SARCOGLYCAN IN MYOPATHY AND MUSCLE MEMBRANE STABILITY

肌营养不良蛋白的缺陷被认为破坏了肌肉细胞的细胞骨架、膜和细胞外基质之间的机械联系，导致肌肉营养不良和心肌病。肌营养不良蛋白结合了一种蛋白质复合体，该复合体将其附着在肌肉膜上，进而结合细胞外基质蛋白-层粘连蛋白。层粘连蛋白突变也会导致肌病，但层粘连蛋白缺陷肌肉中的膜似乎不像dystrophin缺陷膜那样受到物理上的破坏，这意味着一个独特的分子机制。在肌营养不良蛋白-糖蛋白复合体中有一种多亚单位蛋白，即肌聚糖，当肌营养不良蛋白发生突变时，它会继而减少。最近，人们发现肌聚糖基因的突变是人类肌病的主要原因。肌聚糖的功能尚不清楚，其预测的结构表明它是细胞表面的受体。为了研究缺乏肌聚糖的肌肉的膜缺陷，我们使用胚胎干细胞中的同源重组来产生缺乏不同亚基的肌聚糖的小鼠。γ-肌聚糖和β-肌聚糖是一类相关的35 kDa糖基化跨膜蛋白，通常在心肌和骨骼肌中表达。我们产生了缺乏γ-肌聚糖的小鼠和杂合性缺乏β-肌聚糖的胚胎干细胞。初步数据显示，缺乏γ-肌糖的小鼠表现出一种严重的营养不良模式，类似于由肌糖突变引起的人类肌病。将使用各种免疫细胞化学方法来鉴定肌聚糖缺陷小鼠的特征。将调查由于肌聚糖缺乏而导致的基因表达的变化。最后，我们将进行旨在破坏多个膜-细胞外基质连接的遗传学实验，以评估这些不同连接的相对贡献及其在正常和异常心脏和肌肉功能中的作用。

Defects in dystrophin are thought to disrupt a mechanical link between the cytoskeleton, the membrane and the extracellular matrix of muscle cells resulting in muscular dystrophy and cardiomyopathy. Dystrophin binds a complex of proteins that anchor its attachment to the muscle membrane and in turn, bind the extracellular matrix protein, laminin. Laminin mutations also result in myopathy, but the membrane in laminin-deficient muscle does not seem to be as physically disrupted as dystrophin-deficient membranes implying a distinct molecular mechanism. Within the dystrophin-glycoprotein complex is a multisubunit protein, sarcoglycan, that is secondarily decreased when dystrophin is mutated. Recently, it was discovered that mutations in sarcoglycan genes are a primary cause of human myopathy. The function of sarcoglycan is unknown and its predicted structure suggests a cell surface receptor. To investigate membrane defects in muscle lacking sarcoglycan, we are using homologous recombination in embryonic stem cells to generate mice lacking different subunits of sarcoglycan. gamma-sarcoglycan and delta-sarcoglycan are related 35 kDa glycosylated, transmembrane proteins normally expressed in cardiac and skeletal muscle. We have generated mice lacking gamma-sarcoglycan and embryonic stem cells heterozygously lacking delta-sarcoglycan. Preliminary data indicate mice lacking gamma-sarcoglycan show a severe dystrophic pattern that resembles human myopathy arising from sarcoglycan mutations. Sarcoglycan-deficient mice will be characterized using a variety of immunocytochemical approaches. Changes in gene expression that result from sarcoglycan deficiency will be investigated. Lastly, we will perform genetic experiments aimed at disrupting multiple membrane-extracellular matrix connections to assess the relative contribution of these different attachments and their role in normal and abnormal heart and muscle function.