Regulation of PPARgamma and Adipogenesis by MLL3/MLL4 complex

MLL3/MLL4 复合物对 PPARgamma 和脂肪生成的调节

• 批准号: 8939678
• 负责人: Kai Ge
• 金额: $ 72.5万
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8939678
• 关键词: Accounting; Binding; Biological; Biological Models; CCAAT-Enhancer-Binding Proteins; Cell Differentiation process; Cell Nucleus; Cells; Complex; Defect; Diabetes Mellitus; Embryo; Embryonic Development; Enhancers; Enzymes; Exhibits; Fatty acid glycerol esters; Gene Expression; Generations; Genomics; Histones; Knock-out; Lead; Ligands; Mediator of activation protein; Methyltransferase; Molecular; Mono-S; Morbidity - disease rate; Mus; Non-Insulin-Dependent Diabetes Mellitus; Nuclear Protein; Nuclear Receptors; Obesity; PPAR gamma; Peroxisome Proliferator-Activated Receptors; Physiological; Play; Polymerase; Proteins; Regulation; Risk Factors; Role; Staging; Tissues; activating transcription factor; cell type; embryonic stem cell; lipid biosynthesis; member; mortality; myogenesis; novel; novel strategies; obesity treatment; protein complex; stem cell differentiation; transcription factor

My lab previously identified H3K4 methyltransferases MLL3 and MLL4 (Cho YW, JBC 2007) and H3K27 demethylases UTX and JMJD3 (Hong S, PNAS 2007). Using a nuclear protein PTIP as the bait, we isolated from cell nuclei a protein complex that contains H3K4 methyltransferases MLL3/MLL4, H3K27 demethylase UTX, PTIP and a novel protein PA1 (Cho YW, JBC 2007). Further, we show that PTIP is required for PPARγ and C/EBPα expression and adipogenesis (Cho, YW, Cell Metab 2009). To understand the physiological roles of MLL3/MLL4 and associated factors, we have knocked out MLL3, MLL4, UTX and PA1 in mice. Because of their embryonic lethality, we have generated conditional knockout (KO) of MLL4, UTX and PA1. By crossing these conditional KO with Myf5-Cre mice, we found that MLL4, PTIP and PA1 are essential for adipogenesis while UTX is dispensable. To investigate UTX function, we turned to mouse embryonic stem (ES) cells. We found that UTX controls ES cell differentiation and early embryonic development independent of H3K27 demethylase activity (Wang C, PNAS 2012). To understand the biological role of the H3K27 demethylase activity of UTX, we have generated enzyme-dead UTX knockin mice. We recently identify MLL3/MLL4 as H3K4 mono- and di-methyltransferases that are essential for enhancer activation during cell differentiation (Lee JE, eLife 2013). Enhancers play a central role in cell-type-specific gene expression and are marked by H3K4me1/2. Active enhancers are further marked by H3K27ac. However, the methyltransferases responsible for H3K4me1/2 on enhancers remain elusive. Furthermore, how these enzymes function on enhancers to regulate cell-type-specific gene expression is unclear. We identify MLL4 as a major mammalian H3K4 mono- and di-methyltransferase with partial functional redundancy with MLL3. Using adipogenesis and myogenesis as model systems, we show that MLL4 exhibits cell-type- and differentiation-stage-specific genomic binding and is predominantly localized on enhancers. MLL4 co-localizes with lineage-determining transcription factors (TFs) on active enhancers during differentiation. Deletion of MLL4 markedly decreases H3K4me1/2, H3K27ac, Mediator and Polymerase II (Pol II) levels on enhancers and leads to severe defects in cell-type-specific gene expression and cell differentiation. Together, these findings identify MLL3/MLL4 as major mammalian H3K4 mono- and di-methyltransferases essential for enhancer activation during cell differentiation.

我的实验室先前鉴定了H3 K4甲基转移酶MLL 3和MLL 4（Cho YW，JBC 2007）和H3 K27脱甲基酶UTX和JMJD 3（Hong S，PNAS 2007）。使用核蛋白PTIP作为诱饵，我们从细胞核中分离了含有H3 K4甲基转移酶MLL 3/MLL 4、H3 K27脱甲基酶UTX、PTIP和新蛋白PA 1的蛋白复合物（Cho YW，JBC 2007）。此外，我们表明PTIP是PPAR和C/EBP表达和脂肪形成所需的（Cho，YW，Cell Metab 2009）。为了了解MLL 3/MLL 4和相关因子的生理作用，我们在小鼠中敲除了MLL 3、MLL 4、UTX和PA 1。由于它们的胚胎致死性，我们已经产生了MLL 4、UTX和PA 1的条件性敲除（KO）。通过将这些条件性KO与Myf 5-Cre小鼠杂交，我们发现MLL 4、PTIP和PA 1对于脂肪形成是必需的，而UTX是必需的。为了研究UTX的功能，我们转向小鼠胚胎干细胞（ES）。我们发现UTX控制ES细胞分化和早期胚胎发育，而不依赖于H3 K27脱甲基酶活性（Wang C，PNAS 2012）。为了理解UTX的H3 K27脱甲基酶活性的生物学作用，我们产生了酶死亡的UTX敲入小鼠。我们最近将MLL 3/MLL 4鉴定为H3 K4单甲基转移酶和二甲基转移酶，其对于细胞分化期间的增强子活化是必需的（Lee JE，eLife 2013）。增强子在细胞类型特异性基因表达中起核心作用，并由H3 K4 me 1/2标记。活性增强子进一步由H3 K27 ac标记。然而，负责增强子上的H3 K4 me 1/2的甲基转移酶仍然难以捉摸。此外，这些酶如何在增强子上发挥作用以调节细胞类型特异性基因表达尚不清楚。我们确定MLL 4作为一个主要的哺乳动物H3 K4单-和二甲基转移酶与MLL 3的部分功能冗余。使用脂肪生成和肌生成作为模型系统，我们表明MLL 4表现出细胞类型和分化阶段特异性基因组结合，并主要定位于增强子上。MLL 4在分化期间与谱系决定转录因子（TF）共定位在活性增强子上。MLL 4的缺失显著降低增强子上的H3 K4 me 1/2、H3 K27 ac、介体和聚合酶II（Pol II）水平，并导致细胞类型特异性基因表达和细胞分化的严重缺陷。总之，这些发现将MLL 3/MLL 4鉴定为细胞分化期间增强子活化所必需的主要哺乳动物H3 K4单甲基转移酶和二甲基转移酶。