State Key Laboratory of Molecular Biology, Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, 320 Yueyang Road, Shanghai 200031, China.
J Cell Biochem. 2012 May;113(5):1663-70. doi: 10.1002/jcb.24035.
Jumonji C-terminal (JmjC) domain-containing proteins are protein hydroxylases and histone demethylases that control gene expression. Jumonji domain-containing protein 6 (Jmjd6) is indispensable for embryonic development and has both histone arginine demethylase and lysyl-hydroxylase activities. The protein undergoes post-translational homo-oligomerization, but the underlying mechanism remains unknown. In this study, we examined the enzymatic activity of Jmjd6 and uncovered the mechanism underlying its homo-oligomerization. An in vitro enzymatic assay monitored by matrix-assisted laser desorption-ionization time-of-flight (MALDI-TOF) mass spectrometry indicates that Jmjd6 is unable to remove the methyl group from histone arginine residues but can hydroxylate the histone H4 tail at lysine residues in a 2-oxoglutarate (2-OG)- and Fe (II)-dependent manner. A mutational analysis reveals that the homo-oligomerization of Jmjd6 requires its enzymatic activity and the N- and C-termini. Using an in vitro enzymatic assay, we further demonstrate that Jmjd6 can hydroxylate its N-terminus but not its C-terminus. In summary, we did not detect arginine demethylase activity for Jmjd6, but we did confirm that it could catalyze the lysyl-hydroxylation of histone peptides. In addition, we demonstrated that the homo-oligomerization of Jmjd6 requires its own enzymatic activity and the N- and C-termini. We propose that Jmjd6 forms intermolecular covalent bonds between its N- and C-termini via autohydroxylation.
Jumonji C 末端（JmjC）结构域蛋白是一类蛋白羟化酶和组蛋白去甲基酶，它们可以调控基因表达。Jumonji 结构域蛋白 6（Jmjd6）在胚胎发育过程中不可或缺，具有组蛋白精氨酸去甲基酶和赖氨酸羟化酶活性。该蛋白可发生翻译后同源寡聚化，但具体机制尚不清楚。本研究通过检测 Jmjd6 的酶活性，揭示了其同源寡聚化的机制。基质辅助激光解吸电离飞行时间（MALDI-TOF）质谱分析的体外酶促实验表明，Jmjd6 无法去除组蛋白精氨酸残基上的甲基，但可以在 2-氧戊二酸（2-OG）和 Fe（II）依赖性条件下，羟化组蛋白 H4 尾部赖氨酸残基。突变分析显示，Jmjd6 的同源寡聚化需要其酶活性和 N 端及 C 端。通过体外酶促实验，我们进一步证实 Jmjd6 可以羟化其 N 端，但不能羟化 C 端。综上所述，我们并未检测到 Jmjd6 的精氨酸去甲基酶活性，但证实其可以催化组蛋白肽的赖氨酸羟化。此外，我们证实 Jmjd6 的同源寡聚化需要其自身的酶活性以及 N 端和 C 端。我们提出 Jmjd6 通过自身的羟化作用在 N 端和 C 端之间形成了分子间的共价键。
