Understanding the role of IFN-gamma and CIITA in skeletal muscle

 描述（由申请人提供）：本研究的目的是确定炎症细胞因子γ干扰素（IFN-γ）如何调节肌肉功能和修复，γ干扰素（IFN-γ）在响应创伤或疾病引起的肌肉损伤时上调。骨骼肌的修复对人类健康至关重要，修复效率低下会导致运动能力丧失，最终导致死亡。炎症反应在骨骼肌对损伤的反应和启动骨骼肌修复中起着重要作用。IFN-γ在肌肉发生中起积极和消极作用，并且是体内有效肌肉再生所需的。我们已经发现IFN-γ通过II类反式激活因子CIITA通过抑制肌纤维形成所需的生肌蛋白（Myogenic Regulatory Factor，MRF）的表达或活性而起作用。在我们理解CIITA抑制机制的研究中，我们发现IFN-γ通过CIITA起维持多梳复合物PRC 2表达的作用，PRC 2在分化过程中通常是沉默的. CIITA将PRC 2复合物募集到受抑制的基因启动子，PRC 2通过催化组蛋白H3赖氨酸27（H3 K27 me 3）的甲基化来抑制基因表达。我们认为，瞬时表达的IFN-γ，通过CIITA，调节肌发生和促进肌肉修复，但建议这种信号的失调有助于肌肉疾病，改变基因表达谱的肌纤维受到IFN-γ刺激。在这个提议中，我们将通过确定在IFN-γ存在下激活因子如MyoD的募集是否被阻断，确定在抑制中使用的表观遗传修饰和确定PRC 2复合物是否是抑制所需的，来确定IFN-γ和CIITA如何抑制肌细胞生成蛋白.接下来，我们将确定已知在分化时改变H3 K27 me 3模式的基因是否响应于IFN-γ而甲基化并依赖于肌细胞生成素。这些数据将与EZH 2的募集和基因表达的抑制相关。最后，我们将在体内证实我们的发现，并确定慢性IFN-γ是否可以抑制肌肉修复。这项工作将利用一种新的小鼠模型，工程表达低慢性水平的IFN-γ。使用该小鼠模型，我们将表征CIITA和PRC 2复合物的表达，确定IFN-γ靶基因是否失调，并表征慢性IFN-γ存在下的修复。为了理解IFN-γ作为炎性浸润的一部分的影响，将在mdx小鼠中测定IFN-γ调节的基因靶的甲基化谱和基因表达，已知mdx小鼠具有增强的IFN-γ表达作为慢性炎症反应的一部分.拟议的工作将加强我们对骨骼肌炎症过程中发生的分子事件的理解。这些信息不仅有助于阐明修复和再生的正常过程，而且还扩展了我们对慢性炎症潜在有害影响的了解，增强了对肌肉萎缩症，恶病质和炎性肌病等疾病的理解，并允许开发针对这些疾病的创新疗法。

 DESCRIPTION (provided by applicant): The objective of this study is to identify how the inflammatory cytokine, interferon gamma (IFN-, which is up regulated in response to muscle injury resulting from trauma or disease, modulates muscle function and repair. The repair of skeletal muscle is essential to human health and inefficient repair leads to a loss of locomotion and eventually, to death. The inflammatory response plays an important role in responding to injury and initiating skeletal muscle repair. IFN- plays both positive and negative roles in myogenesis and is required for efficient muscle regeneration in vivo. We have discovered that IFN- acts through the class II transactivator, CIITA, by repressing the expression or activity o myogenin, the Myogenic Regulatory Factor (MRF) required for myofiber formation. In our studies to understand the mechanism of CIITA repression, we have discovered that IFN-, through CIITA, acts to maintain the expression of the polycomb complex, PRC2, which is normally silenced during differentiation. CIITA recruits the PRC2 complex to repressed gene promoters and PRC2 represses gene expression by catalyzing the methylation of histone H3, lysine 27 (H3K27me3). We propose that transient expression of IFN-, through CIITA, modulates myogenesis and promotes muscle repair, but suggest that deregulation of this signaling contributes to muscle disease by altering the gene expression profile in myofibers subject to IFN- stimulation. In this proposal, we will determine how IFN- and CIITA repress myogenin by determining if recruitment of activating factors such as MyoD are blocked in the presence of IFN-, defining the epigenetic modifications used in the repression and determining whether the PRC2 complex is required for repression. Next, we will determine if genes known to be altered in H3K27me3 patterns upon differentiation are methylated in response to IFN- and dependent on myogenin. These data will be correlated with the recruitment of EZH2 and the repression of gene expression. Lastly, we will confirm our findings in vivo and determine if chronic IFN- can inhibit muscle repair. This work will utilize a new mouse model engineered to express low chronic levels of IFN-. Using this mouse model, we will characterize the expression of CIITA and the PRC2 complex, determine if IFN- target genes are deregulated, and characterize repair in the presence of chronic IFN-. To understand the impact of IFN- as part of the inflammatory infiltrate, the methylation profile and gene expression of IFN- regulated gene targets will be assayed in mdx mice, which are known to have enhanced IFN- expression as part of a chronic inflammation response. The proposed work will strengthen our understanding of the molecular events that occur during inflammation in skeletal muscle. This information will not only help elucidate the normal process of repair and regeneration, but also extend our knowledge about the potential deleterious effects of chronic inflammation, enhancing the understanding of diseases such as muscular dystrophy, cachexia and inflammatory myopathies and allowing the development of innovative therapies for these diseases.