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

 描述(申请人提供)：这项研究的目的是确定炎症细胞因子干扰素-(干扰素-γ)是如何调节肌肉功能和修复的，它在应对创伤或疾病造成的肌肉损伤时上调。骨骼肌的修复对人类健康至关重要，效率低下的修复会导致运动能力的丧失，最终导致死亡。炎症反应在骨骼肌损伤反应和启动骨骼肌修复中起着重要作用。干扰素-在肌肉发生中既有积极的作用，也有消极的作用，是体内有效的肌肉再生所必需的。我们已经发现，干扰素-通过II类反式激活因子CIITA，通过抑制肌生成素的表达或活性来发挥作用，肌生成素是肌纤维形成所需的生肌调节因子。在我们对CIITA抑制机制的研究中，我们发现干扰素-通过CIITA维持多梳复合体PrC2的表达，而PrC2在分化过程中通常是沉默的。CIITA招募PRC2复合体来抑制基因启动子，而PRC2通过催化组蛋白H3，赖氨酸27(H3K27me3)的甲基化来抑制基因表达。我们认为，干扰素-的瞬时表达通过CITA调节肌肉发生并促进肌肉修复，但我们认为，该信号的解除调控通过改变干扰素-刺激下肌肉纤维中的基因表达谱而导致肌肉疾病。在这项建议中，我们将通过确定在干扰素-存在的情况下是否阻止诸如MyoD等激活因子的招募，定义用于抑制的表观遗传修饰，以及确定是否需要PrC2复合体来抑制肌生成素，从而确定干扰素-和CIITA是如何抑制肌生成素的。接下来，我们将确定已知的在H3K27me3模式中分化时发生变化的基因是否因干扰素-而发生甲基化，并依赖于肌生成素。这些数据将与EZH2的招募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，并确定慢性干扰素-是否会抑制肌肉修复。这项工作将利用一种新的小鼠模型，该模型被设计为表达低水平的慢性干扰素-。利用这个小鼠模型，我们将表征CITA和PrC2复合体的表达，确定干扰素-靶基因是否被解除调控，并表征在慢性干扰素-存在的情况下的修复。为了了解干扰素-作为炎症渗入的一部分的影响，将在mdx小鼠中检测干扰素-调节基因靶标的甲基化情况和基因表达，众所周知，作为慢性炎症反应的一部分，mdx小鼠会增加干扰素-的表达。这项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，还将扩大我们对慢性炎症潜在有害影响的认识，加强对肌肉营养不良、恶病质和炎症性肌病等疾病的了解，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.