COLLABORATIVE RESEARCH: Development of the Triad Junction inSkeletal Muscle

三联结是一种复杂的结构，涉及血浆和肌浆网膜以及至少7种蛋白质，是电兴奋被转导释放细胞内钙的部位，因此在电兴奋与肌肉收缩的耦合中起关键作用。此外，它可能与细胞骨架相互作用，形成肌肉特异性形态。在胚胎鸡骨骼肌分化过程中，二氢吡啶受体（DHPR）和红嘌呤受体（RR）在骨骼肌三联体的两个关键组成部分表达较早。该项目的目的是确定这些蛋白如何被纳入三联结并在骨骼肌发育过程中细胞内钙的释放中变得活跃，以及由三联结介导的钙释放事件是否影响肌肉发育的其他方面。具体目的是：(1)通过建立DHPR和RR在发育中的鸡胚骨骼肌中的表达时间，明确实验的具体内容；(2)通过验证DHPR和RR作为三联结组装和整合到成熟肌肉结构的组织者的假设，研究这些蛋白质是否影响骨骼肌的发育。这将通过定义这些和其他连接成分结合到连接中的时间过程来完成，并评估连接蛋白和细胞骨架元件之间相互作用的重要性。埃利斯曼博士对该项目的贡献将主要集中在形态学方面。骨骼肌为移动骨骼和其他结构提供收缩力，影响器官或整个生物体的运动。“告诉”肌肉收缩并确保沿着肌肉长度同步收缩的信号涉及神经肌肉连接处产生的电波，该电波沿着质膜沿着肌肉纤维的长度传播。电信号向收缩的转化是通过电信号向“第二信使”信号的转导发生的，即钙从细胞内储存（肌浆网）释放到细胞质中。在肌细胞的“三联结”处，肌浆网膜与称为横小管的质膜的特殊结构紧密相连。这个高度特化的区域包含一组独特的肌浆网膜蛋白，这些蛋白与紧密相连的横小管膜相互作用，影响电信号与钙通道打开的耦合，钙通道允许钙从肌浆网释放到细胞质。无论是信号转导事件的机制，还是这种复杂的亚细胞结构的细胞发育，都没有得到很好的理解。这项研究的结果将增加我们对脊椎动物肌肉中这两个非常重要的过程的理解。

The triad junction is a complex structure involving plasma and sarcoplasmic reticulum membranes and at least seven proteins, and is the site where electrical excitation is transduced to release intracellular calcium, and thus has a critical role in coupling electrical excitation to muscle contraction. Additionally, it may interact with the cellular cytoskeleton to form muscle-specific morphology. Two key components of the triad junction, the dihydropyridine (DHPR) and ryanodine receptors (RR), are expressed early during embryonic chick skeletal muscle differentiation. The aim of this project is to determine how these proteins are incorporated into the triad junction and become active in the release of intracellular calcium in developing skeletal muscle, and whether the calcium release events mediated by the triad junction influence other aspects of muscle development. The specific aims are to: (1), define the experimental in detail by establishing the time course of the expression of the DHPR and RR in developing embryonic chick skeletal muscle; and (2), investigate whether these proteins influence skeletal muscle development by testing the hypothesis that DHPR and RR serve as organizers for the assembly of the triad junction and its integration into mature muscle structure. This will be done by defining the time course of incorporation of these and other junctional components into the junction, and assess the significance of interactions between junctional proteins and cytoskeletal elements. Dr. Ellisman's contribution to the project will be primarily with regard to the morphological aspects. Skeletal muscle provides the contractile force to move bones and other structures, effecting organ or whole organism movement. The signal which "tells" the muscle to contract and ensures synchronous contraction along the length of the muscle involves an electrical wave generated at the neuromuscular junction, which travels down the length of the muscle fiber along the plasma membrane. The conversion of the electrical signal to contraction occurs by transduction of the electrical signal to a "second messenger" signal, namely release of calcium from intracellular stores (sarcoplasmic reticulum) into the cytoplasm. At the "triad junction" of the muscle cell, the membrane of the sarcoplasmic reticulum is closely apposed to a specialized structure of the plasma membrane known as the transverse tubule. This highly specialized region contains a set of unique sarcoplasmic reticulum membrane proteins which interact with the closely apposed transverse tubule membrane to effect the coupling of the electrical signal to the opening of a calcium channel which allows the release of calcium from the sarcoplasmic reticulum to the cytoplasm. Neither the mechanism of this signal transduction event, nor the cellular development of this intricate subcellular structure, is well understood. The results of this research will increase our understanding of both of these very important processes in vertebrate muscle.