Dept. of Veterinary Physiology, Chonnam National Univ., Gwangju, Korea.
Am J Physiol Cell Physiol. 2010 May;298(5):C1159-69. doi: 10.1152/ajpcell.00496.2009. Epub 2010 Jan 20.
Laminin is the first extracellular matrix (ECM) component to be expressed in the developing mammalian embryo. However, the roles of laminin or the related signal pathways are not well known in mouse embryonic stem cells (mESCs). Presently, we examined the effect of laminin on mESC migration. Laminin (10 microg/ml) decreased cell aggregation, whereas migration was increased. Laminin bound alpha6beta1 integrin and laminin receptor 1 (LR1), decreasing their mRNA levels. Laminin increased focal adhesion kinase (FAK) and paxillin phosphorylation, cAMP intracellular concentration, and the protein levels of exchange factor directly activated by cAMP (Epac1) and Rap1. These increases were completely blocked by alpha6beta1 integrin and LR1 neutralizing antibody, indicating that laminin-bound LR1 assists laminin-induced alpha6beta1 integrin activity and initiates signal. As a downstream signal molecule, laminin activated small G protein such as Rac1/cdc42 and its effector protein p21-activated kinase (PAK). Subsequently, laminin stimulated E-cadherin complex disruption. Inhibition of each pathway such as those for alpha6beta1 integrin and LR1, FAK, Rap1, and PAK1 blocked laminin-induced migration. We conclude that laminin binds both alpha6beta1 integrin and LR1 and induces signaling FAK/paxillin and cAMP/Epac1/Rap1. These signaling merge at Rac1/cdc42 subsequently activate PAK1. Activated PAK1 enhances E-cadherin complex disruption and finally increases mESCs migration.
层粘连蛋白是哺乳动物胚胎中最早表达的细胞外基质（ECM）成分之一。然而，层粘连蛋白或相关信号通路在小鼠胚胎干细胞（mESCs）中的作用尚不清楚。目前，我们研究了层粘连蛋白对 mESC 迁移的影响。层粘连蛋白（10μg/ml）降低了细胞聚集，而迁移增加。层粘连蛋白结合α6β1 整合素和层粘连蛋白受体 1（LR1），降低其 mRNA 水平。层粘连蛋白增加了粘着斑激酶（FAK）和桩蛋白的磷酸化、cAMP 细胞内浓度以及 cAMP 直接激活的交换因子（Epac1）和 Rap1 的蛋白水平。这些增加完全被α6β1 整合素和 LR1 中和抗体阻断，表明层粘连蛋白结合的 LR1 辅助层粘连蛋白诱导的α6β1 整合素活性并启动信号。作为下游信号分子，层粘连蛋白激活了小 G 蛋白，如 Rac1/cdc42 及其效应蛋白 p21 激活激酶（PAK）。随后，层粘连蛋白刺激 E-钙粘蛋白复合物解体。抑制α6β1 整合素和 LR1、FAK、Rap1 和 PAK1 等每条途径均可阻断层粘连蛋白诱导的迁移。我们得出结论，层粘连蛋白结合α6β1 整合素和 LR1 并诱导 FAK/paxillin 和 cAMP/Epac1/Rap1 信号。这些信号在 Rac1/cdc42 处融合，随后激活 PAK1。激活的 PAK1 增强 E-钙粘蛋白复合物解体，最终增加 mESCs 的迁移。
