Degradation of NFAT5, a transcriptional regulator of osmotic stress-related genes, is a critical event for doxorubicin-induced cytotoxicity in cardiac myocytes
Degradation of NFAT5, a transcriptional regulator of osmotic stress-related genes, is a critical event for doxorubicin-induced cytotoxicity in cardiac myocytes
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DOI：
10.1074/jbc.m609547200
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发表时间：
2007-01-12
影响因子：
4.8
通讯作者：
Azuma, Junichi
中科院分区：
文献类型：
--
作者：
Ito, Takashi;Fujio, Yasushi;Azuma, Junichi
核因子激活的T细胞5（NFAT 5）是NFAT蛋白家族的新成员，最初被鉴定为负责适应高渗胁迫的转录因子。虽然NFAT 5广泛表达，但其生物学功能仍有待阐明，特别是在未暴露于高渗的组织中，包括心脏。在本研究中，我们重点关注NFAT 5对心脏毒性抗肿瘤剂阿霉素（Dox）的心脏保护作用。在培养的心肌细胞中，高渗诱导基因的转录，如牛磺酸转运蛋白（TauT）和钠/肌醇转运蛋白，被Dox下调。有趣的是，在暴露于Dox的心肌细胞中，NFAT 5蛋白而不是mRNA减少。蛋白酶体抑制剂MG-132或蛋白酶体特异性抑制剂1的治疗防止了Dox介导的NFAT 5蛋白的减少。此外，在用MG-132和/或Dox处理的培养的心肌细胞中未检测到泛素缀合的NFAT 5，如通过免疫沉淀测定所评估的，表明Dox通过泛素非依赖性蛋白酶体途径诱导降解。重要的是，通过显性负性NFAT 5的过表达抑制NFAT 5降低细胞活力并增加肌酸激酶向培养基中的渗漏。因此，针对NFAT 5基因的小干扰RNA增强了心肌细胞死亡。这些发现表明，Dox促进了NFAT 5蛋白的降解，降低了心肌细胞的细胞活力。这是首次证明NFAT 5是心肌细胞存活的正调节因子。
Nuclear factor-activated T cell 5 (NFAT5), a novel member of the NFAT family of proteins, was originally identified as a transcriptional factor responsible for adaptation to hyperosmotic stress. Though NFAT5 is ubiquitously expressed, the biological functions of NFAT5 remain to be clarified, especially in the tissues that are not exposed to hypertonicity, including hearts. In the present study, we focused on the cardioprotective roles of NFAT5 against the cardiotoxic anti-tumor agent doxorubicin (Dox). In cultured cardiomyocytes, transcripts of the hypertonicity-inducible genes, such as taurine transporter (TauT) and sodium/myo-inositol transporter, were down-regulated by Dox. Interestingly, NFAT5 protein, but not mRNA, was decreased in cardiomyocytes exposed to Dox. Treatment of proteasome inhibitors, MG-132 or proteasome-specific inhibitor 1, prevented the Dox-mediated decrease of NFAT5 protein. Further, ubiquitin-conjugated NFAT5 was not detected in cultured cardiomyocytes treated with MG-132 and/or Dox, as assessed by immunoprecipitation assay, suggesting Dox-induced degradation through ubiquitin-independent proteasome pathway. Importantly, inhibition of NFAT5 with overexpression of dominantnegative NFAT5 decreased cell viability and increased creatine kinase leakage into culture medium. Consistently, small interfering RNA targeting NFAT5 gene enhanced myocyte death. These findings suggest that Dox promoted the degradation of NFAT5 protein, reducing cell viability in cardiomyocytes. This is the first demonstration that NFAT5 is a positive regulator of cardiomyocyte survival.