CaMKIIδ splice variants differentially regulate vascular smooth muscle cell motility

Vascular smooth muscle (VSM) cell migration promotes physiological and pathophysiological vascular remodeling. A previous study from our laboratory demonstrated that both CaMKIIδ and FYN, a SRC- family kinase, promote VSM cell migration in vitro. CaMKIIδ co-localized with FYN and physically interacted in an activation-dependent manner, with CaMKIIδ activation resulting in FYN activation and dissociation of the interaction. The CAMK2D gene undergoes extensive alternative splicing, including an exon encoding a 21-amino acid C-terminus, resulting in the expression of CaMKIIδ₂ and related variants that include the 21-AA C-terminal tail, and CaMKIIδ₆ and related variants lacking the sequence. The aforementioned CaMKIIδ/FYN interaction was attributed to the CaMKIIδ₂ variant that predominates in rat aortic VSM cells used in those studies. The purpose of this study was to test the functional equivalency of CaMKIIδ₂ and CaMKIIδ₆ isoforms in mediating signal transduction required for VSM motility. Adenoviral overexpression of CaMKIIδ₆ inhibited VSM cell motility, disrupted CaMKIIδ/FYN interaction_, reduced localization of active CaMKIIδ in lamellipodia, and reduced PDGF-dependent tyrosine phosphorylation of the focal adhesion proteins, P130^Cas and paxillin mediated by FYN. Silencing FYN expression using siRNA phenocopied these effects of over-expressing CaMKIIδ₆. As an alternative approach, application of an exon skipping antisense oligonucleotide targeting the C-terminal exon in the Camk2d gene, resulted in loss of CaMKIIδ₂ expression and a reciprocal increase in expression of CaMKIIδ₆. This manipulation of endogenous CaMKIIδ variant expression also inhibited localization of active CaMKIIδ in lamellipodia and inhibited VSM cell migration. Taken together, these results suggest that the alternatively spliced C-terminal domain in the CaMKIIδ₂ splice variant confers interaction with FYN, localization of the kinases to lamellipodia, and consequent regulation of focal adhesion dynamics to promote VSM cell migration.