Direct methylation of FXR by Set7/9, a lysine methyltransferase, regulates the expression of FXR target genes
Direct methylation of FXR by Set7/9, a lysine methyltransferase, regulates the expression of FXR target genes
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DOI：
10.1152/ajpgi.00441.2011
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发表时间：
2012-05-01
影响因子：
4.5
通讯作者：
Suchy, Frederick J.
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文献类型：
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作者：
Balasubramaniyan, Natarajan;Ananthanarayanan, Meena;Suchy, Frederick J.
Balasubramaniyan N，Ananthanarayanan M，Suchy FJ. Set 7/9（一种赖氨酸甲基转移酶）对FXR的直接甲基化调节FXR靶基因的表达。美国生理学杂志胃肠和肝脏生理学302：G937-G947，2012年。首次发表于2012年2月16日; doi：10.1152/ajpgi.00441.2011。法尼醇X受体（FXR）是一种配体（胆汁酸）依赖性核受体，调节参与胆汁酸稳态各个方面的靶基因。在配体结合后，FXR募集一系列共激活因子和相关蛋白，其中一些具有修饰组蛋白或甚至转录复合物组分的内在酶活性。在这项研究中，我们显示了在FXR反应元件（FXRE）的Set 7/9甲基转移酶的染色质占有率和FXR在体内和体外赖氨酸206的直接甲基化。Huh-7肝细胞系中Set 7/9的siRNA消耗减少了FXR靶基因、短异源二聚体伴侣（SHP）和胆盐输出泵（BSEP）的内源性mRNA。将FXR的K206处的甲基化位点突变为精氨酸阻止了Set 7/9的甲基化。泛甲基赖氨酸抗体识别野生型FXR，但不识别K206 R突变体形式。电迁移率变化测定表明，Set 7/9的甲基化增强了FXR/视黄酸X受体-α与FXRE的结合。通过免疫共沉淀、GST pull down和哺乳动物双杂交实验证实了FXR铰链区（含K206）与Set 7/9的相互作用。Set 7/9在Huh-7细胞中的过表达以配体依赖性方式显著增强野生型FXR对SHP和BSEP启动子的反式激活，但对K206 R突变型FXR没有增强作用。甲基转移酶活性缺陷的Set 7/9突变体也不能有效增加BSEP启动子的反式激活。这些研究表明，通过Set 7/9的FXR的翻译后甲基化有助于FXR靶基因的转录激活。
Balasubramaniyan N, Ananthanarayanan M, Suchy FJ. Direct methylation of FXR by Set7/9, a lysine methyltransferase, regulates the expression of FXR target genes. Am J Physiol Gastrointest Liver Physiol 302: G937-G947, 2012. First published February 16, 2012; doi:10.1152/ajpgi.00441.2011.-The farnesoid X receptor (FXR) is a ligand (bile acid)-dependent nuclear receptor that regulates target genes involved in every aspect of bile acid homeostasis. Upon binding of ligand, FXR recruits an array of coactivators and associated proteins, some of which have intrinsic enzymatic activity that modify histones or even components of the transcriptional complex. In this study, we show chromatin occupancy by the Set7/9 methyltransferase at the FXR response element (FXRE) and direct methylation of FXR in vivo and in vitro at lysine 206. siRNA depletion of Set7/9 in the Huh-7 liver cell line decreased endogenous mRNAs of the FXR target genes, the short heterodimer partner (SHP) and bile salt export pump (BSEP). Mutation of the methylation site at K206 of FXR to an arginine prevented methylation by Set7/9. A pan-methyllysine antibody recognized the wild-type FXR but not the K206R mutant form. An electromobility shift assay showed that methylation by Set7/9 enhanced binding of FXR/retinoic X receptor-alpha to the FXRE. Interaction between hinge domain of FXR (containing K206) and Set7/9 was confirmed by coimmunoprecipitation, GST pull down, and mammalian two-hybrid experiments. Set7/9 overexpression in Huh-7 cells significantly enhanced transactivation of the SHP and BSEP promoters in a ligand-dependent fashion by wild-type FXR but not the K206R mutant FXR. A Set7/9 mutant deficient in methyltransferase activity was also not effective in increasing transactivation of the BSEP promoter. These studies demonstrate that posttranslational methylation of FXR by Set7/9 contributes to the transcriptional activation of FXR-target genes.