Bijian Krikor, Takano Tomoko, Papillon Joan, Le Berre Ludmilla, Michaud Jean-Louis, Kennedy Chris R J, Cybulsky Andrey V
Department of Medicine, McGill University Health Centre, Montreal, Quebec, Canada.
Am J Physiol Renal Physiol. 2005 Dec;289(6):F1313-23. doi: 10.1152/ajprenal.00106.2005. Epub 2005 Jul 12.
Adhesion of rat glomerular epithelial cells (GEC) to collagen activates focal adhesion kinase (FAK) and the Ras-extracellular signal-regulated kinase (ERK) pathway and supports survival (prevents apoptosis). The present study addresses the relationship between actin organization and the survival phenotype. Parental GEC (adherent to collagen) and GEC stably transfected with constitutively active mutants of mitogen-activated protein kinase kinase (R4F-MEK) or FAK (CD2-FAK) (on plastic) showed ERK activation, low levels of apoptosis, and a cortical distribution of F-actin. Parental GEC adherent to plastic showed increased apoptosis, disorganization of cortical F-actin, and formation of prominent stress fibers. Assembly of cortical F-actin was, at least in part, mediated via ERK. However, disruption of the actin cytoskeleton with cytochalasin D or latrunculin B in parental GEC (on collagen) and in GEC that express R4F-MEK or CD2-FAK (on plastic) decreased ERK activation and increased apoptosis. Expression of a constitutively active RhoA (L(63)RhoA) induced assembly of cortical F-actin, promoted ERK activation, and supplanted the requirement of collagen for survival. Adhesion of GEC to collagen increased phosphatidylinositol-4,5-bisphosphate (PIP(2)). Downregulation or sequestration of PIP(2) by transfection with an inositol 5'-phosphatase or the plextrin-homology domain of phospholipase C-delta1 decreased F-actin content and survival. Moreover, overexpression of wild-type or K256E mutant alpha-actinin-4 with increased affinity for F-actin increased apoptosis. These results demonstrate a reciprocal relationship between collagen-induced cortical F-actin assembly and collagen-dependent survival signaling, including ERK activation. Appropriate remodeling of the actin cytoskeleton may be necessary for facilitating survival, as both disassembly and excessive crosslinking affect survival adversely.
大鼠肾小球上皮细胞（GEC）与胶原蛋白的黏附激活了黏着斑激酶（FAK）和Ras-细胞外信号调节激酶（ERK）通路，并支持细胞存活（防止细胞凋亡）。本研究探讨肌动蛋白组织与存活表型之间的关系。亲代GEC（黏附于胶原蛋白）以及稳定转染有丝裂原活化蛋白激酶激酶（R4F-MEK）或FAK（CD2-FAK）组成型活性突变体的GEC（在塑料上）显示出ERK激活、低水平凋亡以及F-肌动蛋白的皮质分布。黏附于塑料的亲代GEC显示出凋亡增加、皮质F-肌动蛋白紊乱以及明显应力纤维的形成。皮质F-肌动蛋白的组装至少部分是通过ERK介导的。然而，用细胞松弛素D或拉春库林B破坏亲代GEC（在胶原蛋白上）以及表达R4F-MEK或CD2-FAK的GEC（在塑料上）中的肌动蛋白细胞骨架，会降低ERK激活并增加凋亡。组成型活性RhoA（L(63)RhoA）的表达诱导了皮质F-肌动蛋白的组装，促进了ERK激活，并取代了胶原蛋白对存活的需求。GEC与胶原蛋白的黏附增加了磷脂酰肌醇-4,5-二磷酸（PIP(2)）。通过转染肌醇5'-磷酸酶或磷脂酶C-δ1的磷酯酰肌醇结合同源结构域来下调或隔离PIP( )会降低F-肌动蛋白含量和细胞存活。此外，对F-肌动蛋白亲和力增加的野生型或K256E突变体α-辅肌动蛋白-4的过表达会增加凋亡。这些结果表明胶原蛋白诱导的皮质F-肌动蛋白组装与包括ERK激活在内的胶原蛋白依赖性存活信号之间存在相互关系。肌动蛋白细胞骨架的适当重塑可能是促进存活所必需的，因为拆卸和过度交联都会对存活产生不利影响。
