Division of Gastroenterology and Liver Diseases, USC Research Center for Liver Diseases, Keck School of Medicine, University of Southern California, Los Angeles, CA, USA.
Hepatology. 2012 Jun;55(6):1942-53. doi: 10.1002/hep.25594. Epub 2012 Apr 23.
Methionine adenosyltransferases (MATs) are critical enzymes that catalyze the formation of the methyl donor S-adenosyl methionine (SAM). The MAT2A gene, which encodes the catalytic subunit α2, is induced in dedifferentiated liver. We previously demonstrated that MAT2A expression is enhanced in activated hepatic stellate cells (HSCs) and that silencing this gene reduces HSC activation. In this study, we examined the molecular mechanisms responsible for the transcriptional regulation of the MAT2A gene in HSCs. We identified peroxisome proliferator-activated receptor (PPAR) response elements (PPREs) in the rat MAT2A promoter. The PPARγ agonist rosiglitazone (RSG) promoted quiescence in the activated rat HSC cell line (BSC) or culture-activated primary rat HSCs, decreased MAT2A expression and promoter activity, and enhanced PPARγ binding to MAT2A PPREs. In vivo HSC activation in bile duct-ligated rats lowered PPARγ interaction with MAT2A PPREs. Silencing PPARγ increased MAT2A transcription, whereas overexpressing it had the opposite effect, demonstrating that PPARγ negatively controls this gene. Site-directed mutagenesis of PPREs abolished PPARγ recruitment to the MAT2A promoter and its inhibitory effect on MAT2A transcription in quiescent HSCs. PPRE mutations decreased the basal promoter activity of MAT2A in activated HSCs independent of PPARγ, indicating that other factors might be involved in PPRE interaction. We identified PPARβ binding to wild-type but not to mutated PPREs in activated cells. Furthermore, silencing PPARβ inhibited MAT2A expression and promoter activity. Forced expression of MAT2A in RSG-treated HSCs lowered PPARγ and enhanced PPARβ expression, thereby promoting an activated phenotype.
We identified PPARγ as a negative regulator of MAT2A in quiescent HSCs. A switch from quiescence to activation abolishes this control and allows PPARβ to up-regulate MAT2A transcription.
甲硫氨酸腺苷转移酶(MATs)是催化形成甲基供体 S-腺苷甲硫氨酸(SAM)的关键酶。编码催化亚基α2 的 MAT2A 基因在去分化的肝脏中被诱导。我们之前证明,MAT2A 的表达在激活的肝星状细胞(HSCs)中增强,并且沉默该基因可减少 HSC 的激活。在这项研究中,我们研究了负责 HSCs 中 MAT2A 基因转录调控的分子机制。我们在大鼠 MAT2A 启动子中鉴定出过氧化物酶体增殖物激活受体(PPAR)反应元件(PPREs)。PPARγ 激动剂罗格列酮(RSG)促进激活的大鼠 HSC 细胞系(BSC)或培养激活的原代大鼠 HSCs 的静止,降低 MAT2A 的表达和启动子活性,并增强 PPARγ 与 MAT2A PPREs 的结合。胆管结扎大鼠体内 HSC 的激活降低了 PPARγ 与 MAT2A PPREs 的相互作用。沉默 PPARγ 增加了 MAT2A 的转录,而过表达则产生相反的效果,表明 PPARγ 负调控该基因。PPRE 突变消除了 PPARγ 对静止 HSCs 中 MAT2A 启动子的募集及其对 MAT2A 转录的抑制作用。PPRE 突变降低了激活的 HSCs 中 MAT2A 基本启动子活性,而与 PPARγ 无关,表明其他因素可能参与 PPRE 相互作用。我们在激活的细胞中鉴定出 PPARβ 与野生型但不与突变型 PPREs 的结合。此外,沉默 PPARβ 抑制了 MAT2A 的表达和启动子活性。在 RSG 处理的 HSCs 中强制表达 MAT2A 降低了 PPARγ 并增强了 PPARβ 的表达,从而促进了激活表型。
我们鉴定出 PPARγ 是静止 HSCs 中 MAT2A 的负调节剂。从静止到激活的转变消除了这种控制,允许 PPARβ 上调 MAT2A 的转录。