The response regulator-like protein Pos9/Skn7 ofSaccharomyces cerevisiae is involved in oxidative stress resistance
The response regulator-like protein Pos9/Skn7 ofSaccharomyces cerevisiae is involved in oxidative stress resistance
复制标题
DOI：
10.1007/bf02208613
复制
发表时间：
1996-03
期刊：
影响因子：
2.5
通讯作者：
B. Krems;C. Charizanis;K. Entian
中科院分区：
文献类型：
--
作者：
B. Krems;C. Charizanis;K. Entian
我们已经分离出对氧化应激敏感性增加的酵母突变体。所有pos 9突变体（posfor过氧化物敏感性）对甲基紫精、高压氧或过氧化氢超敏感，但在所有其他测试条件下生长与野生型相似。经分离和测序证实，该基因与SKN 7基因完全相同。预测的Skn 7/Pos 9蛋白具有与原核反应调节因子高度同源的结构域。这些调节蛋白是称为“双组分系统”的简单信号级联的一部分，其中组氨酸激酶的磷酸化信号被转移到应答调节剂的保守天冬氨酸残基。为了测试Skn 7/Pos 9蛋白的相应天冬氨酸残基在氧化应激中的功能作用，我们在体外将该残基诱变为丙氨酸、精氨酸和谷氨酸。只有谷氨酸等位基因（D427到E）能够拯救pos 9突变体的过氧化氢敏感性。通过与Gal 4 DNA结合结构域的融合实验，我们确定了分离的反应调节因子样结构域作为一种新的真核生物结构域足以激活基因。而这种混合蛋白激活转录的alacZ报告基因在有氧条件下，没有激活观察到在厌氧条件下，表明反应调节域参与信号传导反应。双杂交研究还表明Pos 9蛋白的寡聚化。我们的研究结果表明，一个双组分系统参与酵母的氧化应激反应。
We have isolated mutants ofSaccharomyces cerevisiaewith an increased sensitivity to oxidative stress. Allpos9mutants (posfor peroxide sensitivity) were hypersensitive to methylviologene, hyperbaric oxygen or hydrogen peroxide, but grew similarly to the wild-type under all other conditions tested. Isolation and sequencing of the respectivePOS9gene revealed that it was identical toSKN7. The predicted Skn7/Pos9 protein possesses a domain with high homology to prokaryotic response regulators. These regulatory proteins are part of a simple signalling cascade termed a “two-component system”, where a phosphorylation signal of a histidine kinase is transferred to a conserved aspartate residue of the response regulator. To test the functional role of the respective aspartate residue of Skn7/Pos9 protein in oxidative stress, we mutagenized this residue in vitro to alanine, arginine and glutamate. Only the glutamate allele (D427to E) was able to rescue the hydrogen peroxide-sensitivity ofpos9mutants. By fusion experiments with the Gal4 DNA-binding domain we identified the isolated response regulator-like domain as a novel eukaryotic domain sufficient for gene activation. Whereas this hybrid protein activated transcription of alacZreporter gene under aerobic conditions, no activation was observed under anaerobic conditions, indicating that the response regulator domain is involved in a signalling reaction. Two-hybrid investigations also suggest an oligomerization of the Pos9 protein. Our results indicate that a two-component system is involved in the oxidative-stress response of yeast.