Further evidence that 3-phosphoinositide-dependent protein kinase-1 (PDK1) is required for the stability and phosphorylation of protein kinase C (PKC) isoforms
Further evidence that 3-phosphoinositide-dependent protein kinase-1 (PDK1) is required for the stability and phosphorylation of protein kinase C (PKC) isoforms
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DOI：
10.1016/s0014-5793(00)02162-1
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发表时间：
2000-11-10
期刊：
影响因子：
3.5
通讯作者：
Alessi, DR
中科院分区：
文献类型：
--
作者：
Balendran, A;Hare, GR;Alessi, DR
蛋白激酶C（PKC）超家族的多位点磷酸化在这些酶的调节中起着重要作用。活化所有PKC亚型所需的关键磷酸化位点之一位于激酶结构域的T环中。最近的体外和转染实验表明，该残基的磷酸化可以由3-磷酸肌醇依赖性蛋白激酶1（PDK 1）介导。在这项研究中，我们证明在缺乏PDK 1的胚胎干细胞（ES细胞）中，（PDK 1-/-细胞），内源性表达的PKC α，PKC β I，PKC γ，PKC δ，PKC β，与对照ES细胞（PDK 1 +/+细胞）相比，PKC相关激酶-1（PRK 1）的表达显著降低。在PDK 1-/- ES细胞中，PKC zeta和PRK 2蛋白的水平仅适度降低。我们证明，在相反的PKC ζ表达PDK 1 +/+ ES细胞，PKC ζ在ES细胞缺乏PDK 1是不是磷酸化的T-环残基。这提供了第一个遗传学证据，证明PKC zeta是PDK 1的生理底物，相反，PRK 2在PDK 1-/-细胞中的T环处仍然部分磷酸化，表明存在PDK 1-独立的PRK 2磷酸化机制。(C)2000年欧洲生物化学学会联合会。由Elsevier Science B，V出版，保留所有权利。
The multi-site phosphorylation of the protein kinase C (PKC) superfamily plays an important role in the regulation of these enzymes. One of the key phosphorylation sites required for the activation of all PKC isoforms lies in the T-loop of the kinase domain. Recent in vitro and transfection experiments indicate that phosphorylation of this residue can be mediated by the 3-phosphoinositide-dependent protein kinase-1 (PDK1), In this study, we demonstrate that in embryonic stem (ES) cells lacking PDK1 (PDK1 -/- cells), the intracellular levels of endogenously expressed PKC alpha, PKC betaI, PKC gamma, PKC delta, PKC epsilon, and PKC-related kinase-1 (PRK1) are vastly reduced compared to control ES cells (PDK1 +/+ cells). The levels of PKC zeta and PRK2 protein are only moderately reduced in the PDK1 -/- ES cells. We demonstrate that in contrast to PKC zeta expressed PDK1 +/+ ES cells, PKC zeta in ES cells lacking PDK1 is not phosphorylated at its T-loop residue. This provides the first genetic evidence that PKC zeta is a physiological substrate for PDK1, In contrast, PRK2 is still partially phosphorylated at its T-loop in PDK1 -/- cells, indicating the existence of a PDK1-independent mechanism for the phosphorylation of PRK2 at this residue. (C) 2000 Federation of European Biochemical Societies. Published by Elsevier Science B,V, All rights reserved.