Involvement of protein kinase Cdelta in contact-dependent inhibition of growth in human and murine fibroblasts.
There is evidence that protein kinase C delta (PKCdelta) is a tumor suppressor, although its physiological role has not been elucidated so far. Since important anti-proliferative signals are mediated by cell-cell contacts we studied whether PKCdelta is involved in contact-dependent inhibition of growth in human (FH109) and murine (NIH3T3) fibroblasts. Cell-cell contacts were imitated by the addition of glutardialdehyde-fixed cells to sparsely seeded fibroblasts. Downregulation of the PKC isoforms alpha, delta, epsilon, and mu after prolonged treatment with 12-O-tetradecanoylphorbol-13-acetate (TPA, 0.1 microM) resulted in a significant release from contact-inhibition in FH109 cells. Bryostatin 1 selectively prevented TPA-induced PKCdelta-downregulation and reversed TPA-induced release from contact-inhibition arguing for a role of PKCdelta in contact-inhibition. In accordance, the PKCdelta specific inhibitor Rottlerin (1 microM) totally abolished contact-inhibition. Interestingly, immunofluorescence revealed a rapid translocation of PKCdelta to the nucleus when cultures reached confluence with a peak in early-mid G1 phase. Nuclear translocation of PKCdelta in response to cell-cell contacts could also be demonstrated after subcellular fractionation by Western blotting and by measuring PKCdelta-activity after immunoprecipitation. Transient transfection of NIH3T3 cells with a dominant negative mutant of PKCdelta induced a transformed phenotype. We conclude that PKCdelta is involved in contact-dependent inhibition of growth.
Heit
I
I
Institute of Toxicology, Johannes Gutenberg-University of Mainz, 55131 Mainz, Germany.
Wieser
R J
RJ
Herget
T
T
Faust
D
D
Borchert-Stuhlträger
M
M
Oesch
F
F
Dietrich
C
C
D016475:3T3 Cells; D000098:Acetophenones / Q000494:pharmacology; D021581:Active Transport, Cell Nucleus; D000818:Animals; D001578:Benzopyrans / Q000494:pharmacology; D054713:Bryostatins; D002453:Cell Cycle; D002455:Cell Division / Q000187:drug effects; D002633:Chemotaxis / Q000187:drug effects; D015536:Down-Regulation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts / Q000378:metabolism*; D005404:Fixatives / Q000494:pharmacology; D005976:Glutaral / Q000494:pharmacology; D006801:Humans; D007527:Isoenzymes / Q000737:chemistry / Q000378:metabolism*; D007783:Lactones / Q000494:pharmacology; D018942:Macrolides; D051379:Mice; D008934:Mitogens / Q000494:pharmacology; D011485:Protein Binding; D020033:Protein Isoforms; D011493:Protein Kinase C / Q000737:chemistry / Q000378:metabolism*; D051745:Protein Kinase C-delta; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013997:Time Factors
D000098:Acetophenones; D001578:Benzopyrans; D054713:Bryostatins; D004791:Enzyme Inhibitors; D005404:Fixatives; D007527:Isoenzymes; D007783:Lactones; D018942:Macrolides; D008934:Mitogens; D020033:Protein Isoforms; C046785:bryostatin 1; C085746:rottlerin; C497259:Prkcd protein, mouse; C497258:PRKCD protein, human; D011493:Protein Kinase C; D051745:Protein Kinase C-delta; D013755:Tetradecanoylphorbol Acetate; D005976:Glutaral