Division of Novel Food and Immunochemistry, National Institute of Health Sciences, 1-18-1 Kamiyoga, Setagaya, Tokyo 158-8501, Japan.
J Biol Chem. 2010 Apr 23;285(17):13079-91. doi: 10.1074/jbc.M109.044206. Epub 2010 Feb 22.
Poly(ADP-ribose)polymerase-1 (PARP-1) is thought to be required for apoptosis-inducing factor (AIF) release from mitochondria in caspase-independent apoptosis. The mechanism by which AIF is released through PARP-1 remains unclear. Here, we provide evidence that PARP-1-independent AIF release and cell death are induced by a trienoic fatty acid, alpha-eleostearic acid (alpha-ESA). Alpha-ESA induced the caspase-independent and AIF-initiated apoptotic death of neuronal cell lines, independently of PARP-1 activation. The cell death was inhibited by the MEK inhibitor U0126 and by knockdown of MEK using small interfering RNA. However, inhibitors for JNK, p38 inhibitors, calpain, phospholipase A(2), and phosphatidylinositol 3-kinase, did not block cell death. AIF was translocated to the nucleus after the induction of apoptosis by alpha-ESA in differentiated PC12 cells without activating caspase-3 and PARP-1. The alpha-ESA-mediated cell death was not inhibited by PARP inhibitor 3,4-dihydro-5-[4-(1-piperidinyl)butoxyl]-1(2H)-isoquinoline and by knockdown of PARP-1 using small interfering RNA. Unlike N-methyl-N'-nitro-N-nitrosoguanidine treatment, histone-phosphorylated histone 2AX was not phosphorylated by alpha-ESA, which suggests no DNA damage. Overexpression of Bcl-2 did not inhibit the cell death. alpha-ESA caused a small quantity of superoxide production in the mitochondria, resulting in the reduction of mitochondrial membrane potential, both of which were blocked by a trace amount of alpha-tocopherol localized in the mitochondria. Our results demonstrate that alpha-ESA induces PARP-1-independent AIF release and cell death without activating Bax, cytochrome c, and caspase-3. MEK is also a key molecule, although the link between ERK, AIF release, and cell death remains unknown. Finding molecules that regulate AIF release may be an important therapeutic target for the treatment of neuronal injury.
聚（ADP-核糖）聚合酶 1（PARP-1）被认为是细胞色素 c 和 caspase-3 依赖性细胞凋亡中凋亡诱导因子（AIF）从线粒体释放所必需的。AIF 通过 PARP-1 释放的机制尚不清楚。在这里，我们提供的证据表明，三烯脂肪酸α-桐油酸（α-ESA）诱导 PARP-1 非依赖性 AIF 释放和细胞死亡。α-ESA 诱导神经元细胞系 caspase 非依赖性和 AIF 起始的凋亡性死亡，而不依赖于 PARP-1 的激活。该细胞死亡可被 MEK 抑制剂 U0126 和使用小干扰 RNA 对 MEK 的敲低所抑制。然而，JNK 抑制剂、p38 抑制剂、钙蛋白酶、磷脂酶 A2 和磷脂酰肌醇 3-激酶抑制剂不能阻断细胞死亡。在分化的 PC12 细胞中，α-ESA 诱导细胞凋亡后，AIF 易位到细胞核，而不激活 caspase-3 和 PARP-1。PARP 抑制剂 3,4-二氢-5-[4-（1-哌啶基）丁氧基]-1（2H）-异喹啉和使用小干扰 RNA 对 PARP-1 的敲低都不能抑制 α-ESA 介导的细胞死亡。与 N-甲基-N'-亚硝基-N-亚硝基胍处理不同，α-ESA 不使组蛋白磷酸化组蛋白 2AX 磷酸化，这表明没有 DNA 损伤。Bcl-2 的过表达不能抑制细胞死亡。α-ESA 在线粒体中引起少量的超氧化物产生，导致线粒体膜电位降低，这两者都被线粒体中少量的α-生育酚所阻断。我们的结果表明，α-ESA 诱导 PARP-1 非依赖性 AIF 释放和细胞死亡，而不激活 Bax、细胞色素 c 和 caspase-3。MEK 也是一个关键分子，尽管 ERK、AIF 释放和细胞死亡之间的联系尚不清楚。寻找调节 AIF 释放的分子可能是治疗神经元损伤的重要治疗靶点。
