佛波酯引起蛋白激酶C下降调节的专一性

The Specificity of Down Regulation of Protein Kinase C Induced by Phorbol Ester

探讨了佛波酯（ＰＭＡ）对蛋白激酶的下降调节是否有激酶专一性及亚型专一性．用组蛋白Ｈ１作为蛋白激酶Ｃ（ＰＫＣ）和蛋白激酶Ａ（ＰＫＡ）的受体底物，加入ＰＫＣ和ＰＫＡ的特异性激活剂区分ＰＫＣ和ＰＫＡ，用聚谷酪（４１）为酪氨酸蛋白激酶（ＴＰＫ）的专一性受体底物，以３２Ｐ－ＡＴＰ为３２Ｐ共同供体底物测定三种蛋白激酶的活力，并用免疫组化法测定ＰＫＣ亚型．结果发现ＰＭＡ对人７７２１肝癌细胞只引起ＰＫＣ而不引起ＰＫＡ和ＴＰＫ的下降调节，ＰＫＣ的非特异性抑制剂槲皮素和特异性抑制剂Ｄ－鞘氨醇能大部分取消ＰＭＡ对ＰＫＣ的下降调节，但ＴＰＫ抑制剂ｇｅｎｅｓｔｅｉｎ则没有阻断下降调节的作用．用ＨＬ－６０细胞还证明ＰＭＡ只对含量丰富的ＰＫＣα和ＰＫＣβⅡ亚型而不对含量很少的ＰＫＣβⅠ亚型发生下降调节．上述结果说明ＰＭＡ对蛋白激酶的下降调节有激酶和亚型专一性．

The questions whether the down regulation of protein kinase by phorbol ester(PMA) are specific for one protein kinase and its subtype were studied.Using histone H1 as the acceptor substrates of protein kinase C(PKC) and protien kinase A(PKA),adding specific activators for PKC and PKA assays,and using poly Glu·Tyr(4∶1) as the acceptor substrate of tyrosine proteion kinase(TPK),the three protien kinases were detected with 32 P ATP as the common donor of 32 P.The subtypes of PKC were determined by immunohistochemical method.It was discovered that PMA only induced the down regulation of PKC,but not PKA and TPK in 7721 human hepaticarcinoma cell line.Quercetin,a non specific inhibitor of PKC,and D sphingosine,a specific inhibitor of PKC,partially eliminated the down regulation of PKC induced by PMA,while the inhibitor of TPK,genestein,did not show any blocking effect on the down regulation of PKC.It was also found that PMA only down regulated the abundant PKC α and PKC β Ⅱ subtype of PKC,but not the rare PKC β Ⅰ subtype in HL 60 cells.