Ligation of cell surface-associated glucose-regulated protein 78 by receptor-recognized forms of α2-macroglobulin:: Activation of p21-activated protein kinase-2-dependent signaling in murine peritoneal macrophages

Previous studies of the plasma proteinase inhibitor alpha(2)-niacroglobulin (alpha(2)M) demonstrated that alpha(2)M-proteinase complexes (alpha(2)M*) modulate immune responses and promotes macrophage locomotion and chemotaxis. a2M* binds to cell surface-associated glucoseregulated protein 78 (GRP78), which activates downstream signaling events. The role of p21-activated protein kinase-1 and -2 (PAK-1 and -2) in promoting cellular motility is well documented. In the current study, we examined the ability of alpha(2)M* to activate PAK-1 and PAK-2. Upon macrophage stimulation with alpha(2)M*, PAK-2 is autophosphorylated, resulting in increased kinase activity; however, PAK-1 is negligibly affected. alpha(2)M*-stimulated macrophages showed a marked elevation in the levels of Rac-GTP. Receptor tyrosine phosphorylation upon binding of a2M* to GRP78, recruits PAK-2 to the plasma membrane via the adaptor protein NCK. Consistent with this hypothesis, silencing of GRP78 gene expression greatly attenuated the levels of membrane-associated PAK-2 and NCK. PAK-2 activity was markedly decreased by inhibition of tyrosine kinases and PI3K before a2M* stimulation. We further demonstrate that phosphorylation of Lin-11, Isl-1, Mec-3 (LIM) kinase and cofilin is promoted by treating macrophages with alpha(2)M*. Thus, alpha(2)M* regulates activation of the PAK-2-dependent motility mechanism in these cells.