Effects of ethanol on protein kinase C alpha activity induced by association with Rho GTPases.
Previous studies have shown that n-alkanols have biphasic chain length-dependent effects on protein kinase C (PKC) activity induced by association with membranes or with filamentous actin [Slater, S. J., et al. (1997) J. Biol. Chem. 272, 6167-6173; Slater, S. J., et al. (2001) Biochim. Biophys. Acta 1544, 207-216]. Recently, we showed that PKCalpha is also activated by a direct membrane lipid-independent interaction with Rho GTPases. Here, the effects of ethanol and 1-hexanol on Rho GTPase-induced activity were investigated using an in vitro assay system to provide further insight into the mechanism of the effects of n-alkanols on PKC activity. Both ethanol and 1-hexanol were found to have two competing concentration-dependent effects on the Ca(2+)- and phorbol ester- or diacylglycerol-dependent activities of PKCalpha associated with either RhoA or Cdc42, consisting of a potentiation at low alcohol levels and an attenuation of activity at higher levels. Measurements of the Ca(2+), phorbol ester, and diacylglycerol concentration-response curves for Cdc42-induced activation indicated that the activating effect corresponded to a shift in the midpoints of each of the curves to lower activator concentrations, while the attenuating effect corresponded to a decrease in the level of activity induced by maximal activator levels. The presence of ethanol enhanced the interaction of PKCalpha with Cdc42 within a concentration range corresponding to the potentiating effect, whereas the level of binding was unaffected by higher ethanol levels that were found to attenuate activity. Thus, ethanol may either enhance activation of PKCalpha by Rho GTPases by enhancing the interaction between the two proteins or attenuate the level of activity of Rho GTPase-associated PKCalpha by inhibiting the ensuing activating conformational change. The results also suggest that the effects of ethanol on Rho GTPase-induced activity may switch between an activation and inhibition depending on the concentration of Ca(2+) and other activators.
Slater
Simon J
SJ
Department of Pathology, Anatomy and Cell Biology, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
Cook
Anthony C
AC
Seiz
Jodie L
JL
Malinowski
Steve A
SA
Stagliano
Brigid A
BA
Stubbs
Christopher D
CD
D001667:Binding, Competitive / Q000187:drug effects; D002118:Calcium / Q000737:chemistry / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D000431:Ethanol / Q000494:pharmacology*; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism; D006165:Guanylyl Imidodiphosphate / Q000737:chemistry; D000441:Hexanols / Q000494:pharmacology; D006801:Humans; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism*; D051571:Protein Kinase C-alpha; D013755:Tetradecanoylphorbol Acetate / Q000737:chemistry / Q000494:pharmacology; D020764:cdc42 GTP-Binding Protein / Q000737:chemistry* / Q000378:metabolism*; D020742:rhoA GTP-Binding Protein / Q000737:chemistry* / Q000378:metabolism*
D020662:Guanine Nucleotide Exchange Factors; D000441:Hexanols; D007527:Isoenzymes; D006165:Guanylyl Imidodiphosphate; D000431:Ethanol; C036260:1-hexanol; C496628:PRKCA protein, human; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D020764:cdc42 GTP-Binding Protein; D020742:rhoA GTP-Binding Protein; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium