Dephosphorylation of activated protein kinase c contributes to
downregulation by bryostatin.
Lee, Hyeon-Woo, Lucinda Smith, George R. Pettit, and Jeffrey Bingham
Smith.
Department of Pharmacology and Toxicology, Schools of Medicine and
Dentistry, University of Alabama at Birmingham, Birmingham, AL 35294,
and the Cancer Research Institute and Department of Chemistry,
Arizona State University, Tempe, AZ 85287
APStracts 3:0040C, 1996.
Bryostatin 1 (Bryo), like PMA, acutely activated PKC as indicated by
autophosphorylation and translocation of PKC-[alpha], the predominant
PMA-sensitive isoform expressed by LLC-MK2 renal epithelial cells.
Bryo concomitantly increased the 32P labeling of 80 kDa PKC-[alpha]
by autophosphorylation and produced a 76 kDa form of PKC-[alpha] that
lacked detectable 32P. The 76 kDa form was in the particulate, rather
than the cytosolic, fraction, which suggests that it was produced
from activated kinase. Alkaline phosphatase treatment of
immunoprecipitated PKC-[alpha] converted the 80 kDa form to 76 kDa,
but had no effect on the mobility of the 76 kDa form suggesting that
it was not phosphorylated. Pulse-chase labeling of PKC-[alpha] with
35S-Met/Cys indicated that there is a precursor-product relationship
between the 80 and 76 kDa forms, respectively. Inhibition of protein
synthesis had no effect on the production of 76 kDa PKC-[alpha] by
Bryo. PMA also produced 76 kDa PKC-[alpha], but was less potent than
Bryo. Bryo produced a more rapid loss of 80 kDa PKC-[alpha] protein
and total Ca2+- and phospholipid-dependent PKC activity than PMA.
Dephosphorylation contributes to rapid PKC-[alpha] downregulation
because the 76 kDa form lacked 32P under conditions that
autophosphorylated the 80 kDa form. We suggest that dephosphorylation
predisposes PKC to proteolysis.
Received 26 September 1995; accepted in final form 17 January
1996.
APS Manuscript Number C588-5.
Article publication pending Am. J. Physiol. (Cell Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 8 February 96