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, P130Cas 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.