BH-3-only BIK functions at the endoplasmic reticulum to stimulate cytochrome c release from mitochondria
BH-3-only BIK functions at the endoplasmic reticulum to stimulate cytochrome c release from mitochondria
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DOI：
10.1074/jbc.m201235200
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发表时间：
2002-05-17
影响因子：
4.8
通讯作者：
Shore, GC
中科院分区：
文献类型：
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作者：
Germain, M;Mathai, JP;Shore, GC
P53对细胞凋亡的刺激伴随着BH-3诱导BCL-2家族的唯一促凋亡成员BIK，以及在P53缺失的细胞中异位表达BIK导致细胞色素c从线粒体释放并激活caspase，这依赖于BH-3的功能结构域。BIK的很大一部分被发现插入到内质网(ER)膜中，大部分蛋白质面向胞浆，在其COOH末端含有一个预测的跨膜片段。用细胞色素b(5)的内质网选择性膜锚取代其跨膜片段，限制BIK对该膜的作用，也保留了BIK的细胞色素c释放和诱导细胞死亡的活性。而半胱氨酸天冬氨酸氨基转移酶抑制剂benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone，对BIk诱导的细胞死亡有强烈的抑制作用，但不影响BIk刺激细胞色素c从线粒体外流的能力。这一benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone-insensitive刺激线粒体释放细胞色素c的途径被成功地在体外重组，并确定了这一活性所必需的光膜(ER)和胞浆中的成分。总的来说，这些结果确认BIK是从内质网某个位置操作的线粒体释放细胞色素a的启动者。
Stimulation of apoptosis by p53 is accompanied by induction of the BH-3-only proapoptotic member of the BCL-2 family, BIK, and ectopic expression of BIK in p53-null cells caused the release of cytochrome c from mitochondria and activation of caspases, dependent on a functional BH-3 domain. A significant fraction of BIK, which contains a predicted transmembrane segment at its COOH terminus, was found inserted in the endoplasmic reticulum (ER) membrane, with the bulk of the protein facing the cytosol. Restriction of BIK to this membrane by replacing its transmembrane segment with the ER-selective membrane anchor of cytochrome b(5) also retained the cytochrome c release and cell death-inducing activity of BIK. Whereas induction of cell death by BIK was strongly inhibited by the caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone, the inhibitor was without effect on the ability of BIK to stimulate egress of cytochrome c from mitochondria. This benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone-insensitive pathway for stimulating cytochrome c release from mitochondria by ER BIK was successfully reconstituted in vitro and identified the requirement for components present in the light membrane (ER) and cytosol as necessary for this activity. Collectively, the results identify BIK as an initiator of cytochrome a release from mitochondria operating from a location at the ER.