MLL3/4-complexes in nuclear receptor-mediated metabolism

描述(申请人提供)：胆汁酸(BA)信号的动态平衡调节需要一个复杂的核受体(NRs)网络，但它们的辅助激活因子重塑染色质和调节基因转录以响应BA信号的机制还知之甚少。我们对NR共激活子ASC-2的发现导致了我们随后分离纯化了“ASC-2-复合体”(ASCOM)，这是第一个包含H3赖氨酸4甲基转移酶(H3K4MT)MLL3或其类似物MLL4的哺乳动物复合体。后来，又发现Ascom含有H3-赖氨酸27-脱甲基酶(H3K27 DM)UTX。三甲基化的H3K4和三甲基化的H3K27分别标记转录活性和非活性染色质。因此，Ascom含有两种类型的酶，它们与转录激活有关。令人兴奋的是，我们发现Ascom的主要生理功能是在各种不同的条件下调节代谢，这主要是由于Asc-2能够将Ascom招募到多个代谢NRs，包括PPAR？、LXR和FXR，即BAS的NR。特别是，我们发现Ascom在调节BA合成的过程中是FXR的关键辅助激活因子。一致地，我们的MLL3突变小鼠最显著的表型之一是BA水平显著增加，这表明MLL3介导的Ascom的H3K4MT活性对于维持BA的动态平衡是必不可少的。有趣的是，我们的MLL3突变小鼠也显示出良好的代谢特征，我们认为这是由于Ascom拮抗Bas的质膜受体TGR5信号的能力存在缺陷。TGR5激活一条信号通路，导致依赖于环磷酸腺苷的甲状腺激素激活酶2碘甲腺原氨酸脱碘酶(D2)表达上调，并促进胰岛素促分泌剂--胰高血糖素样肽-1(GLP-1)的分泌，从而增加能量消耗，改善血糖稳态。我们的初步结果表明，Ascom不仅通过降低BA水平抑制TGR5信号，而且直接通过编码TGR5信号的关键修饰物DPP4(DPP4)的基因调节，DPP4使GLP-1失活。综上所述，这些结果支持了本研究的中心假说：Ascom通过控制调节BA代谢和信号的多个途径中基因的表达，发挥着‘BA信号的动态平衡调节’的主要辅助激活作用。此次更新有两个目标：1)我们将利用上一次资助期间的发现，测试ASCOM在NR激活(主要是FXR和RAR)中的作用机制。2)通过重点研究Ascom在BA内稳态中的作用，我们将建立一个新的范式来理解Ascom的不同代谢作用。这是一项综合良好的研究，因为研究的第一部分对于理解Ascom在BA动态平衡和信号转导中的功能的分子基础至关重要(研究的第二部分)。我们将利用生物化学、细胞和遗传方法的组合，从三个具体目标来解决这两个问题。这项研究将有助于我们了解NRs调控转录和代谢的分子基础。

DESCRIPTION (provided by applicant): The homeostatic regulation of bile acid (BA) signaling requires a complex network of nuclear receptors (NRs), but their coactivators that remodel chromatin and regulate gene transcription in response to BA signaling are poorly understood. Our discovery of the NR coactivator ASC-2 led to our subsequent purification of 'ASC-2- complex' (ASCOM), the first mammalian complex that contains the H3 lysine 4 methyltransferase (H3K4MT) MLL3 or its paralogue MLL4. Later, ASCOM has also been found to contain the H3-lysine 27-demethylase (H3K27DM) UTX. Trimethylated H3K4 and trimethylated H3K27 mark transcriptionally active and inactive chromatin, respectively. Thus, ASCOM contains two types of enzymes that are linked to transcriptional activation. Excitingly, we found that the major physiological function of ASCOM is to regulate metabolism under a variety of different conditions primarily attributed by the ability of ASC-2 to recruit ASCOM to multiple metabolic NRs, including PPAR?, LXRs, and FXR, the NR for BAs. In particular, we discovered that ASCOM functions as a critical coactivator for FXR in regulating BA synthesis. Consistently, one of the most salient phenotypes of our MLL3 mutant mice was a significant increase in BA levels, suggesting that MLL3-mediated H3K4MT activity of ASCOM is essential for maintaining BA homeostasis. Intriguingly, our MLL3 mutant mice also displayed favorable metabolic profiles, which we propose is via defects in the ability of ASCOM to antagonize signaling by Tgr5, the plasma membrane receptor for BAs. Tgr5 triggers a signaling pathway that leads to upregulation of 'the cyclic-AMP-dependent thyroid hormone activating enzyme type 2 iodothyronine deiodinase' (D2) and to enhance secretion of glucagon-like peptide-1 (GLP-1), an insulin secretagogue, thereby resulting in enhanced energy expenditure and improved glucose homeostasis. Our preliminary results suggest that ASCOM inhibits Tgr5 signaling not only through decreasing BA levels but also directly through the regulation of a gene encoding a key modifier of Tgr5 signaling, 'dipeptidyl peptidase-4' (Dpp4), which inactivates GLP-1. Together, these results support the central hypothesis of this study: ASCOM functions as a master coactivator of 'the homeostatic regulation of BA signaling' by controlling the expression of genes in multiple pathways that regulate BA metabolism and signaling. This renewal has two objectives: 1) We will test the mechanisms of action for ASCOM in NR transactivation (primarily for FXR and RAR) by using the discoveries made during the previous funding period. 2) By focusing specifically on the role of ASCOM in BA homeostasis, we will establish a new paradigm for understanding the diverse metabolic roles of ASCOM. This is a well-integrated study, as the first part of the study is critical to understand the molecular basis for the functio of ASCOM in BA homeostasis and signaling (the second part of the study). We will tackle these two issues in three specific aims, utilizing a combination of biochemical, cellular and genetic approaches. This study will help us to understand the molecular basis for how NRs regulate transcription and metabolism.