Department of Medicine, Division of Hematology/Oncology, Case Western Reserve University, Cleveland, OH 44106 , USA.
Cancer Biol Ther. 2009 Sep;8(18):1771-86. doi: 10.4161/cbt.8.18.9323. Epub 2009 Sep 20.
Bax is a pro-apoptotic protein that mediates intrinsic cell-death signaling. Using a yeast-based functional screening approach, we identified interferon gamma receptor beta chain (IFNgammaR2) as a new Bax suppressor. IFNgammaR2 is a component of the IFNgamma receptor complex along with the IFNgammaR alpha chain (IFNgammaR1). Upon IFNgamma binding, a conformational change in the receptor complex occurs that activates the Jak2/STAT1 signaling cascade. We found that the C-terminal region (amino acids 296-337) of IFNgammaR2 (IFNgammaR2(296-337)) contains a novel Bax inhibitory domain. This portion does not contain the Jak2-binding domain; therefore, the antiapoptotic function of IFNgammaR2 is independent of JAK/STAT signaling. IFNgammaR2(296-337) rescued human cells from apoptosis induced by overexpression of Bax but not Bak. Overexpression of IFNgammaR2 (wild type and IFNgammaR2(296-337)) rescued cells from etoposide and staurosporine, which are known to induce Bax-mediated cell death. Interestingly, IFNgammaR2 inhibited apoptosis induced by the BH3-only protein Bim-EL, suggesting that IFNgammaR2 inhibits Bax activation through a BH3-only protein. Bax and IFNgammaR2 were co-immunoprecipitated from cell lysates prepared from HEK293 and DAMI cells. Furthermore, direct binding of purified recombinant proteins of Bax and IFNgammaR2 was also confirmed. Addition of recombinant Bcl-2 protein to cell lysates significantly reduced the interaction of IFNgammaR2 and Bax, suggesting that Bcl-2 and IFNgammaR2 bind a similar domain of Bax. We found that the C-terminal fragment (cytoplasmic domain) of IFNgammaR2 is expressed in human cancer cell lines of megakaryocytic cancer (DAMI), breast cancer (MDA-MD-468), and prostate cancer (PC3 cells). The presence of the C-terminal fragment of IFNgammaR2 may confer on cancer cells resistance to apoptotic stresses. Our discovery of the anti-Bax activity of the cytoplasmic domain of IFNgammaR2 may shed new light on the mechanism of how cell death is controlled by IFNgamma and Bax.
Bax 是一种促进细胞凋亡的蛋白，可介导内在的细胞死亡信号。我们采用基于酵母的功能筛选方法，发现干扰素γ受体β链（IFNγR2）是 Bax 的新抑制因子。IFNγR2 是干扰素γ受体复合物的组成部分，与 IFNγR1 共同构成 IFNγ 受体复合物。IFNγ 结合后，受体复合物发生构象变化，激活 Jak2/STAT1 信号级联。我们发现，IFNγR2 的 C 端区域（氨基酸 296-337）（IFNγR2(296-337)）包含一个新的 Bax 抑制结构域。这部分不包含 Jak2 结合结构域；因此，IFNγR2 的抗凋亡功能不依赖于 JAK/STAT 信号。IFNγR2(296-337) 可挽救 Bax 过表达诱导的人细胞凋亡，但不能挽救 Bak 过表达诱导的细胞凋亡。IFNγR2（野生型和 IFNγR2(296-337)）的过表达可挽救依托泊苷和 staurosporine 诱导的细胞死亡，这两种药物已知可诱导 Bax 介导的细胞死亡。有趣的是，IFNγR2 抑制 Bim-EL 诱导的 BH3 仅有蛋白诱导的细胞凋亡，这表明 IFNγR2 通过 BH3 仅有蛋白抑制 Bax 的激活。从 HEK293 和 DAMI 细胞的细胞裂解物中免疫沉淀 Bax 和 IFNγR2。此外，还证实了纯化的 Bax 和 IFNγR2 重组蛋白的直接结合。将重组 Bcl-2 蛋白添加到细胞裂解物中可显著降低 IFNγR2 和 Bax 的相互作用，表明 Bcl-2 和 IFNγR2 结合 Bax 的相似结构域。我们发现，IFNγR2 的 C 末端片段（细胞质结构域）在巨核细胞癌（DAMI）、乳腺癌（MDA-MD-468）和前列腺癌（PC3 细胞）的人癌细胞系中表达。IFNγR2 的 C 末端片段的存在可能使癌细胞对凋亡应激产生抗性。我们发现 IFNγR2 细胞质结构域的抗 Bax 活性，可能为细胞死亡如何受 IFNγ 和 Bax 控制的机制提供新的认识。
