Post-translational modification analysis of Saccharomyces cerevisiae histone methylation enzymes reveals phosphorylation sites of regulatory potential.
Post-translational modification analysis of Saccharomyces cerevisiae histone methylation enzymes reveals phosphorylation sites of regulatory potential.
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DOI：
10.1074/jbc.ra120.015995
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发表时间：
2021-01
期刊：
影响因子：
--
通讯作者：
Wilkins MR
中科院分区：
文献类型：
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作者：
Separovich RJ;Wong MWM;Chapman TR;Slavich E;Hamey JJ;Wilkins MR
组蛋白甲基化是真核生物转录调控的核心。在酿酒酵母中，它由四种甲基转移酶（Set 1 p，Set 2 p，Set 5 p和Dot 1 p）和四种脱甲基酶（Jhd 1 p，Jhd 2 p，Rph 1 p和Gis 1 p）控制。虽然这些酶的组蛋白靶点已被很好地表征，但这些酶与细胞内信号网络的连接以及它们的调节却知之甚少;这也适用于所有其他真核生物。在这里，我们报告的8个酿酒酵母酶的详细表征，并表明，他们携带总共75个磷酸化位点，92个乙酰化位点，和两个泛素化位点。所有的酶都受到磷酸化，虽然脱甲基酶Jhd 1 p和Jhd 2 p分别含有一个和五个位点，而其他酶携带14至36个位点。磷酸化是不存在的或代表性不足的催化和其他领域，但强烈丰富的甲基转移酶的紊乱区域，这表明在蛋白质-蛋白质相互作用的调制中的作用。通过诱变研究，我们发现酸性和无序的N-末端Set 2 p内的磷酸化位点影响H3 K36甲基化水平在体内，说明这些网站的功能的重要性。虽然大多数激酶上游的酵母组蛋白甲基化酶仍然是未知的，我们模拟细胞信号网络和组蛋白为基础的基因调控系统之间的可能的连接，并提出了一个集成的监管结构。我们的研究结果提供了一个基础，为未来的详细探索的作用，特定的激酶和磷酸化位点的调节组蛋白甲基化。
Histone methylation is central to the regulation of eukaryotic transcription. In Saccharomyces cerevisiae, it is controlled by a system of four methyltransferases (Set1p, Set2p, Set5p, and Dot1p) and four demethylases (Jhd1p, Jhd2p, Rph1p, and Gis1p). While the histone targets for these enzymes are well characterized, the connection of the enzymes with the intracellular signaling network and thus their regulation is poorly understood; this also applies to all other eukaryotes. Here we report the detailed characterization of the eight S. cerevisiae enzymes and show that they carry a total of 75 phosphorylation sites, 92 acetylation sites, and two ubiquitination sites. All enzymes are subject to phosphorylation, although demethylases Jhd1p and Jhd2p contained one and five sites respectively, whereas other enzymes carried 14 to 36 sites. Phosphorylation was absent or underrepresented on catalytic and other domains but strongly enriched for regions of disorder on methyltransferases, suggesting a role in the modulation of protein–protein interactions. Through mutagenesis studies, we show that phosphosites within the acidic and disordered N-terminus of Set2p affect H3K36 methylation levels in vivo, illustrating the functional importance of such sites. While most kinases upstream of the yeast histone methylation enzymes remain unknown, we model the possible connections between the cellular signaling network and the histone-based gene regulatory system and propose an integrated regulatory structure. Our results provide a foundation for future, detailed exploration of the role of specific kinases and phosphosites in the regulation of histone methylation.