title: Low- and high-affinity phorbol ester and diglyceride interactions with protein kinase C: 1-O-alkyl-2-acyl-sn-glycerol enhances phorbol ester- and diacylglycerol-induced activity but alone does not induce activity.
abstract: Phorbol ester-induced conventional protein kinase C (PKCalpha, -betaIota/IotaIota, and -gamma) isozyme activities are potentiated by 1,2-diacyl-sn-glycerol. This has been attributed to a "cooperative" interaction of the two activators with two discrete sites termed the low- and high-affinity phorbol ester binding sites, respectively [Slater, S. J., Milano, S. K., Stagliano, B. A., Gergich, K. J., Ho, C., Mazurek, A., Taddeo, F. J., Kelly, M. B., Yeager, M. D., and Stubbs, C. D. (1999) Biochemistry 38, 3804-3815]. Here, we report that the 1-O-alkyl ether diglyceride, 1-O-hexadecyl-2-acetyl-sn-glycerol (HAG), like its 1,2-diacyl counterpart, 1-oleoyl-2-acetyl-sn-glycerol (OAG), also potentiated PKCalpha, -betaI/II, and -gamma activities induced by the phorbol ester 4beta-12-O-tetradecanoylphorbol-13-acetate (TPA). Similar to OAG, HAG was found to bind to the low-affinity phorbol ester binding site and to enhance high-affinity phorbol ester binding, and to decrease the level of Ca(2+) required for phorbol ester-induced activity, while being without effect on the Ca(2+) dependence of membrane association. Thus, similar to OAG, HAG may also potentiate phorbol ester-induced activity by interacting with the low-affinity phorbol ester binding site, leading to a reduced level of Ca(2+) required for the activating conformational change. However, HAG was found not to behave like a 1,2-diacyl-sn-glycerol in that alone it did not induce PKC activity, and also in that it enhanced OAG-induced activity. The results reveal HAG to be a member of a new class of "nonactivating" compounds that modulate PKC activity by interacting with the low-affinity phorbol ester binding site.
authors.affiliation: Department of Pathology, Cell Biology, and Anatomy, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
authors.forename: S J
authors.initials: SJ
authors.lastname: Slater
authors.forename: J L
authors.initials: JL
authors.lastname: Seiz
authors.forename: B A
authors.initials: BA
authors.lastname: Stagliano
authors.forename: A C
authors.initials: AC
authors.lastname: Cook
authors.forename: S K
authors.initials: SK
authors.lastname: Milano
authors.forename: C
authors.initials: C
authors.lastname: Ho
authors.forename: C D
authors.initials: CD
authors.lastname: Stubbs
mesh_terms: D000818:Animals; D001665:Binding Sites / Q000187:drug effects; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D004075:Diglycerides / Q000378:metabolism* / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004357:Drug Synergism; D004789:Enzyme Activation / Q000187:drug effects; D004790:Enzyme Induction / Q000187:drug effects; D005995:Glyceryl Ethers / Q000378:metabolism* / Q000494:pharmacology; D007527:Isoenzymes / Q000378:metabolism; D008081:Liposomes / Q000737:chemistry / Q000378:metabolism; D010703:Phorbol Esters / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D011493:Protein Kinase C / Q000096:biosynthesis / Q000378:metabolism*; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha; D051745:Protein Kinase C-delta; D051381:Rats; D013755:Tetradecanoylphorbol Acetate / Q000378:metabolism* / Q000494:pharmacology
chemical_list: C011439:1,2-diacylglycerol; D004075:Diglycerides; D005995:Glyceryl Ethers; D007527:Isoenzymes; D008081:Liposomes; D010703:Phorbol Esters; C065374:sapintoxin D; C038237:1-oleoyl-2-acetylglycerol; C497260:Prkcd protein, rat; C115197:calcium-independent protein kinase C; D011493:Protein Kinase C; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha; D051745:Protein Kinase C-delta; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium