Department of Microbiology & Immunology, George Williams Hooper Foundation, University of California San Francisco, San Francisco, California, USA; TETRAD Graduate Program, University of California San Francisco, San Francisco, California, USA.
Department of Microbiology & Immunology, George Williams Hooper Foundation, University of California San Francisco, San Francisco, California, USA.
J Biol Chem. 2021 Nov;297(5):101276. doi: 10.1016/j.jbc.2021.101276. Epub 2021 Oct 5.
Unique among metazoan repressive histone methyltransferases, G9a and GLP, which chiefly target histone 3 lysine 9 (H3K9), require dimerization for productive H3K9 mono (me1)- and dimethylation (me2) in vivo. Intriguingly, even though each enzyme can independently methylate H3K9, the predominant active form in vivo is a heterodimer of G9a and GLP. How dimerization influences the central H3K9 methyl binding ("reading") and deposition ("writing") activity of G9a and GLP and why heterodimerization is essential in vivo remains opaque. Here, we examine the H3K9me "reading" and "writing" activities of defined, recombinantly produced homo- and heterodimers of G9a and GLP. We find that both reading and writing are significantly enhanced in the heterodimer. Compared with the homodimers, the heterodimer has higher recognition of H3K9me2, and a striking ∼10-fold increased turnover rate for nucleosomal substrates under multiple turnover conditions, which is not evident on histone tail peptide substrates. Cross-linking Mass Spectrometry suggests that differences between the homodimers and the unique activity of the heterodimer may be encoded in altered ground state conformations, as each dimer displays different domain contacts. Our results indicate that heterodimerization may be required to relieve autoinhibition of H3K9me reading and chromatin methylation evident in G9a and GLP homodimers. Relieving this inhibition may be particularly important in early differentiation when large tracts of H3K9me2 are typically deposited by G9a-GLP, which may require a more active form of the enzyme.
与主要靶向组蛋白 3 赖氨酸 9（H3K9）的后生动物抑制性组蛋白甲基转移酶 G9a 和 GLP 不同，它们需要二聚化才能在体内有效地进行 H3K9 单（me1）和二甲基化（me2）。有趣的是，尽管每种酶都可以独立地甲基化 H3K9，但在体内主要的活性形式是 G9a 和 GLP 的异二聚体。二聚化如何影响 G9a 和 GLP 的中央 H3K9 甲基结合（“读取”）和沉积（“写入”）活性，以及为什么异二聚化在体内是必不可少的，仍然不清楚。在这里，我们检查了 G9a 和 GLP 的定义的、重组产生的同二聚体和异二聚体的 H3K9me“读取”和“写入”活性。我们发现，异二聚体的读取和写入都显著增强。与同二聚体相比，异二聚体对 H3K9me2 的识别能力更高，并且在多次周转条件下，核小体底物的周转率显著提高了约 10 倍，而在组蛋白尾部肽底物上则不明显。交联质谱表明，同二聚体之间的差异和异二聚体的独特活性可能编码在改变的基态构象中，因为每个二聚体显示不同的结构域接触。我们的结果表明，异二聚化可能是缓解 G9a 和 GLP 同二聚体中明显的 H3K9me 读取和染色质甲基化的自身抑制所必需的。当 G9a-GLP 通常沉积大量 H3K9me2 时，解除这种抑制可能特别重要，这可能需要酶的更活跃形式。
