Overexpression of protein kinase C-delta and -epsilon in NIH 3T3 cells induces opposite effects on growth, morphology, anchorage dependence, and tumorigenicity.
We have determined the patterns of mRNA and protein expression of 7 protein kinase C (PKC) isozymes in NIH 3T3 cells. Only PKC-alpha is expressed abundantly in NIH 3T3 cells; endogenous levels of the other 6 PKC isozymes are low or undetectable. We have overexpressed PKC-delta and -epsilon in these cells to observe activation/translocation of these two isozymes and the biological consequences of overexpression. Both PKC-delta and -epsilon, but not PKC-alpha, are partially associated with the insoluble fraction even in the absence of phorbol 12-myristate 13-acetate (PMA). Upon PMA stimulation, both PKC-delta and -epsilon translocate to the insoluble fraction of cell homogenates, as can be observed with the endogenous PKC-alpha. Overexpression of PKC-delta induces significant changes in morphology and causes the cells to grow more slowly and to a decreased cell density in confluent cultures. These changes are accentuated by treatment with PMA. Overexpression of PKC-epsilon does not lead to morphological changes, but causes increased growth rates and higher cell densities in monolayers. None of the PKC-delta overexpressers grow in soft agar with or without PMA, but all the cell lines that overexpress PKC-epsilon grow in soft agar in the absence of PMA, but not in its presence. NIH 3T3 cells that overexpress PKC-epsilon also form tumors in nude mice with 100% incidence. This indicates that high expression of PKC-epsilon contributes to neoplastic transformation.
Mischak
H
H
Laboratory of Genetics, National Cancer Institute, Bethesda, Maryland 20892.
Goodnight
J A
JA
Kolch
W
W
Martiny-Baron
G
G
Schaechtle
C
C
Kazanietz
M G
MG
Blumberg
P M
PM
Pierce
J H
JH
Mushinski
J F
JF
D016475:3T3 Cells; D000818:Animals; D001692:Biological Transport; D015152:Blotting, Northern; D015153:Blotting, Western; D001921:Brain / Q000201:enzymology; D002448:Cell Adhesion / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002471:Cell Transformation, Neoplastic; D003001:Cloning, Molecular; D004789:Enzyme Activation; D004795:Enzyme Stability; D007211:Indoles / Q000494:pharmacology; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008301:Maleimides / Q000494:pharmacology; D051379:Mice; D008810:Mice, Inbred C57BL; D008819:Mice, Nude; D009374:Neoplasms, Experimental / Q000209:etiology; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D007211:Indoles; D007527:Isoenzymes; D008301:Maleimides; D011493:Protein Kinase C; C070515:bisindolylmaleimide I; D013755:Tetradecanoylphorbol Acetate