GROWTH OF MELANOCYTIC CELLS IS ASSOCIATED WITH DOWN-REGULATION OF PROTEIN-KINASE-C ALPHA-ISOFORM, DELTA-ISOFORM, AND EPSILON-ISOFORM - POSSIBLE ROLE OF DIACYLGLYCEROL
G. Brooks et al., GROWTH OF MELANOCYTIC CELLS IS ASSOCIATED WITH DOWN-REGULATION OF PROTEIN-KINASE-C ALPHA-ISOFORM, DELTA-ISOFORM, AND EPSILON-ISOFORM - POSSIBLE ROLE OF DIACYLGLYCEROL, The Journal of biological chemistry, 268(32), 1993, pp. 23868-23875
Protein kinase C (PKC) down-regulation has been shown to correlate wit
h the growth of murine melanocytic cells in culture (Brooks, G., Wilso
n, R. E., Dooley, T. P., Goss, M. W., and Hart, I. R. (1991) Cancer Re
s. 51, 3281-3288). We now show that PKC alpha, delta, epsilon, and zet
a isoforms are present at the protein level in quiescent, non-transfor
med Mel-ab melanocytes, maintained in the absence of phorbol ester. Pr
oliferation of Mel-ab cells, achieved by incubation in the continual p
resence of phorbol 12,13-dibutyrate, was associated with a down-regula
tion of the PKC alpha, delta, and epsilon isozymes. Examination of two
transformed syngeneic lines (the B16 murine melanoma and the long ter
minal repeat Ras.2 line), that grew in the absence of exogenous phorbo
l esters, showed that PKC alpha protein levels were either partially d
own-regulated or unaffected, the PKC delta and epsilon isoforms were d
own-regulated completely, and the levels of PKC zeta protein remained
unaltered relative to quiescent Mel-ab cells. Basal levels of total di
acylglycerol were elevated 5-fold in B16 melanoma cells compared with
levels found in quiescent or proliferating Mel-ab melanocytes and appe
ar to arise largely from the breakdown of phosphatidylinositol phospho
lipids accompanied by a significant rise in phospholipase C activity.
Hourly treatments of quiescent Mel-ab melanocytes with the synthetic d
iacylglycerol analogue, 1,2-dioctanoyl-sn-glycerol, for 24 h, resulted
in an induction of DNA synthesis which was associated with a signific
ant down-regulation of PKC levels mediated largely via post-translatio
nal rather than transcriptional mechanisms. These results show for the
first time that specific isoforms of PKC are down-regulated at the pr
otein level during proliferation of murine melanocytic cells and sugge
st that the constitutive down-regulation of PKC in transformed melanom
a cells may arise as a consequence of elevated endogenous phosphatidyl
inositol-derived diacylglycerol levels.