Transport of protein kinase C isoforms to the nucleus of PC12 cells by nerve growth factor: association of atypical zeta-PKC with the nuclear matrix.
In an effort to understand the role of protein kinase C (PKC) in nerve growth factor-induced differentiation, we studied the expression of PKC using isoform-specific antibodies. Western blot analysis on whole cell lysates showed that alpha,beta,gamma,epsilon,zeta, iota/lambda and mu were expressed in PC12 cells, except for theta which was absent. In nuclei obtained from control PC12 cells, small amounts of delta, epsilon, iota/lambda and zeta were detected. A computer-assisted search algorithm was used to search for the presence of bipartite nuclear targeting motifs. In classical PKC isoforms alpha,beta,gamma, two bipartite motifs were present, while atypical iota/lambda and zeta-PKC displayed one motif, whereas novel PKC isoforms did not exhibit any bipartite motif structure. Treatment of cells with differentiating doses of nerve growth factor (NGF) resulted in changes of differential magnitude for all of the nuclear PKC isoforms in response to NGF. However, little change in gamma-PKC was observed in response to NGF. This analysis indicated that other factors may contribute to transport of PKC into the nucleus, in addition to the bipartite motif itself. Atypical zeta-PKC is required for NGF-induced neurite outgrowth of PC12 cells (Coleman and Wooten: J Mol Neurosci 5:39-57, 1994). Increases in nuclear zeta-PKC were NGF dose-dependant with a concomitant decrease in cytoplasmic immunoreactivity. The localization of zeta-PKC was investigated by means of immunoelectron microscopy which revealed the localization of this isoform within the inner nuclear matrix bound to chromatin. Taken together, these findings suggest that zeta-PKC may be involved in the regulation of nuclear processes.
Wooten
M W
MW
Department of Zoology, Auburn University, Alabama 36849, USA. mwwooten@acesag.auburn.edu
Zhou
G
G
Wooten
M C
MC
Seibenhener
M L
ML
D000818:Animals; D001692:Biological Transport / Q000502:physiology; D015153:Blotting, Western; D004815:Epidermal Growth Factor / Q000378:metabolism; D007527:Isoenzymes / Q000032:analysis / Q000378:metabolism*; D008854:Microscopy, Electron; D009414:Nerve Growth Factors / Q000494:pharmacology*; D015530:Nuclear Matrix / Q000187:drug effects / Q000201:enzymology* / Q000648:ultrastructure; D016716:PC12 Cells; D011493:Protein Kinase C / Q000032:analysis / Q000378:metabolism*; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha; D051745:Protein Kinase C-delta; D051381:Rats; D000099293:Protein Kinase C-lambda; D000099221:Protein Kinase C zeta
D004815:Epidermal Growth Factor; D007527:Isoenzymes; D009414:Nerve Growth Factors; D011493:Protein Kinase C; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha; D051745:Protein Kinase C-delta; D000099293:Protein Kinase C-lambda; D000099221:Protein Kinase C zeta; C497260:Prkcd protein, rat; C087857:protein kinase C gamma; C089212:protein kinase D