Yao Lina, Fan Peidong, Jiang Zhan, Gordon Adrienne, Mochly-Rosen Daria, Diamond Ivan
CV Therapeutics, Inc., 3172 Porter Drive, Palo Alto, CA 94304, USA.
Mol Pharmacol. 2008 Apr;73(4):1105-12. doi: 10.1124/mol.107.042580. Epub 2008 Jan 17.
We found previously that neural responses to ethanol and the dopamine D2 receptor (D2) agonist 2,10,11-trihydroxy-N-propylnorapomorphine hydrobromide (NPA) involve both epsilon protein kinase C (epsilonPKC) and cAMP-dependent protein kinase A (PKA). However, little is known about the mechanism underlying ethanol- and D2-mediated activation of epsilonPKC and the relationship to PKA activation. In the present study, we used a new epsilonPKC antibody, 14E6, that selectively recognized active epsilonPKC when not bound to its anchoring protein epsilonRACK (receptor for activated C-kinase), and PKC isozyme-selective inhibitors and activators to measure PKC translocation and catalytic activity. We show here that ethanol and NPA activated epsilonPKC and induced translocation of both epsilonPKC and its anchoring protein, epsilonRACK to a new cytosolic site. The selective epsilonPKC agonist, pseudo-epsilonRACK, activated epsilonPKC but did not cause translocation of the epsilonPKC/epsilonRACK complex to the cytosol. These data suggest a step-wise activation and translocation of epsilonPKC after NPA or ethanol treatment, where epsilonPKC first translocates and binds to its RACK and subsequently the epsilonPKC/epsilonRACK complex translocates to a new subcellular site. Direct activation of PKA by adenosine-3',5'-cyclic monophosphorothioate, Sp-isomer (Sp-cAMPS), prostaglandin E1, or the adenosine A2A receptor is sufficient to cause epsilonPKC translocation to the cytosolic compartment in a process that is dependent on PLC activation and requires PKA activity. These data demonstrate a novel cross-talk mechanism between epsilonPKC and PKA signaling systems. PKA and PKC signaling have been implicated in alcohol rewarding properties in the mesolimbic dopamine system. Cross-talk between PKA and PKC may underlie some of the behaviors associated with alcoholism.
我们之前发现，对乙醇和多巴胺D2受体（D2）激动剂2,10,11 - 三羟基 - N - 丙基去甲阿朴吗啡氢溴酸盐（NPA）的神经反应涉及ε蛋白激酶C（εPKC）和环磷酸腺苷依赖性蛋白激酶A（PKA）。然而，关于乙醇和D2介导的εPKC激活机制以及与PKA激活的关系却知之甚少。在本研究中，我们使用了一种新的εPKC抗体14E6，当未与其锚定蛋白εRACK（活化C激酶受体）结合时，它能选择性识别活性εPKC，还使用了PKC同工酶选择性抑制剂和激活剂来测量PKC易位和催化活性。我们在此表明，乙醇和NPA激活了εPKC，并诱导εPKC及其锚定蛋白εRACK易位至一个新的胞质位点。选择性εPKC激动剂伪εRACK激活了εPKC，但未导致εPKC/εRACK复合物易位至胞质溶胶。这些数据表明，在NPA或乙醇处理后，εPKC会逐步激活和易位，其中εPKC首先易位并与其RACK结合，随后εPKC/εRACK复合物易位至一个新的亚细胞位点。通过腺苷 - 3',5' - 环磷硫代酸酯Sp - 异构体（Sp - cAMPS）、前列腺素E1或腺苷A2A受体直接激活PKA足以导致εPKC在依赖磷脂酶C激活且需要PKA活性的过程中易位至胞质区室。这些数据证明了εPKC和PKA信号系统之间一种新的相互作用机制。PKA和PKC信号传导与中脑边缘多巴胺系统中酒精的奖赏特性有关。PKA和PKC之间的相互作用可能是一些与酒精中毒相关行为的基础。
