A functional genome-wide RNAi screen identifies TAF1 as a regulator for apoptosis in response to genotoxic stress
A functional genome-wide RNAi screen identifies TAF1 as a regulator for apoptosis in response to genotoxic stress
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DOI：
10.1093/nar/gkn506
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发表时间：
2008-09-01
影响因子：
14.9
通讯作者：
Yoshida, Kiyotsugu
中科院分区：
文献类型：
--
作者：
Kimura, Junko;Nguyen, Su Tien;Yoshida, Kiyotsugu
逃避凋亡性细胞死亡是癌症的一个特征;调节这一过程的基因可能是治疗攻击的最佳选择。因此，确定细胞凋亡的关键调节因子是癌症治疗的中心目标。在这里，我们描述了一个功能丧失的屏幕，使用RNA干扰库，以确定诱导细胞凋亡所需的基因。我们使用了一个短发夹RNA表达载体，具有高基因表达沉默活性，含有胎脑cDNA。从遗传毒性应激中存活的细胞被分离以确定对诱导细胞凋亡至关重要的分子的敲低。我们确定了TBP-associated factor 1（TAF 1），这是一个以前被认为是转录机制的重要组成部分的基因。TAF 1的消耗与氧化以及遗传毒性应激诱导的细胞凋亡的实质性衰减相关。微阵列分析进一步表明，在TAF 1沉默的细胞中，许多基因的转录下降。令人惊讶的是，敲低TAF 1表现出p27（Kip 1）表达的显着降低，使细胞抵抗氧化应激诱导的凋亡。这些结果表明，TAF 1通过控制p27（Kip 1）的表达来调节细胞凋亡。我们的系统提供了一种新的方法来确定候选基因，调节细胞凋亡。
Evasion from apoptotic cell death is a characteristic of cancer; genes that modulate this process may be optimal for therapeutic attack. Identifying key regulators of apoptosis is thus a central goal in cancer therapy. Here, we describe a loss-of-function screen that uses RNA interference libraries to identify genes required for induction of apoptosis. We used a short-hairpin RNA expressing vector with high gene-expression silencing activity that contained fetal brain cDNAs. Survived cells from genotoxic stress were isolated to determine knock-down of molecules that are crucial for induction of apoptosis. We identified TBP-associated factor 1 (TAF1), a gene previously implicated as an essential component of transcription machinery. Depletion of TAF1 was associated with substantial attenuation of apoptosis induced by oxidative as well as genotoxic stress. Microarray analysis further demonstrated that a number of genes were transcriptionally declined in cells silenced for TAF1. Surprisingly, knocking down TAF1 exhibited a marked decrease in p27(Kip1) expression, allowing cells resistant from oxidative stress-induced apoptosis. These results suggest that TAF1 regulates apoptosis by controlling p27(Kip1) expression. Our system provides a novel approach to identifying candidate genes that modulate apoptosis.