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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >Posttranslational control of transcription factor FixK_2, a key regulator for the Bradyrhizobium japonicum-soybean symbiosis
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Posttranslational control of transcription factor FixK_2, a key regulator for the Bradyrhizobium japonicum-soybean symbiosis

机译：转录因子FixK_2的翻译后控制，这是日本根瘤菌与大豆共生的关键调控因子

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摘要

Rhizobial FixK-like proteins play essential roles in activating genes for endosymbiotic life in legume root nodules, such as genes for micro-oxic respiration. In the facultative soybean symbiont, Bradyrhizobium japonicum, the FixK_2 protein is the key player in a complex regulatory network. The fixK_2 gene itself is activated by the 2-component regulatory system FixLJ in response to a moderate decrease of the oxygen tension, and the FixK_2 protein distributes and amplifies this response to the level of approximately 200 target genes. Unlike other members of the cAMP receptor protein family, to which FixK_2 belongs, the FixK_2 protein does not appear to be modulated by small effector molecules. Here, we show that a critical, single cysteine residue (C183) near the DNA-binding domain of FixK_2 confers sensitivity to oxidizing agents and reactive oxygen species. Oxidation-dependent inactivation occurs not only in vitro, as shown with cell-free transcription assays, but also in vivo, as shown by microarray-assisted transcriptome analysis of the FixK_2 regulon. The oxidation mechanism may involve a reversible dimerization by intermolecular disulfide-bridge formation and a direct, irreversible oxidation at the cysteine thiol, depending on the oxidizing agent. Mutational exchange of C183 to alanine renders FixK_2 resistant to oxidation, yet allows full activity, shown again both in vitro and in vivo. We hypothesize that posttranslational modification by reactive oxygen species is a means to counterbalance the cellular pool of active FixK_2, which would otherwise fill unrestrictedly through FixLJ-dependent synthesis.
机译：根瘤菌FixK样蛋白在激活豆科植物根瘤内共生生命的基因（例如用于微氧呼吸的基因）中起着至关重要的作用。在兼性大豆共生体日本根瘤菌中，FixK_2蛋白是复杂调控网络中的关键角色。 fixK_2基因本身被2组分调节系统FixLJ激活，以响应氧张力的适度降低，并且FixK_2蛋白将这种响应分布并放大到大约200个靶基因的水平。与FixK_2所属的cAMP受体蛋白家族的其他成员不同，FixK_2蛋白似乎不受小效应分子的调节。在这里，我们表明，FixK_2的DNA结合域附近的一个关键的单个半胱氨酸残基（C183）赋予了对氧化剂和活性氧物种的敏感性。氧化依赖性失活不仅发生在体外（如无细胞转录分析所示），而且还发生在体内（如FixK_2 regulon的微阵列辅助转录组分析所示）。取决于氧化剂，氧化机理可能涉及通过分子间二硫键桥形成的可逆二聚作用和在半胱氨酸硫醇处的直接，不可逆氧化。 C183向丙氨酸的突变交换使FixK_2具有抗氧化性，但仍具有完整的活性，在体外和体内均再次显示。我们假设通过活性氧进行翻译后修饰是一种抵消细胞中活跃的FixK_2平衡的方法，否则该细胞会通过FixLJ依赖的合成不受限制地填充。

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