Defining the role and essential interactions of the centrosomal protein pericentrin in microtubule formation during myogenesis

肌纤维是由成肌细胞在称为肌发生的过程中融合形成的长的多核细胞。细胞核沿着肌纤维的周边规则排列。正确的核定位涉及核膜蛋白、中心体蛋白和马达蛋白之间的相互作用。异常定位，如核簇与肌营养不良症，表明正确的核定位是重要的健康功能的骨骼肌的形成。然而，参与核定位的分子机制和蛋白质的确切作用的细节尚不清楚。在成肌细胞中，微管组织中心（MTOC）的蛋白质位于中心体。已经表明，分化后仅数小时，MTOC的标记物从中心体释放，而是定位在核周表面。这些蛋白质保留其作为微管成核位点的作用。我们的假设是，中心体蛋白定位到核周表面结合nesprin-1，核包膜蛋白，是LINC复合物的一部分，物理连接细胞核的细胞骨架成分。LINC复合物组分中的突变可引起核簇集的事实支持了这一点。一种解释可能是因为MTOC蛋白向核膜的募集和随后的微管重组受损。由于马达蛋白使用微管来移动货物，因此核向细胞周边的移动受到影响。这个项目将测试我们的假设，导致一个全面的了解中心体蛋白质的作用，在微管成核肌发生过程中。我们将确定参与微管成核和核定位的特定蛋白质的功能，并使用一系列细胞生物学和结构技术来定义蛋白质相互作用。这将建立在我们目前的肌肉发生的分子模型。

Muscle fibers are long multinucleated cells formed from the fusion of myoblasts in a process called myogenesis. Nuclei are arranged regularly along the periphery of the myofiber. Correct nuclear positioning involves interactions between nuclear envelope proteins, centrosomal proteins and motor proteins. Aberrant positioning such as nuclear clustering is associated with muscular dystrophies, suggesting that correct nuclear postioning is important for the formation of healthy functioning skeletal muscle. Yet, the details of the molecular mechanisms and the precise roles of proteins involved in nuclear positioning are unclear. In myoblasts, proteins of the microtubule organising centre (MTOC) are localised at the centrosome. It has been shown that just hours after differentiation, markers of the MTOC are released from the centrosome and instead localise at the perinuclear surface. These proteins retain their role as a microtubule nucleation site. Our hypothesis is that centrosomal proteins localise to the perinuclear surface by binding to nesprin-1, a nuclear envelope protein that is part of the LINC complex that physically links the nucleus to cytoskeletal components. This is supported by the fact that mutations in LINC complex components can cause nuclear clustering. An explanation could be because recruitment of MTOC proteins to the nuclear envelope and subsequent microtubule reorganisation becomes impaired. Since motor proteins use microtubules to move cargo, nuclear movement to the cell periphery becomes affected. This project will test our hypothesis, leading to a comprehensive understanding of the roles of centrosomal proteins in microtubule nucleation during myogenesis. We will identify the function of specific protein involved in microtubule nucleation and nuclear positioning, and define the protein interactions using a range of cell biology and structural techniques. This will build upon our current molecular model of myogenesis.