The major chemical-detoxifying system of UDP-glucuronosyltransferases requires regulated phosphorylation supported by protein kinase C
The major chemical-detoxifying system of UDP-glucuronosyltransferases requires regulated phosphorylation supported by protein kinase C
复制标题
DOI：
10.1074/jbc.m800032200
复制
发表时间：
2008-08-22
影响因子：
4.8
通讯作者：
Owens, Ida S.
中科院分区：
文献类型：
--
作者：
Basu, Nikhil K.;Kole, Labanyamoy;Owens, Ida S.
在姜黄素处理后发现LS 180细胞中人UDP-葡萄糖醛酸转移酶（UGT）的快速可逆下调导致发现UGT需要磷酸化。UGT主要分布于肝脏、肾脏和胃肠道，是一种芳香类代谢物和大量的饮食和环境化学物质，可降低毒性、致突变和致癌的风险。我们的目的是确定相关的激酶和机制（S）调节组成型UGT在LS 180细胞和10个不同的人UGT cDNA转染COS-1系统的磷酸化。在用姜黄素或PKC抑制剂calphostin-C处理LS 180细胞后，UGT和蛋白激酶C中免疫可检测的[P-33]正磷酸盐的时间和浓度依赖性抑制是（PKC是）的一个要素，表明UGT磷酸化受到活性PKC的支持。UGT转染细胞的免疫荧光和免疫共沉淀研究表明，UGT 1A 7 His和PKC的共定位是UGT 1A 10 His和PKC α或PKC δ的一个元素。通过PKC抑制UGT活性是特异性拮抗肽的一个要素，或者通过PKC是靶向破坏的一个要素，PKC是特异性小干扰RNA的一个要素，并且用典型的PKC激动剂激活姜黄素下调的UGT证实了PKC在葡萄糖醛酸化中的中心作用。此外，新生UGT 1A 7 His通过PKC的体外磷酸化是确认其为真正的PKC底物的要素。最后，过氧化氢酶或herbimycin-A抑制组成或过氧化氢激活的UGT表明，活性氧相关的氧化剂作为第二信使，在维持组成PKC依赖的信号传导，显然维持UGT磷酸化和活性。由于细胞共同使用信号转导来检测和适当地响应环境变化，本报告结合我们先前的证明，即UGT 1A 7中的特定磷酸基团决定底物选择，表明调节磷酸化允许UGT对差异磷酸盐利用的适应有效发挥作用。
Finding rapid, reversible down-regulation of human UDP-glucuronosyltransferases (UGTs) in LS180 cells following curcumin treatment led to the discovery that UGTs require phosphorylation. UGTs, distributed primarily in liver, kidney, and gastrointestinal tract, inactivate aromatic-like metabolites and a vast number of dietary and environmental chemicals, which reduces the risk of toxicities, mutagenesis, and carcinogenesis. Our aim here is to determine relevant kinases and mechanism(s) regulating phosphorylation of constitutive UGTs in LS180 cells and 10 different human UGT cDNA-transfected COS-1 systems. Time- and concentration-dependent inhibition of immunodetectable [P-33] orthophosphate in UGTs and protein kinase C is an element of(PKC is an element of), following treatment of LS180 cells with curcumin or the PKC inhibitor calphostin-C, suggested UGT phosphorylation is supported by active PKC( s). Immunofluorescent and co-immunoprecipitation studies with UGT-transfected cells showed co-localization of UGT1A7His and PKC is an element of and of UGT1A10His and PKC alpha orPKC delta. Inhibition of UGT activity by PKC is an element of-specific antagonist peptide or by PKC is an element of-targeted destruction with PKC is an element of-specific small interference RNA and activation of curcumin-down-regulated UGTs with typical PKC agonists verified a central PKC role in glucuronidation. Moreover, in vitro phosphorylation of nascent UGT1A7His by PKC is an element of confirms it is a bona fide PKC substrate. Finally, catalase or herbimycin-A inhibition of constitutive or hydrogen peroxide-activated UGTs demonstrated that reactive oxygen species-related oxidants act as second messengers in maintaining constitutive PKC-dependent signaling evidently sustaining UGT phosphorylation and activity. Because cells use signal transduction collectively to detect and respond appropriately to environmental changes, this report, combined with our earlier demonstration that specific phospho-groups in UGT1A7 determined substrate selections, suggests regulated phosphorylation allows adaptations regarding differential phosphate utilization by UGTs to function efficiently.