PKA rapidly enhances proteasome assembly and activity in in vivo canine hearts
PKA rapidly enhances proteasome assembly and activity in in vivo canine hearts
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DOI：
10.1016/j.yjmcc.2008.11.001
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发表时间：
2009-04-01
影响因子：
5
通讯作者：
Kitakaze, Masafumi
中科院分区：
文献类型：
--
作者：
Asai, Mitsutoshi;Tsukamoto, Osamu;Kitakaze, Masafumi
蛋白酶体通过消除泛素化蛋白来调节多种细胞功能。蛋白激酶A(PKA)是蛋白酶体活性的关键调节因子。然而，目前还不清楚PKA是如何调节蛋白酶体活性的，以及它是否控制了活体心脏中的蛋白酶体活性。体外和凝胶内肽酶检测均表明，PKA作用30min可剂量依赖性地激活纯化的26S蛋白酶体。同时，通过非还原天然聚丙烯酰胺凝胶电泳法检测，PKA处理促进了纯化的26S蛋白酶体的磷酸化和组装，这两种作用都被PKA抑制剂H-89预先钝化。在体犬心脏，外源性或内源性PKA刺激30min后，冠脉内注射异丙肾上腺素或Forsklin 30min，或缺血预适应(IP)加4次重复缺血5min，可增强蛋白酶体的组装和活性。冠脉内注射H-89可钝化IP对蛋白酶体组装和活性的增强作用。缺血结束时，心肌蛋白酶体活性较对照组降低，但与对照组相比无明显差异。IP减少了犬缺血/再灌注心肌中泛素化蛋白的积聚，这种作用可被冠脉内给予蛋白酶体抑制剂环氧米星所钝化。然而，IP激活的蛋白酶体并不参与其限制梗塞大小的作用。这些发现表明，PKA迅速增强了活体心脏中蛋白酶体的组装和活性。还需要进一步的研究来阐明PKA介导的蛋白酶体激活在缺血/再灌流心脏中的病理生理作用。(C)2008 Elsevier Inc.保留所有权利。
Proteasome regulates diverse cellular functions by eliminating ubiquitinated proteins. Protein kinase A (PKA) is a key regulator of proteasome activity. However, it remains unknown how PKA regulates proteasome activity and whether it controls proteasome activity in in vivo hearts. Both the in vitro peptidase assay and the in-gel peptidase assays showed that the treatment with PKA for 30 min dose-dependently activated purified 26S proteasome. Simultaneously, PKA treatment enhanced phosphorylation and assembly of purified 26S proteasome evaluated by non-reducing native polyacrylamide gel electrophoresis, either of which was blunted by the pretreatment with a PKA inhibitor, H-89. In in vivo canine hearts, proteasome assembly and activity were enhanced 30 min after the exogenous or endogenous stimulation of PKA by the intracoronary administration of isoproterenol or forskolin for 30 min or by ischemic preconditioning (IP) with 4 times of repeated 5 min of ischemia. The intracoronary administration of H-89 blunted the enhancement of proteasome assembly and activity by IP. Myocardial proteasome activity at the end of ischemia was decreased compared with the control, however, it did not differ from the control in dogs with IP. IP decreased the accumulation of ubiquitinated proteins in the canine ischemia/reperfusion myocardium, which was blunted by the intracoronary administration of a proteasome inhibitor, epoxomicin. However, proteasome activation by IP was not involved in its infarct size-limiting effects. These findings indicate that PKA rapidly enhances proteasome assembly and activity in in vivo hearts. Further investigation will be needed to clarify pathophysiological roles of PKA-mediated proteasome activation in ischemia/reperfusion hearts. (C) 2008 Elsevier Inc. All rights reserved.