Dnmt3 and G9a cooperate for tissue-specific development in zebrafish.
Dnmt3 and G9a cooperate for tissue-specific development in zebrafish.
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DOI：
10.1074/jbc.m109.073676
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发表时间：
2010-02-05
期刊：
影响因子：
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通讯作者：
Jones DA
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文献类型：
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作者：
Rai K;Jafri IF;Chidester S;James SR;Karpf AR;Cairns BR;Jones DA
尽管DNA甲基化对正常的胚胎和组织特异性发育至关重要，但不同的DNA甲基转移酶如何影响组织特异性发育及其靶点仍不清楚。我们在斑马鱼中通过基于反义的morpholino敲低Dnmt3和Dnmt1来解决这个问题。我们的数据显示Dnmt3是正常神经发生所必需的，它的缺失会导致大脑和视网膜的严重缺陷。有趣的是，其他器官如肠道未受影响，这表明Dnmt3的组织特异性需求。此外，Dnmt1基因敲低表型与Dnmt3基因敲低表型的比较表明，这两个家族具有不同的功能。与此观点一致的是，Dnmt1不能补充Dnmt3缺陷，Dnmt3也不能补充Dnmt1缺陷。在Dnmt3的下游，我们发现了一个神经发生调节因子lef1，它是Dnmt3特异性靶基因，在Dnmt3突变体中被去甲基化和上调。敲除lef1可挽救Dnmt3缺失导致的神经发生缺陷。在机制上，我们发现Dnmt3和H3K9甲基转移酶G9a在调节lef1中的合作。此外，就像Dnmt1-Suv39h1的协同作用一样，Dnmt3和G9a似乎在组织特异性发育中共同起作用。G9a敲低而非Suv39h1缺失，表型的dnmt3 morphants和G9a过表达提供了dnmt3 morphants表型的显著拯救，而Suv39h1过表达失败，支持特异性dnmt -组蛋白甲基转移酶网络的概念。与该模型一致的是，dnmt3和g9a变形体中lef1启动子上的H3K9me3水平降低，其敲除挽救了g9a变形体的神经发生缺陷。我们提出了一个模型，其中特定的dnmt -组蛋白甲基转移酶网络被利用来沉默细胞命运的关键调节因子，以组织特异性的方式。
Although DNA methylation is critical for proper embryonic and tissue-specific development, how different DNA methyltransferases affect tissue-specific development and their targets remains unknown. We address this issue in zebrafish through antisense-based morpholino knockdown of Dnmt3 and Dnmt1. Our data reveal that Dnmt3 is required for proper neurogenesis, and its absence results in profound defects in brain and retina. Interestingly, other organs such as intestine remain unaffected suggesting tissue-specific requirements of Dnmt3. Further, comparison of Dnmt1 knockdown phenotypes with those of Dnmt3 suggested that these two families have distinct functions. Consistent with this idea, Dnmt1 failed to complement Dnmt3 deficiency, and Dnmt3 failed to complement Dnmt1 deficiency. Downstream of Dnmt3 we identify a neurogenesis regulator, lef1, as a Dnmt3-specific target gene that is demethylated and up-regulated in dnmt3 morphants. Knockdown of lef1 rescued neurogenesis defects resulting from Dnmt3 absence. Mechanistically, we show cooperation between Dnmt3 and an H3K9 methyltransferase G9a in regulating lef1. Further, like Dnmt1-Suv39h1 cooperativity, Dnmt3 and G9a seemed to function together for tissue-specific development. G9a knockdown, but not Suv39h1 loss, phenocopied dnmt3 morphants and G9a overexpression provided a striking rescue of dnmt3 morphant phenotypes, whereas Suv39h1 overexpression failed, supporting the notion of specific DNMT-histone methyltransferase networks. Consistent with this model, H3K9me3 levels on the lef1 promoter were reduced in both dnmt3 and g9a morphants, and its knockdown rescued neurogenesis defects in g9a morphants. We propose a model wherein specific DNMT-histone methyltransferase networks are utilized to silence critical regulators of cell fate in a tissue-specific manner.