Modulation of anti-cancer drug sensitivity through the regulation of mitochondrial activity by adenylate kinase 4.
Modulation of anti-cancer drug sensitivity through the regulation of mitochondrial activity by adenylate kinase 4.
复制标题
DOI：
10.1186/s13046-016-0322-2
复制
发表时间：
2016-03-16
期刊：
影响因子：
--
通讯作者：
Sakaida I
中科院分区：
文献类型：
--
作者：
Fujisawa K;Terai S;Takami T;Yamamoto N;Yamasaki T;Matsumoto T;Yamaguchi K;Owada Y;Nishina H;Noma T;Sakaida I
腺苷酸激酶是活细胞高能磷酸转移反应中的关键酶。这种酶的一个亚型，腺苷酸激酶4(AK4)，定位于线粒体基质中，被认为与应激、耐药、癌症的恶性转化和ATP调节有关。然而，AK4功能的分子基础仍有待确定。将AK4小干扰RNA(SiRNA)、AK4短发夹状RNA(ShRNA)、对照shRNA、AK4表达载体和对照表达载体瞬时导入HeLa细胞，检测AK4表达对细胞增殖、对抗癌药物敏感性、代谢产物、基因表达和线粒体活性的影响。AK4基因敲除细胞经短发夹状RNA处理后，ATP产量增加，对缺氧和抗癌药物顺铂(CDDP)表现出更高的敏感性。裸鼠皮下移植AK4基因敲除细胞后发现，移植细胞对顺铂的反应表现出较慢的增殖速度和较小的肿瘤体积。AK4基因敲除细胞在FCCP处理后耗氧率增加，而AK4过表达则降低耗氧率。代谢组学分析表明，AK4基因敲除细胞中三羧酸循环中间产物富马酸和苹果酸的水平增加，而AK4过表达则降低了它们的水平。电子显微镜检测到AK4基因敲除细胞线粒体数量增加。基因芯片分析检测到TCA循环中的两个关键酶--琥珀酸脱氢酶A(SDHA)和氧化戊二酸脱氢酶L(OGDHL)的基因表达增加，这两种酶是琥珀酸脱氢酶复合体和氧化戊二酸脱氢酶复合体的组成部分，支持代谢组学结果。我们发现AK4参与耐缺氧、抗肿瘤药物耐药和线粒体活性的调节。这些发现为通过控制AK4的表达而进行有效的抗癌治疗提供了一个新的潜在靶点。本文的在线版本(doi：10.1186/s130460160322-2)包含补充材料，授权用户可以使用。
Adenylate kinase is a key enzyme in the high-energy phosphoryl transfer reaction in living cells. An isoform of this enzyme, adenylate kinase 4 (AK4), is localized in the mitochondrial matrix and is believed to be involved in stress, drug resistance, malignant transformation in cancer, and ATP regulation. However, the molecular basis for the AK4 functions remained to be determined. HeLa cells were transiently transfected with an AK4 small interfering RNA (siRNA), an AK4 short hairpin RNA (shRNA) plasmid, a control shRNA plasmid, an AK4 expression vector, and a control expression vector to examine the effect of the AK4 expression on cell proliferation, sensitivity to anti-cancer drug, metabolome, gene expression, and mitochondrial activity. AK4 knockdown cells treated with short hairpin RNA increased ATP production and showed greater sensitivity to hypoxia and anti-cancer drug, cis-diamminedichloro-platinum (II) (CDDP). Subcutaneous grafting AK4 knockdown cells into nude mice revealed that the grafted cells exhibited both slower proliferation and reduced the tumor sizes in response to CDDP. AK4 knockdown cell showed a increased oxygen consumption rate with FCCP treatment, while AK4 overexpression lowered it. Metabolome analysis showed the increased levels of the tricarboxylic acid cycle intermediates, fumarate and malate in AK4 knockdown cells, while AK4 overexpression lowered them. Electron microscopy detected the increased mitochondrial numbers in AK4 knockdown cells. Microarray analysis detected the increased gene expression of two key enzymes in TCA cycle, succinate dehydrogenase A (SDHA) and oxoglutarate dehydrogenease L (OGDHL), which are components of SDH complex and OGDH complex, supporting the metabolomic results. We found that AK4 was involved in hypoxia tolerance, resistance to anti-tumor drug, and the regulation of mitochondrial activity. These findings provide a new potential target for efficient anticancer therapies by controlling AK4 expression. The online version of this article (doi:10.1186/s13046-016-0322-2) contains supplementary material, which is available to authorized users.