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.
Samuels
I S
IS
Department of Biological Sciences, Program in Cellular and Molecular Biosciences, Auburn University, Auburn, Alabama 36849, USA.
Seibenhener
M L
ML
Neidigh
K B
KB
Wooten
M W
MW
D000818:Animals; D015153:Blotting, Western; D000081247:CSK Tyrosine-Protein Kinase; D002352:Carrier Proteins / Q000378:metabolism*; D002454:Cell Differentiation / Q000187:drug effects*; D002460:Cell Line; D048429:Cell Size / Q000187:drug effects; D011992:Endosomes / Q000187:drug effects* / Q000378:metabolism*; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D008856:Microscopy, Fluorescence; D020932:Nerve Growth Factor / Q000494:pharmacology*; D016501:Neurites / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D016716:PC12 Cells; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding / Q000187:drug effects; D011493:Protein Kinase C / Q000378:metabolism*; D021381:Protein Transport / Q000187:drug effects; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D051381:Rats; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D019061:src-Family Kinases
D002352:Carrier Proteins; D007527:Isoenzymes; D019000:Phosphotyrosine; D020932:Nerve Growth Factor; D011505:Protein-Tyrosine Kinases; D000081247:CSK Tyrosine-Protein Kinase; D019061:src-Family Kinases; C000631346:CSK protein, human; D011493:Protein Kinase C