PKCε, via its regulatory domain and independently of its catalytic domain, induces neurite-like processes in neuroblastoma cells

To investigate the role of protein kinase C (PKC) isoforms in regulation of neurite outgrowth, PKC alpha, beta II, delta, and epsilon fused to enhanced green fluorescent protein (EGFP) were transiently overexpressed in neuroblastoma cells. Overexpression of PKC epsilon-EGFP induced cell processes whereas the other isoforms did not. The effect of PKC epsilon-EGFP was not suppressed by the PKC inhibitor GF109203X. Instead, process formation was more pronounced when the regulatory domain was introduced. Overexpression of various fragments from PKC epsilon regulatory domain revealed that a region encompassing the pseudosubstrate, the two C1 domains, and Darts of the V3 region were necessary and sufficient for induction of processes, By deleting the second C1 domain from this construct, a dominant-negative protein was generated which suppressed processes induced by full-length PKC epsilon and neurites induced during retinoic acid- and growth factor-induced differentiation. As with neurites in differentiated neuroblastoma cells, processes induced by the PKC epsilon-PSC1V3 protein contained alpha-tubulin, neurofilament-160, and F-actin, but the PKC epsilon-PSC1V3-induced processes lacked the synaptic markers synaptophysin and neuropeptide Y. These data suggest that PKC epsilon, through its regulatory domain, can induce immature neurite-like processes via a mechanism that appears to be of importance for neurite outgrowth during neuronal differentiation.