Department of Pharmacology and Laboratory of Aging and Nervous Diseases, Jiangsu Key Laboratory of Translational Research and Therapy for Neuro-Psycho-Diseases, College of Pharmaceutical Sciences, Soochow University School of Pharmaceutical Science, Suzhou, China.
Cell Death Dis. 2015 Jan 29;6(1):e1624. doi: 10.1038/cddis.2014.546.
DRAM1 (DNA damage-regulated autophagy modulator 1) is a TP53 target gene that modulates autophagy and apoptosis. We previously found that DRAM1 increased autophagy flux by promoting lysosomal acidification and protease activation. However, the molecular mechanisms by which DRAM1 regulates apoptosis are not clearly defined. Here we report a novel pathway by which DRAM1 regulates apoptosis involving BAX and lysosomes. A549 or HeLa cells were treated with the mitochondrial complex II inhibitor, 3-nitropropionic acid (3NP), or an anticancer drug, doxorubicin. Changes in the protein and mRNA levels of BAX and DRAM1 and the role of DRAM1 in BAX induction were determined. The interaction between DRAM1 and BAX and its effect on BAX degradation, BAX lysosomal localization, the release of cathepsin B and cytochrome c by BAX and the role of BAX in 3NP- or doxorubicin-induced cell death were studied. The results showed that BAX, a proapoptotic protein, was induced by DRAM1 in a transcription-independent manner. BAX was degraded by autophagy under basal conditions; however, its degradation was inhibited when DRAM1 expression was induced. There was a protein interaction between DRAM1 and BAX and this interaction prolonged the half-life of BAX. Furthermore, upregulated DRAM1 recruited BAX to lysosomes, leading to the release of lysosomal cathepsin B and cleavage of BID (BH3-interacting domain death agonist). BAX mediated the release of mitochondrial cytochrome c, activation of caspase-3 and cell death partially through the lysosome-cathepsin B-tBid pathway. These results indicate that DRAM1 regulates apoptosis by inhibiting BAX degradation. In addition to mitochondria, lysosomes may also be involved in BAX-initiated apoptosis.
DRAM1（DNA 损伤调节自噬调节剂 1）是一种 TP53 靶基因，可调节自噬和细胞凋亡。我们之前发现，DRAM1 通过促进溶酶体酸化和蛋白酶激活来增加自噬通量。然而，DRAM1 调节细胞凋亡的分子机制尚不清楚。在这里，我们报告了一种涉及 BAX 和溶酶体的新型 DRAM1 调节细胞凋亡的途径。用线粒体复合物 II 抑制剂 3-硝基丙酸（3NP）或抗癌药物阿霉素处理 A549 或 HeLa 细胞。测定 BAX 和 DRAM1 的蛋白和 mRNA 水平的变化以及 DRAM1 在 BAX 诱导中的作用。研究了 DRAM1 与 BAX 的相互作用及其对 BAX 降解、BAX 溶酶体定位、BAX 诱导的组织蛋白酶 B 和细胞色素 c 释放以及 BAX 在 3NP 或阿霉素诱导的细胞死亡中的作用。结果表明，促凋亡蛋白 BAX 被 DRAM1 以转录非依赖性方式诱导。BAX 在基础条件下通过自噬降解；然而，当 DRAM1 表达被诱导时，其降解被抑制。DRAM1 与 BAX 之间存在蛋白相互作用，这种相互作用延长了 BAX 的半衰期。此外，上调的 DRAM1 将 BAX 招募到溶酶体，导致溶酶体组织蛋白酶 B 的释放和 BID（BH3 相互作用结构域死亡激动剂）的切割。BAX 通过溶酶体-组织蛋白酶 B-tBid 途径部分介导线粒体细胞色素 c 的释放、半胱天冬酶-3 的激活和细胞死亡。这些结果表明，DRAM1 通过抑制 BAX 降解来调节细胞凋亡。除了线粒体，溶酶体也可能参与 BAX 起始的细胞凋亡。
