Nerve growth factor stimulates the interaction of ZIP/p62 with atypical protein kinase C and targets endosomal localization: evidence for regulation of nerve growth factor-induced differentiation

Atypical protein kinase Cs zeta and lambda/iota play a functional role in the regulation of NGF-induced differentiation and survival of pheochromocytoma, PC12 cells [Coleman and Wooten, 1994; Wooten et al., 1999]. Here we demonstrate an NGF-dependent interaction of aPKC with its binding protein, ZIP/p62. Although, ZIP/p62 was not a PKC-iota substrate, the formation of a ZIP/p62-aPKC complex in PC12 cells by NGF occurred post activation of PKC-iota and was regulated by the tyrosine phosphorylation state of aPKC. Furthermore, NGF-dependent localization of ZIP/p62 was observed within vesicular structures, identified as late endosomes by colocalization with a Rab7 antibody. Both ZIP/p62 as well as PKC-iota colocalized with Rab7 upon NGF stimulation. Inhibition of the tyrosine phosphorylation state of PKC-iota did not prevent movement of ZIP/p62 to the endosomal compartment. These observations indicate that the subcellular localization of ZIP/p62 does not depend entirely upon activation of aPKC itself. Of functional importance, transfection of an antisense p62 construct into PC12 cells significantly diminished NGF-induced neurite outgrowth. Collectively, these findings demonstrate that ZIP/p62 acts as a shuttling protein involved in routing activated aPKC to an endosomal compartment and is required for mediating NGF's biological properties.