MLL3/4-complexes in nuclear receptor-mediated metabolism

描述（由申请人提供）：胆汁酸（BA）信号的稳态调节需要一个复杂的核受体（nr）网络，但它们的共激活因子重塑染色质并调节基因转录以响应BA信号，目前对它们的共激活因子知之甚少。我们发现了NR共激活剂ASC-2，随后纯化了“ASC-2复合物”（ASCOM），这是第一个含有H3赖氨酸4甲基转移酶（H3K4MT） MLL3或其同源物MLL4的哺乳动物复合物。后来，ASCOM也被发现含有h3 -赖氨酸27-去甲基化酶（H3K27DM） 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包含两种与转录激活相关的酶。令人兴奋的是，我们发现ASCOM的主要生理功能是调节多种不同条件下的代谢，这主要归因于ASC-2招募ASCOM到多种代谢NRs的能力，包括PPAR？lxr和FXR是BAs的NR。特别是，我们发现ASCOM在调节BA合成中作为FXR的关键辅激活剂起作用。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 H3K4MT活性对于维持BA稳态至关重要。有趣的是，我们的MLL3突变小鼠也表现出良好的代谢特征，我们认为这是通过ASCOM拮抗BAs质膜受体Tgr5信号传导的能力缺陷。Tgr5触发一个信号通路，导致“循环amp依赖性甲状腺激素激活酶2型碘甲状腺原氨酸去碘酶”（D2）的上调，并增强胰高血糖素样肽-1 （GLP-1）的分泌，胰高血糖素样肽-1是一种胰岛素分泌剂，从而导致能量消耗增加，改善葡萄糖稳态。我们的初步结果表明，ASCOM不仅通过降低BA水平来抑制Tgr5信号传导，还直接通过调控编码Tgr5信号传导关键修饰因子“二肽基肽酶-4”（Dpp4）的基因来抑制GLP-1。总之，这些结果支持了本研究的中心假设：ASCOM通过控制多种途径中调节BA代谢和信号传导的基因表达，作为“BA信号稳态调节”的主辅激活因子。这次更新有两个目标：1)我们将利用之前资助期间的发现，测试ASCOM在NR交易中的作用机制（主要是FXR和RAR）。2)通过特别关注ASCOM在BA稳态中的作用，我们将建立一个新的范式来理解ASCOM的多种代谢作用。这是一项整合良好的研究，因为研究的第一部分对于了解ASCOM在BA稳态和信号传导中的功能的分子基础至关重要（研究的第二部分）。我们将在三个具体目标中解决这两个问题，利用生物化学，细胞和遗传方法的组合。这项研究将有助于我们了解rna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.