MMP-9-dependent proteolysis of histone H3 N-terminal tail (H3NT) is an important mechanism for activation of gene expression during osteoclast differentiation. Like other enzymes targeting their substrates within chromatin structure, MMP-9 enzymatic activity toward H3NT is tightly controlled by histone modifications such as H3K18 acetylation (H3K18ac) and H3K27 monomethylation (H3K27me1). Growing evidence indicates that DNA methylation is another epigenetic mechanism controlling osteoclastogenesis, but whether DNA methylation is also critical for regulating MMP-9-dependent H3NT proteolysis and gene expression remains unknown. We show here that treating RANKL-induced osteoclast progenitor (OCP) cells with the DNMT inhibitor 5-Aza-2′-deoxycytidine (5-Aza-CdR) induces CpG island hypomethylation and facilitates MMP-9 transcription. This increase in MMP-9 expression results in a significant enhancement of H3NT proteolysis and OCP cell differentiation. On the other hand, despite an increase in levels of H3K18ac, treatment with the HDAC inhibitor trichostatin A (TSA) leads to impairment of osteoclastogenic gene expression. Mechanistically, TSA treatment of OCP-induced cells stimulates H3K27ac with accompanying reduction in H3K27me1, which is a key modification to facilitate stable interaction of MMP-9 with nucleosomes for H3NT proteolysis. Moreover, hypomethylated osteoclastogenic genes in 5-Aza-CdR-treated cells remain transcriptionally inactive after TSA treatment, because H3K27 is highly acetylated and cannot be modified by G9a. These findings clearly indicate that DNA methylation and histone modification are important mechanisms in regulating osteoclastogenic gene expression and that their inhibitors can be used as potential therapeutic tools for treating bone disorders.

中文翻译：

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DNMT 和 HDAC 抑制剂调节 MMP-9 依赖性 H3 N 末端尾部蛋白水解和破骨细胞生成

组蛋白 H3 N 末端尾 (H3NT) 的 MMP-9 依赖性蛋白水解是破骨细胞分化过程中基因表达激活的重要机制。与其他针对染色质结构内底物的酶一样，MMP-9 对 H3NT 的酶活性受到组蛋白修饰（例如 H3K18 乙酰化 (H3K18ac) 和 H3K27 单甲基化 (H3K27me1)）的严格控制。越来越多的证据表明 DNA 甲基化是控制破骨细胞生成的另一种表观遗传机制，但 DNA 甲基化是否对于调节 MMP-9 依赖性 H3NT 蛋白水解和基因表达也至关重要仍不清楚。我们在此表明​​，用 DNMT 抑制剂 5-Aza-2'-脱氧胞苷 (5-Aza-CdR) 处理 RANKL 诱导的破骨细胞祖细胞 (OCP) 会诱导 CpG 岛低甲基化并促进 MMP-9 转录。MMP-9 表达的增加导致 H3NT 蛋白水解和 OCP 细胞分化的显着增强。另一方面，尽管 H3K18ac 水平增加，但用 HDAC 抑制剂曲古抑菌素 A (TSA) 治疗会导致破骨细胞基因表达受损。从机制上讲，TSA 处理 OCP 诱导的细胞会刺激 H3K27ac，并伴随 H3K27me1 的减少，这是促进 MMP-9 与核小体稳定相互作用以进行 H3NT 蛋白水解的关键修饰。此外，5-Aza-CdR 处理的细胞中低甲基化的破骨细胞基因在 TSA 处理后仍保持转录失活，因为 H3K27 高度乙酰化并且不能被 G9a 修饰。这些发现清楚地表明DNA甲基化和组蛋白修饰是调节破骨细胞基因表达的重要机制，并且它们的抑制剂可用作治疗骨疾病的潜在治疗工具。