Hsp70-DnaJ chaperone pair prevents nitric oxide- and CHOP-induced apoptosis by inhibiting translocation of Bax to mitochondria
Hsp70-DnaJ chaperone pair prevents nitric oxide- and CHOP-induced apoptosis by inhibiting translocation of Bax to mitochondria
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DOI：
10.1038/sj.cdd.4401369
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发表时间：
2004-04-01
影响因子：
12.4
通讯作者：
Mori, M
中科院分区：
文献类型：
--
作者：
Gotoh, T;Terada, K;Mori, M
我们报道了内质网（ER）应激途径，涉及CHOP，C/EBP转录因子家族的成员，在一氧化氮（NO）介导的巨噬细胞和胰腺β细胞凋亡中起着关键作用。我们还发现，hsp 70和dj 1（hsp 40/hdj-1）或dj 2（HSDJ/hdj-2）的胞质伴侣对阻止NO介导的细胞色素c从线粒体释放的凋亡上游。为了分析分子伴侣对在防止细胞凋亡中的作用，建立了稳定表达hsp 70和dj 1或dj 2的RAW 264.7巨噬细胞。分子伴侣对阻止CHOP诱导下游的LPS/IFN-γ诱导的和NO介导的细胞凋亡。缺乏ATP酶结构域或C-末端EEVD序列的hsp 70突变蛋白不能有效地阻止CHOP诱导的细胞凋亡。缺乏C-末端异戊烯化CaaX基序的突变体dj 2也是无效的。当用LPS/IFN-γ处理野生型RAW 264.7细胞时，诱导NO介导的凋亡，并且促凋亡Bcl-2家族蛋白Bax从胞质溶胶易位到线粒体。在稳定表达hsp 70/dj 2的细胞和CHOP敲除细胞中，这种易位被阻止。CHOP在野生型细胞中的过表达也诱导Bax易位，并且这种易位在表达hsp 70/dj 2的细胞中被阻止。CHOP诱导的细胞凋亡被Bax敲低所阻止。免疫共沉淀实验表明，Bax与热休克蛋白70和dj 1/dj 2相互作用。hsp 70的ATPase结构域是与Bax结合所必需的。这些结果表明，CHOP诱导的细胞凋亡是由Bax从胞质溶胶易位到线粒体介导的，hsp 70/dj 1或dj 2伴侣对通过与Bax相互作用并阻止易位到线粒体来防止细胞凋亡。
We reported that the endoplasmic reticulum ( ER) stress pathway involving CHOP, a member of the C/EBP transcription factor family, plays a key role in nitric oxide (NO)-mediated apoptosis of macrophages and pancreatic beta cells. We also showed that the cytosolic chaperone pair of hsp70 and dj1 (hsp40/hdj-1) or dj2 (HSDJ/hdj-2) prevents NO-mediated apoptosis upstream of cytochrome c release from mitochondria. To analyze roles of the chaperone pair in preventing apoptosis, RAW 264.7 macrophages stably expressing hsp70 and dj1 or dj2 were established. The chaperone pair prevented LPS/IFN-gamma-induced and NO-mediated apoptosis downstream of CHOP induction. hsp70 mutant protein lacking the ATPase domain or the C-terminal EEVD sequence were not effective in preventing CHOP-induced apoptosis. A mutant dj2 lacking the C-terminal prenylation CaaX motif, was also not effective. When wild-type RAW 264.7 cells were treated with LPS/IFN-gamma, NO-mediated apoptosis was induced, and proapoptotic Bcl-2 family protein Bax was translocated from cytosol to mitochondria. This translocation was prevented in cells stably expressing hsp70/dj2, and in CHOP knockout cells. Overexpression of CHOP in wild-type cells also induced translocation of Bax and this translocation was prevented in cells expressing hsp70/dj2. CHOP-induced apoptosis was prevented by Bax knock-down. Coimmunoprecipitation experiments showed that Bax interacts with both hsp70 and dj1/dj2. ATPase domain of hsp70 was necessary for the binding with Bax. These findings indicate that CHOP-induced apoptosis is mediated by translocation of Bax from the cytosol to the mitochondria, and hsp70/dj1 or dj2 chaperone pair prevents apoptosis by interacting with Bax and preventing translocation to the mitochondria.