Regulation of the yeast Yap1p nuclear export signal is mediated by redox signal-induced reversible disulfide bond formation
Regulation of the yeast Yap1p nuclear export signal is mediated by redox signal-induced reversible disulfide bond formation
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DOI：
10.1128/mcb.21.18.6139-6150.2001
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发表时间：
2001-09-01
影响因子：
5.3
通讯作者：
Nomoto, A
中科院分区：
文献类型：
--
作者：
Kuge, S;Arita, M;Nomoto, A
Yap 1 p是酿酒酵母（Saccharomycescerevisiae）氧化应激反应中的一个关键转录因子，在非应激条件下被转运入核和出核。核输出步骤被H2 O2或巯基氧化剂二酰胺特异性抑制，导致Yap 1 p核积累并诱导其靶基因的转录。在这里，我们提供的证据，传感过氧化氢和二酰胺介导的二硫键形成的C-末端富含半胱氨酸的区域（c-CRD），其中包含3个保守的半胱氨酸和核输出信号（内斯）。H2 O2或二酰胺诱导的c-CRD体内氧化与诱导的Yap 1 p核定位相关。这两个都开始在1分钟内应用氧化应激，细胞内的氧化还原状态的硫氧还蛋白和谷胱甘肽的影响之前。半胱氨酸残基在中间区域的Yap 1 p（n-CRD）是需要延长核定位的Yap 1 p响应H2 O2，因此也需要最大的转录活性。使用质谱分析，H2 O2诱导的c-CRD体外氧化被检测为第一个（Cys（598））和第二个（Cys（620））半胱氨酸残基之间的分子内二硫键;该键可被硫氧还蛋白还原。相比之下，二酰胺诱导c-CRD中的每对二硫键，但在这种情况下，n-CRD中的半胱氨酸残基似乎对反应不利。我们的数据提供了证据的氧化还原信号传感的分子机制，通过硫醇-二硫化物氧化还原循环与硫氧还蛋白系统在Yap 1 p内斯。
Yap1p, a crucial transcription factor in the oxidative stress response of Saccharomyces cerevisiae, is transported in and out of the nucleus under nonstress conditions. The nuclear export step is specifically inhibited by H2O2 or the thiol oxidant diamide, resulting in Yap1p nuclear accumulation and induction of transcription of its target genes. Here we provide evidence for sensing of H2O2 and diamide mediated by disulfide bond formation in the C-terminal cysteine-rich region (c-CRD), which contains 3 conserved cysteines and the nuclear export signal (NES). The H2O2 or diamide-induced oxidation of the c-CRD in vivo correlates with induced Yap1p nuclear localization. Both were initiated within 1 min of application of oxidative stress, before the intracellular redox status of thioredoxin and glutathione was affected. The cysteine residues in the middle region of Yap1p (n-CRD) are required for prolonged nuclear localization of Yap1p in response to H2O2 and are thus also required for maximum transcriptional activity. Using mass spectrometry analysis, the H2O2-induced oxidation of the c-CRD in vitro was detected as an intramolecular disulfide linkage between the first (Cys(598)) and second (Cys(620)) cysteine residues; this linkage could be reduced by thioredoxin. In contrast, diamide induced each pair of disulfide linkage in the c-CRD, but in this case the cysteine residues in the n-CRD appeared to be dispensable for the response. Our data provide evidence for molecular mechanisms of redox signal sensing through the thiol-disulfide redox cycle coupled with the thioredoxin system in the Yap1p NES.