Regulation of axonal extension and growth cone motility by calmodulin-dependent protein kinase I

Calcium and calmodulin (CaM) are important signaling molecules that regulate axonal or dendritic extension and branching. The Ca2+-dependent stimulation of neurite elongation has generally been assumed to be mediated by CaM-kinase II (CaMKII), although other members of the CaMK family are highly expressed in developing neurons. We have examined this assumption using a combination of dominant-negative CaMKs (dnCaMKs) and other specific CaMK inhibitors. Here we report that inhibition of cytosolic CaMKI, but not CaMKII or nuclear CaMKIV, dramatically decreases axonal outgrowth and branching in cultured neonatal hippocampal and postnatal cerebellar granule neurons. CaMKI is found throughout the cell cytosol, including the growth cone. Growth cones of neurons expressing dnCaMI or dnCaMKK, the upstream activator of CaMKI, exhibit collapsed morphology with a prominent reduction in lamellipodia. Live-cell imaging confirms that these morphological changes are associated with a dramatic decrease in growth cone motility. Treatment of neurons with 1,8-naphthoylene benzimidazole-3-carboxylic acid (STO-609), an inhibitor of CaMKK, causes a similar change in morphology and reduction in growth cone motility, and this inhibition can be rescued by transfection with an STO-609-insensitive mutant of CaMKK or by transfection with constitutively active CaMKI. These results identify CaMKI as a positive transducer of growth cone motility and axon outgrowth and provide a new physiological role for the CaMKK-CaMKI pathway.