1990 Fiscal Year Final Research Report Summary
Function of Calcium-Dependent Protease in Down Regulation of Protein Dinase C
| Project/Area Number |
01480526
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
物質生物化学
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| Research Institution | Tokyo Metropolitan Institute of Medical Science |
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Principal Investigator |
SUZUKI Koichi Tokyo Metropol. Inst. Med. Sci. Dept. Mol. Bio., Director, 遺伝情報部門, 部長 (80011948)
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| Co-Investigator(Kenkyū-buntansha) |
SAIDO Takaomi Tokyo Metropol. Inst. Med. Sci. Dept. Mol. Bio., researcher, 遺伝情報部門, 研究員 (80205690)
OHNO Shigeo Tokyo Metropol. Inst, Med. Sci. Dept. Mol. Bio., researcher, 遺伝情報部門, 研究所 (10142027)
AKITA Yoshiko Tokyo Metropol. Inst. Med. Sci. Dept. Mol. Bio., researcher, 遺伝情報部門, 研究員 (40124432)
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| Project Period (FY) |
1989 – 1990
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| Keywords | protein kinase C / calcium protease / phorbol ester |
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| Research Abstract |
1. Phosphorylation of protein kinase C (PKC) is essential for its down-regulation
PKCalpha cDNA was expressed transiently in COS cells or stably in rat fibroblast 3Yl cells. The down regulation of PKCalpha was examined using these cells. A point mutation where Lys-368 at the putative ATP-binding site is replaced with arginine confers enhanced phorbol ester binding activity to both COS and 3Yl cells. Like endogenous and exogenously expressed wild type PKCalpha, the mutant is translocated from the cytosol to the particulate fraction when cells are treated with a phorbol ester (TPA). The mutant PKC is, however, not degraded after the TPA treatment, making a clear contrast to wild type PKC. The mutant is resistant to TPA-induced down-regulation. The mutant lacks kinase activity as expected, as judged by autophosphorylation and by a kinase assay. Autophosphorylation of PKC is a requisite for proteolytic cleavage associated with down-regulation of PKC.
2. Novel calcium independent protein kinase (nPKC) also transduce physiological signal
In GH4Cl cells, nPKCepsilon is expressed together with PKCalpha and beta II. nPKC epsilon is translocated to the cell membrane from the cytosol and down regulated when cells are treated with Thyrotropin Releasing Hormone (TRH), whereas other conventional PKC's are not down-regulated. Thus in GH4Cl cells, nPKCepsilon plays a crucial role distinct from that meadiated by calcium-dependent PKCalpha and beta II in cellular response elicited by both TRH and phorbol esters.
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