Moon Haejeong, Baek Dongwon, Lee Boyoung, Prasad D Theertha, Lee Sang Yeol, Cho Moo Je, Lim Chae Oh, Choi Myung Suk, Bahk Jeongdong, Kim Myeong Ok, Hong Jong Chan, Yun Dae-Jin
Division of Applied Life Science (BK21 Program), Gyeongsang National University, Jinju 660-701, Korea.
Biochem Biophys Res Commun. 2002 Jan 11;290(1):457-62. doi: 10.1006/bbrc.2001.6208.
Bax, a mammalian proapoptotic member of the Bcl-2 family, can induce cell death when expressed in yeast or plant cells. To identify plant Bax inhibitors, we cotransformed a soybean cDNA library and the Bax gene into yeast cells and screened for expressed genes that prevented Bax-induced apoptosis. From the Bax-inhibiting genes isolated, ascorbate peroxidase (sAPX) was selected for characterization. The transcription of sAPX in plants was specifically induced by oxidative stress. Moreover, overexpression of sAPX partially suppressed the H(2)O(2)-sensitive phenotype of yeast cytosolic catalase T (Deltactt)- and thermosensitive phenotype of cytochrome c peroxidase (Deltaccp)-deleted mutant cells. Examination of reactive oxygen species (ROS) production using the fluorescence method of dihydrorhodamine 123 oxidation revealed that expression of Bax in yeast cells generated ROS, which was greatly reduced by coexpression with sAPX. Our results collectively suggest that sAPX inhibits the generation of ROS by Bax, which in turn suppresses Bax-induced cell death in yeast.
Bax是Bcl-2家族的一种哺乳动物促凋亡成员，当在酵母或植物细胞中表达时可诱导细胞死亡。为了鉴定植物Bax抑制剂，我们将大豆cDNA文库和Bax基因共转化到酵母细胞中，并筛选能够阻止Bax诱导的细胞凋亡的表达基因。从分离得到的抑制Bax的基因中，选择了抗坏血酸过氧化物酶（sAPX）进行特性分析。植物中sAPX的转录受到氧化应激的特异性诱导。此外，sAPX的过表达部分抑制了酵母胞质过氧化氢酶T（Deltactt）缺失突变体细胞对H₂O₂敏感的表型以及细胞色素c过氧化物酶（Deltaccp）缺失突变体细胞的热敏表型。使用二氢罗丹明123氧化荧光法检测活性氧（ROS）的产生，结果显示Bax在酵母细胞中的表达会产生活性氧，而与sAPX共表达时活性氧会大大减少。我们的结果共同表明，sAPX抑制了Bax诱导的活性氧生成，进而抑制了酵母中Bax诱导的细胞死亡。
