膜的褶皱，表明PIPKIpha和PIP2参与了褶皱的形成。为了探索这一点，PIPKIpha在血清饥饿的细胞中过表达，并被PDGF刺激。在血清饥饿的细胞中，PIPKIpha的表达不会刺激肌动蛋白的重塑，但当PDGF刺激这些细胞时，肌动蛋白迅速重组成斑点，但不会出现膜褶皱。PIPKIpha介导的肌动蛋白焦点的形成不依赖于rac1和磷脂酰肌醇3-激酶的活性。值得注意的是，在PDGF刺激的细胞中，显性活性的rac1和PIPKIpha的共表达导致了细胞膜的褶皱。PDGF和rac1刺激的褶皱被蛋白激酶死亡的PIPKIpha的表达抑制。综合考虑，这些数据支持一个模型，在该模型中，PIPKIpha局部产生PIP2是肌动蛋白重塑所必需的，而膜褶皱的形成需要RAC信号。
Membrane ruffle formation requires remodeling of cortical actin filaments, a process dependent upon the small G-protein Rac. Growth factors stimulate actin remodeling and membrane ruffling by integration of signaling pathways that regulate actin-binding proteins. Phosphatidylinositol 4,5-bisphosphate (PIP2) regulates the activity of many actin-binding proteins and is produced by the type I phosphatidylinositol phosphate kinases (PIPKIs). Here we show in MG-63 cells that only the PIPKIalpha isoform is localized to platelet-derived growth factor (PDGF)-induced membrane ruffles. Further, expression of kinase dead PIPKIalpha, which acts as a dominant negative mutant, blocked membrane ruffling, suggesting that PIPKIalpha and PIP2 participate in ruffling. To explore this, PIPKIalpha was overexpressed in serum-starved cells and stimulated with PDGF. In serum-starved cells, PIPKIalpha expression did not stimulate actin remodeling, but when these cells were stimulated with PDGF, actin rapidly reorganized into foci but not membrane ruffles. PIPKIalpha-mediated formation of actin foci was independent of both Rac1 and phosphatidylinositol 3-kinase activities. Significantly, coexpression of dominant active Rac1 with PIPKIalpha in PDGF-stimulated cells resulted in membrane ruffling. The PDGF- and Rac1-stimulated ruffling was inhibited by expression of kinase-dead PIPKIalpha. Combined, these data support a model where the localized production of PIP2 by PIPKIalpha is necessary for actin remodeling, whereas formation of membrane ruffles required Rac signaling.