Role of protein kinase C in the regulation of gap junctional communication.
We have examined the effect of protein kinase (PKC) depletion in SV40-transformed Djungarian hamster fibroblasts (DM15 cells) on the level of gap junction permeability. Cx43 electrophoretic mobility, and cell sensitivity to different uncoupling stimuli. After 24 hr exposure to 12-O-tetradecanoyl-phorbol-13-acetate (TPA), the total PKC activity in DM15 cells was reduced to 20-25% in comparison with intact cells. In PKC-depleted cells the level of dye coupling was 30-40% higher than in the same untreated cultures. Western blot analysis revealed multiple forms of the gap junction protein connexin 43, which correspond to known phosphorylated and dephosphorylated forms of this protein. No decrease in the level of connexin 43 phosphorylation after PKC depletion was observed. TPA (10(-7) g/ml), mezerein (10(-7) g/ml), teleocidin (10(-8) g/ml), Ca-ionophore A23187 (10(-6) g/ml), insecticide 1,1,1-trichloro-2,2-bis-(p-chlorphenyl)-ethane (DDT) (10(-4) g/ml), and nigericin (10(-5) M in hydrolysate lactalbumin solution, pH 6.3) induced a four-to six-fold decrease in the number of recipient cells in the dye-coupling assay. PKC-depleted cells became almost completely resistant to the uncoupling effect of mezerein, teleocidin, and A23187, as well as to new exposure to TPA, and became partially resistant to the effect of DDT. Nigericin inhibited intercellular communication between PKC-depleted cells to the same extent as between control cells. Thus, in the cell system studied, PKC plays a certain role in maintaining the basal level of gap junction permeability and has an important significance as a mediator of the uncoupling effects of such substances as TPA, mezerein, teleocidin, and Ca2+.
Budunova
I V
IV
Laboratory of Carcinogen Screening Methods, National Cancer Center, Moscow, Russia.
Mittelman
L A
LA
Miloszewska
J
J
D000818:Animals; D002450:Cell Communication / Q000187:drug effects / Q000502:physiology; D002461:Cell Line, Transformed; D002472:Cell Transformation, Viral / Q000502:physiology*; D006224:Cricetinae; D015536:Down-Regulation / Q000502:physiology; D004586:Electrophoresis; D005456:Fluorescent Dyes; D017629:Gap Junctions / Q000187:drug effects / Q000502:physiology*; D007546:Isoquinolines; D011493:Protein Kinase C / Q000502:physiology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D005456:Fluorescent Dyes; D007546:Isoquinolines; C017475:lucifer yellow; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate