Adenylyl cyclase type VI gene transfer reduces phospholamban expression in cardiac myocytes via activating transcription factor 3
Adenylyl cyclase type VI gene transfer reduces phospholamban expression in cardiac myocytes via activating transcription factor 3
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DOI：
10.1074/jbc.m405701200
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发表时间：
2004-09-10
影响因子：
4.8
通讯作者：
Hammond, HK
中科院分区：
文献类型：
--
作者：
Gao, MH;Tang, T;Hammond, HK
腺苷酸环化酶VI型（AC（VI））的心脏定向表达增加了刺激的cAMP产生，改善了心脏功能，并增加了心肌病患者的存活率。相反，增加细胞内cAMP水平的药物对心脏功能和存活具有不利影响。我们想知道是否独立于cAMP的效应可能是与AC（VI）表达相关的这些有益结果的原因。因此，我们进行了一系列的实验，重点是如何基因转录的影响AC（VI）在培养的新生大鼠心肌细胞，特别关注的基因，可能会影响心脏功能。我们发现AC（VI）的过表达下调了受磷蛋白（一种肌浆网Ca 2 +-ATP酶抑制剂）的mRNA和蛋白表达。我们确定受磷蛋白（PLB）启动子中的cAMP响应样元件对于AC（VI）的下调至关重要。AC（VI）的过表达不改变CREB、CREM、ATF 1、ATF 2或ATF 4蛋白的表达。相反，AC（VI）的过表达增加了转录抑制因子ATF 3蛋白的表达。AC（VI）基因转移后，当用异丙肾上腺素或NKH 477（一种直接刺激AC的水溶性毛喉素类似物）刺激心肌细胞时，ATF 3蛋白的表达增加更多，这与PLB表达减少相关。然后，我们发现AC（VI）诱导的ATF 3蛋白与PLB启动子上的cAMP反应样元件结合，心肌细胞中ATF 3的过表达抑制PLB启动子活性。这些发现表明，AC（VI）对基因转录的影响并不直接依赖于cAMP的产生。
Cardiac-directed expression of adenylyl cyclase type VI (AC(VI)) increases stimulated cAMP production, improves heart function, and increases survival in cardiomyopathy. In contrast, pharmacological agents that increase intracellular levels of cAMP have detrimental effects on cardiac function and survival. We wondered whether effects that are independent of cAMP might be responsible for these salutary outcomes associated with AC(VI) expression. We therefore conducted a series of experiments focused on how gene transcription is influenced by AC(VI) in cultured neonatal rat cardiac myocytes, with a particular focus on genes that might influence cardiac function. We found that overexpression of AC(VI) down-regulated mRNA and protein expression of phospholamban, an inhibitor of the sarcoplasmic reticulum Ca2+-ATPase. We determined that the cAMP-responsive-like element in the phospholamban (PLB) promoter was critical for down-regulation by AC(VI). Overexpression of AC(VI) did not alter the expression of CREB, CREM, ATF1, ATF2, or ATF4 proteins. In contrast, overexpression of AC(VI) increased expression of ATF3 protein, a suppressor of transcription. Following AC(VI) gene transfer, when cardiac myocytes were stimulated with isoproterenol or NKH477, a water-soluble forskolin analog that directly stimulates AC, expression of ATF3 protein was increased even more, which correlated with reduced expression of PLB. We then showed that AC(VI)-induced ATF3 protein binds to the cAMP-responsive-like element on the PLB promoter and that overexpression of ATF3 in cardiac myocytes inhibits PLB promoter activity. These findings indicate that AC(VI) has effects on gene transcription that are not directly dependent on cAMP generation.