H3K9MTase G9a is essential for the differentiation and growth of tenocytes in vitro
H3K9MTase G9a is essential for the differentiation and growth of tenocytes in vitro
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H3K9MTase G9a 对于肌腱细胞的体外分化和生长至关重要
DOI：
10.1007/s00418-015-1318-2
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发表时间：
2015
期刊：
影响因子：
--
通讯作者：
Nifuji A.
中科院分区：
文献类型：
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作者：
Wada S;Ideno H;Shimada A;Kamiunten T;Nakamura Y;Nakashima K;Kimura H;Shinkai Y;Tachibana M;Nifuji A.
细胞分化由特定的转录因子控制。这些转录因子的功能和表达水平受表观遗传修饰的调节，例如基因组的组蛋白修饰和胞嘧啶甲基化。在肌腱组织中，肌腱特异的转录因子已被证明在肌腱细胞分化的调节中发挥功能作用。然而，表观遗传修饰对肌腱细胞基因表达和分化的影响尚不清楚。在这项研究中，我们研究了腱细胞分化的表观遗传学调控，重点是介导组蛋白3赖氨酸9(H3K9)甲基化的酶。在原代培养的小鼠腱细胞中，G9a、G9a样蛋白(GLP)、PR结构域锌指蛋白2(PRDM2)、SUV39H1、SUV39H2和SETDB1/ESET等6个H3K9甲基转移酶(H3K9MTase)基因均有表达，且在肌腱细胞分化过程中表达水平升高。在小鼠胚胎中，G9a和Prdm2mRNAs在腱细胞前体细胞中表达，这些前体细胞与表达腱细胞特异性标记的腱调节蛋白的细胞重叠或相邻。使用从G9a-FLOX/FLOX小鼠分离的腱细胞，我们通过感染表达Cre的腺病毒来删除G9a。G9a缺失的肌腱细胞的增殖明显低于GFP表达的腺病毒感染的对照细胞。此外，在G9a缺失的肌腱细胞中，肌腱转录因子基因Sclaxis(SCX)、Mohawk(MKX)、Egr1、SIX1和Six2的表达水平均受到抑制。肌腱相关基因Col1a1、肌腱调节蛋白和骨膜蛋白也下调。与此一致的是，Western印迹分析表明，G9a缺失显著抑制了Tenomodrin蛋白的表达。这些结果表明，H3K9MTase G9a的表达对肌腱细胞的分化和生长是必需的，H3K9MTase可能在肌腱形成中发挥重要作用。
Cell differentiation is controlled by specific transcription factors. The functions and expression levels of these transcription factors are regulated by epigenetic modifications, such as histone modifications and cytosine methylation of the genome. In tendon tissue, tendon-specific transcription factors have been shown to play functional roles in the regulation of tenocyte differentiation. However, the effects of epigenetic modifications on gene expression and differentiation in tenocytes are unclear. In this study, we investigated the epigenetic regulation of tenocyte differentiation, focusing on the enzymes mediating histone 3 lysine 9 (H3K9) methylation. In primary mouse tenocytes, six H3K9 methyltransferase (H3K9MTase) genes, i.e.,G9a,G9a-like protein(GLP),PR domain zinc finger protein 2(PRDM2),SUV39H1,SUV39H2, andSETDB1/ESETwere all expressed, with increased mRNA levels observed during tenocyte differentiation. In mouse embryos,G9aandPrdm2mRNAs were expressed in tenocyte precursor cells, which were overlapped with or were adjacent to cells expressing a tenocyte-specific marker, tenomodulin. Using tenocytes isolated from G9a-flox/flox mice, we deleted G9a by infecting the cells with Cre-expressing adenoviruses. Proliferation of G9a-null tenocytes was significantly decreased compared with that of control cells infected with GFP-expressing adenoviruses. Moreover, the expression levels of tendon transcription factors gene, i.e.,Scleraxis(Scx),Mohawk(Mkx),Egr1,Six1, andSix2were all suppressed in G9a-null tenocytes. The tendon-related genesCol1a1,tenomodulin, andperiostinwere also downregulated. Consistent with this, Western blot analysis showed that tenomodulin protein expression was significantly suppressed by G9a deletion. These results suggested that expression of the H3K9MTase G9a was essential for the differentiation and growth of tenocytes and that H3K9MTases may play important roles in tendinogenesis.