Regulation of Store-Operated Calcium Entry During Muscle Aging

项目摘要/摘要 骨质疏松症是指骨骼肌群在衰老过程中逐渐丧失，是导致虚弱的主要因素。 使人衰弱的伤害，丧失独立性，降低老年人的生活质量。不幸的是，骨质疏松症的进展 尽管采取了诸如增加体力活动和改善饮食等干预措施。在石棺减少期间，与年龄相关的下降 肌肉力量的下降是由于肌肉质量丧失(萎缩)和肌肉比力降低(即， 单位横截面积的肌力)。越来越多的人认为，伴随而来的是肌肉无力 骨骼减少，而不是肌肉大小本身的减少，是导致残疾的主要原因。我们最近发现， 骨骼肌收缩力产生的要素依赖于存储操作的钙内流(SOCE)，并且 这种能力在老化的骨骼肌中消失。这种改变的SOCE直接与减少的 Mitsugumin 29(MG29)是突触素(SYPL2)家族中的一种肌肉特异性蛋白，它包含一种 Marvel结构域参与胆固醇结合、脂质处理和这些蛋白质低聚体的形成。MG29是 对于横管(TT)系统的正确形成和有效的SOCE来说是必不可少的。这导致了两者之间的联系 MG29在老年肌肉中的表达减少，并在骨质疏松症中发生肌肉功能障碍。骨骼肌 来自年轻的mg29-/-小鼠的与来自老年野生型(WT)小鼠的相似之处在于它们表现出的特异性降低 肌收缩力、SOCE降低，SR钙释放减少，MG29含量减少，TT形态改变。 因此，我们的数据表明，在mg29-/-小鼠中存在骨骼肌衰老的关键方面，从而形成了这个模型。 适用于这一领域的研究。在这个提案中，我们特别假设MG29是正确的TT所必需的 形成以及SOCE在骨骼肌中通过特定蛋白质结构域的作用发挥作用。减少MG29 正常衰老肌肉中的含量会导致SOCE缺陷，从而导致钙的利用率降低 肌肉萎缩不能解释肌肉的收缩能力和与年龄相关的肌肉力量丧失。我们将检验这一假设 通过两个具体的目标：a)检查MG29在骨骼肌中作用的机制基础，b)建立 MG29如何适应老龄化和E-C耦合的多模式方面的更广泛背景，以及c)提供 对治疗骨质疏松症的翻译价值。目标1将阐明MG29对SOCE、SR-Ca的贡献 电穿孔MG29基因或RNAi结构改变老年骨骼肌的释放和收缩 MG29的表达及其与SOCE功能、肌浆网钙释放、收缩力和基因表达的关系 级别。在目标2中，我们使用了一种新的小鼠模型，结合分子生物学和生化方法来解决 负责MG29作用机制的蛋白质基序在脂质调节中的分子机制的建立 MG29对骨骼肌SOCE调节的研究以及这些机制在衰老过程中如何改变。

Project Summary/Abstract Sarcopenia is the progressive loss of skeletal muscle mass during aging and is a leading factor contributing to frailty, debilitating injuries, loss of independence, and reduced quality of life in the elderly. Unfortunately, sarcopenia progresses despite interventions such as increased physical activity and improved diet. During sarcopenia, the age-related decreases in muscle strength result from a combination of loss of muscle mass (atrophy) and reduced muscle specific force (i.e., muscle force per unit of cross-sectional area). Increasingly, it is thought that it is the muscle weakness that accompanies sarcopenia, rather than the loss of muscle size per se, as the principal contributor to disability. We recently found that elements of contractile force generation in skeletal muscle are dependent on store-operated Ca2+ entry (SOCE) and that this capacity is lost in aging skeletal muscle. This altered SOCE is directly associated with a reduced content of mitsugumin 29 (MG29), a muscle-specific protein belonging to the synaptophysin (SYPL2) family which contains a MARVEL domain involved in cholesterol binding, lipid handling and formation of oligomers of these proteins. MG29 is essential for the proper formation of the transverse-tubule (TT) system and efficient SOCE. This leads to a link between reduced MG29 expression in aged muscle and the development of muscle dysfunction in sarcopenia. Skeletal muscles from young mg29-/- mice are similar to those from aged wild-type (WT) mice in that they demonstrate decreased specific contractile force, reduced SOCE, reduced Ca release from the SR, reduced content of MG29, and altered TT morphology. Therefore, our data demonstrate that key aspects of skeletal muscle aging are present in mg29-/- mice, making this model applicable for this line of research. In this proposal we specifically hypothesize that MG29 is required for proper TT formation and that SOCE functions in skeletal muscle through the action of specific protein domains. Decreased MG29 content in normal aged muscle leads to defective SOCE, which results in the decreased availability of Ca2+ for contractility and age-related loss of muscle strength not accounted for by muscle atrophy. We will test this hypothesis through two specific aims that will: a) examine the mechanistic basis for MG29 function in skeletal muscle, b) establish how MG29 fits into the broader context of the multi-modal aspects of aging and E-C coupling, and c) provide translational value for the treatment of sarcopenia. Aim 1 will elucidate the contribution of MG29 to SOCE, SR-Ca release and contractility in aged skeletal muscle by using electroporation of MG29 cDNA or RNAi constructs to alter the expression of MG29 and concomitantly evaluate SOCE function, SR-Ca release, contractile force and gene expression levels. In Aim 2 we use a novel mouse model supported by molecular biology and biochemical approaches to resolve the protein motifs responsible for the mechanism of MG29 function in lipid handling to establish the molecular mechanisms of SOCE regulation by MG29 in skeletal muscle and how these mechanisms are altered in aging.