酿酒酵母NDI1基因编码一个位于线粒体内膜面向基质侧的泛醌氧化还原酶，该酶在线粒体呼吸链中将电子从NADH传递给泛醌，与哺乳动物细胞中的线粒体复合物Ⅰ功能类似。在我们以往的研究中，经序列比对发现酵母NDI1是人的AIF（凋亡诱导因子）和AMID（凋亡诱导因子同源的，与线粒体相连的死亡诱导因子）的同源基因，其过表达在呼吸抑制型培养基上表现出明显的促凋亡活性，也参与了衰老所引起的凋亡过程(Li et al., 2006)。本研究发现，酵母Ndi1参与了所检测的各种凋亡诱导剂，如过氧化氢，锰和乙酸，引起的凋亡，且不依赖于Z-VAD-fmk（一个广谱的caspase抑制剂）的抑制作用及酵母Yca1。尽管Ndi1参与了线粒体的呼吸作用，但它的凋亡活性不依赖于其本身的NADH脱氢酶活性，也不依赖于线粒体电子传递链活性。在凋亡过程中，Ndi1的N端在线粒体中被切割下来，从而形成有活性的Ndi1，该激活的Ndi1从线粒体转移到细胞质中以行使其凋亡功能。我们研究发现没有经切割活化的全长Ndi1在细胞质中的凋亡活性大大降低，推断N端切割活化对Ndi1的完全激活来说是至关重要的。通过构建Ndi1 N端和C端的截短突变体，检测它们的凋亡活性发现，C端对Ndi1的凋亡活性至关重要，N端截短了72个氨基酸残基的Ndi1直接表达在细胞质中完全重复出将野生型Ndi1过表达到线粒体中的凋亡表型，这说明Ndi1的切割很有可能发生在N端的70~80个氨基酸残基附近。本研究的结果表明，在正常的生长条件下，Ndi1将电子传递到线粒体呼吸链中，以启动呼吸过程来生产ATP，为生物体提供能量，但是在面临压力的情况下，它N端的毒性抑制小帽被切割掉，从线粒体释放出来成为一个自杀因子。本研究的结果为深入了解Ndi1在酵母凋亡过程中的分子作用机制提供了实验证据。Ndi1在凋亡过程中的作用既具有保守性，又具有特殊性：和其同源物AIF一样，Ndi1在转移出线粒体之前经过了N端的切割活化，不同之处在于AIF从线粒体中转移出来之后进入细胞核促进染色质凝集和片段化，而Ndi1本身不具备细胞核定位序列，从线粒体中转移出来之后进入细胞质，引发下游凋亡反应。
Saccharomyces cerevisiae NDI1 codes for the internal mitochondrial ubiquinone oxidoreductase, which transfers electrons from NADH to ubiquinone in the respiratory chain. By sequence comparison, we previously revealed that Ndi1 is a yeast homologue of AIF (apoptosis-inducing factor) and AMID (apoptosis-inducing factor-homologous mitochondrion-associated inducer of death)(Li et al., 2006), and Ndi1 overexpression strain displays potent proapoptotic activity in respiration-restricted medium, it is also involved in aging-related apoptosis. Here we show that S. cerevisiae NDI1 is involved in apoptosis induced by various stimuli tested, including H2O2, Mn and acetate acid, independent of Z-VAD-fmk (a caspase inhibitor) inhibition and yeast Yca1. Although Ndi1 also participates in respiration, its proapoptotic property is separable from its original NADH dehydrogenase activity and the mitochondrial electron transport chain (ETC). During apoptosis, the N-terminal of Ndi1 is cleaved off in the mitochondria and this activated form then escapes out to execute its apoptotic function. The N-terminal cleavage appears essential for the manifestation of the full apoptotic activity as the uncleaved form of Ndi1 exhibits much less growth-inhibitory activity. By testing the proapoptotic phenotype of Ndi1-truncated mutants, we found that the C-terminal is critical for its apoptotic activity, and a truncated mutant lack 72 amino acids of N-terminal overexpressed in cytosol completely reproduced the proapoptotic phenotype of wild-type Ndi1 overexpressed in mitochondria. It indicates that this cleavage of Ndi1 may involve about the first 70~80 amino acids of Ndi1. Our results thus indicate an important role of Ndi1 in the switch of life and death fates in yeast: during the normal growth, Ndi1 assimilates electrons to the electron transport chain and initiates the respiration process to make ATP, whereas under stresses, it cleaves the toxicity-sequestering N-terminal cap, gets released from the mitochondria and becomes a cell killer.The results of this study provided experimental evidence to illustrate the molecular mechanism of Ndi1 in promoting apoptosis. The proapoptotic mechanism of Ndi1 is conserved but also special. Ndi1 is N-terminal cleaved before release from mitochondria as its homologue AIF. The difference is that AIF translocates to nucleus to promote chromatin condensation and fragmentation, while Ndi1 lacks the nucleus locating sequence and translocates into cytosol to activate downstream apoptotic factors.
