Signaling Mechanisms of the Dystrophin-Glycoprotein Complex

描述(申请人提供)：Duchenne肌营养不良症(DMD)是儿童中最常见的致命性遗传病，是一种由dystrophin基因突变引起的严重肌肉萎缩疾病。肌营养不良蛋白是肌营养不良蛋白-糖蛋白复合体(DGC)的核心成分，它被认为通过在内部肌动蛋白细胞骨架和细胞外基质之间提供强大的、灵活的连接来赋予肌肉纤维机械稳定性。许多研究都研究了dystrophin的功能和DGC的核心成分，然而，对DGC中与dystrophin的C末端部分结合的其他成员的功能知之甚少。其中一些蛋白质，包括合成素和神经元型一氧化氮合酶(NNOS)，具有已知的信号特性，并可能在肌肉组织中发挥重要的信号作用。我们之前建立了骨骼肌表达DPI 16的转基因小鼠，并将此转基因引入Dstrophin缺陷型MDX小鼠，作为DMD的模型。DPI 16是dystrophin的非肌肉亚型，缺乏肌动蛋白结合结构域和全长dystrophin亚型中的大部分杆状结构域。推测DPI 16与肌动蛋白细胞骨架没有直接的机械联系，但仍能在肌膜上组装和稳定DGC。因此，DPI 16应该允许研究dystrophin的C-末端部分及其结合伙伴的功能和结构，而不依赖于dystrophin N-末端和杆状结构域的机械功能。我们已经证明，DPI 16对mdx小鼠肌肉的影响很小，在mdx小鼠的肌肉中，utroin上调以部分补偿dystrophin的缺乏。然而，当在严重营养不良的dystrophin：utroin双基因敲除(MDX：UTUN-/-)小鼠中表达。DPI 16的表达显著延长了患者的寿命，延缓了关节痉挛和后凸的形成和发展。因此，我们建议研究Dp116-Mediate(J)DGC组装导致mdx：uTUN-/-小鼠表型拯救的结构和功能机制，并探索DGC在骨骼肌中可能的信号转导作用。我们将生成删除构建体来识别DPI 16的结构域，这些结构域对于拯救致命性至关重要。我们还将评估骨骼肌生物学的各个方面，包括再生、卫星细胞激活、细胞外基质组织和线粒体功能可能受到DGC的影响。

DESCRIPTION (provided by applicant): Duchenne muscular dystrophy (DMD), the most common lethal genetic disorder in children, is a severe muscle-wasting disease caused by mutations in the dystrophin gene. Dystrophin is a core component of the dystrophin-glycoprotein complex (DGC), which is thought to confer mechanical stability to the muscle fiber by providing a strong, flexible connection between the internal actin cytoskeleton and the extracellular matrix. Numerous studies have examined the function of dystrophin and the core components of the DGC, however, less is known about the function of other members of the DGC that bind the C-terminal portion of dystrophin. Some of these proteins, including syntrophin and neuronal nitric oxide synthase (nNOS), have documented signaling properties and may play important signaling roles in muscle tissue. We previously generated transgenic mice with skeletal muscle expression of Dpi 16 and have introduced this transgene into dystrophin-deficient mdx mice, which serve as a model for DMD. Dpi 16 is a non-muscle isoform of dystrophin that lacks the actin-binding domain and the majority of the rod domain found in the full length dystrophin isoform. It is postulated that Dpi 16 has no direct, mechanical link to the actin cytoskeleton, but can still assemble and stabilize the DGC at the sarcolemma. Thus, Dpi 16 should permit functional and structural studies of both the C-terminal portion of dystrophin and its binding partners, independent of the mechanical function of the dystrophin N-terminus and rod domain. We have shown that Dpi 16 has minimal effect in muscles of mdx mice, in which utrophin is upregulated to partially compensate for the lack of dystrophin. However, when expressed in the severely dystrophic dystrophin:utrophin double knockout (mdx:utrn-/-) mouse. Dpi 16 expression imparts a dramatic increase in longevity and a delay in the formation and progression of joint contractures and kyphosis. Thus, we propose to study the structural and functional mechanism through which Dp116-mediate(j DGC assembly leads to phenotypic rescue in mdx:utrn-/- mice and explore possible signaling roles of the DGC in skeletal muscle. We will generate deletion constructs to identify the domains of Dpi 16 that are critical for rescue of lethality. We will also evaluate aspects of skeletal muscle biology, including regeneration, satellite cell activation, extracellular matrix organization, and mitochondrial function that may be influenced by the DGC.