Modulation of vascular smooth muscle cell migration by calcium/calmodulin-dependent protein kinase II-δ2

Previous studies demonstrated a requirement for multifunctional Ca2+/calmodulin-dependent protein kinase II (CaMKII) in PDGF-stimulated vascular smooth muscle (VSM) cell migration. In the present study, molecular approaches were used specifically to assess the role of the predominant CaMKII isoform (delta(2) or delta(C)) on VSM cell migration. Kinase-negative (K43A) and constitutively active (T287D) mutant forms of CaMKIIdelta(2) were expressed using recombinant adenoviruses. CaMKII activities were evaluated in vitro by using a peptide substrate and in intact cells by assessing the phosphorylation of overexpressed phospholamban on Thr(17), a CaMKII-selective phosphorylation site. Expression of kinase-negative CaMKIIdelta(2) inhibited substrate phosphorylation both in vitro and in the intact cell, indicating a dominant-negative function with respect to exogenous substrate. However, overexpression of the kinase-negative mutant failed to inhibit endogenous CaMKIIdelta(2) autophosphorylation on Thr(287) after activation of cells with ionomycin, and in fact, these subunits served as a substrate for the endogenous kinase. Constitutively active CaMKIIdelta(2) phosphorylated substrate in vitro without added Ca2+/calmodulin and in the intact cell without added Ca2+-dependent stimuli, but it inhibited autophosphorylation of endogenous CaMKIIdelta(2) on Thr(287). Basal and PDGF-stimulated cell migration was significantly enhanced in cells expressing kinase-negative CaMKIIdelta(2), an effect opposite that of KN-93, a chemical inhibitor of CaMKII activation. Expression of the constitutively active CaMKIIdelta(2) mutant inhibited PDGF-stimulated cell migration. These studies point to a role for the CaMKIIdelta(2) isoform in regulating VSM cell migration. An inclusive interpretation of results using both pharmacological and molecular approaches raises the hypothesis that CaMKIIdelta(2) autophosphorylation may play an important role in PDGF-stimulated VSM cell migration.