Identification of Mechanisms Regulating Muscle Size and Shape

描述（由申请人提供）：从允许人们跑步的肌肉到指导眨眼的肌肉，骨骼肌有各种形状和大小。这些肌肉具有不同的功能;此外，它们对肌肉萎缩症和横纹肌肉瘤等疾病也具有不同的易感性。由于修复肌肉萎缩的潜在疗法需要产生特定大小和形状的肌肉的能力，因此开发治疗方法的重要一步将是了解肌肉多样性是如何实现的。在果蝇Drosophila melanogaster中，一组被称为身份基因的转录因子标记肌肉的子集，并且对其形态很重要。然而，这些转录因子的下游靶点很少，关于身份基因如何控制肌肉多样性也知之甚少。这个提议的目的是确定形态信息是如何从身份基因翻译到控制肌肉大小和形状的细胞过程的。这项研究的中心假设是，身份基因相互结合，通过调节细胞骨架重塑，细胞粘附和成肌细胞融合等细胞行为来指导肌肉形态发生。果蝇胚胎的侧横肌（LT）的形态发生将使用突变分析，实时成像和基因组方法进行分析，以确定调节这些肌肉发育的因素。这些实验的重点将是确定身份基因如何结合指导LT肌肉形态。五个身份基因已被证明是表达和影响LT肌肉的形态发生：Kruppel，扭曲，无翅，肌肉节段同源框和caupolican。将检查这些基因中的每一个的突变体，以确定每个身份基因对最终肌肉形态的相对贡献，以及确定这五个转录因子之间的遗传层次。形态发生将通过荧光标记物的实时共聚焦成像来表征，阐明该过程中的步骤并帮助解释遗传数据。最后，荧光激活细胞分选（FACS）将用于分离果蝇胚胎成肌细胞的亚群;微阵列分析将在特定的细胞群上进行，以确定发育中的肌肉内的身份基因的目标。我们将进一步研究预测在细胞形状或肌腱附着中发挥作用的基因。总之，这些数据将为身份基因共同作用以指定肌肉的独特大小和形状的机制提供见解。

DESCRIPTION (provided by applicant): From the muscles allowing people to run to those that direct the blink of an eye, skeletal muscles come in a variety of shapes and sizes. These muscles have different functions; additionally, they also have different susceptibilities to diseases like muscular dystrophy and rhabdomyosarcoma. Since potential therapies to repair muscle wasting will require the ability to generate muscles of a specific size and shape, an important step in developing treatments will be to understand how muscle diversity is achieved. In the fruit fly Drosophila melanogaster, a group of transcription factors, known as identity genes, mark subsets of muscles and are important for their morphology. Few downstream targets of these transcription factors are known, however, and little is known about how identity genes control muscle diversity. The goal of this proposal is to determine how morphological information is translated from the identity genes to the cellular processes that control muscle size and shape. The central hypothesis of this research is that identity genes work in combination with each other to direct muscle morphogenesis by regulating cytoskeletal remodeling, cell adhesion and myoblast fusion, among other cellular behaviors. The morphogenesis of the lateral transverse (LT) muscles of the Drosophila embryo will be analyzed using mutant analysis, live imaging, and genomic approaches to identify factors regulating the development of those muscles. The focus of these experiments will be to determine how identity genes work in combination to direct LT muscle morphology. Five identity genes have been shown to be expressed in and affect morphogenesis of the LT muscles: Kruppel, twist, apterous, muscle segment homeobox and caupolican. Mutants for each of these genes will be examined to determine the relative contributions of each identity gene to the final muscle morphology, as well as to determine the genetic hierarchy beteween these five transcription factors. Morphogenesis will be characterized by live confocal imaging of fluorescent markers, elucidating the steps in this process and aiding in the interpretation of genetic data. Finally, fluorescence activated cell sorting (FACS) will be used to isolate subpopulations of myoblasts from Drosophila embryos; microarray analysis will be performed on particular groups of cells to identify the targets of identity genes within the developing muscles. We will focus further studies on those genes predicted to have roles in cell shape or tendon attachment. Taken together, these data will provide insight to the mechanisms by which identity genes act together to specify a muscle's unique size and shape.