Core D

核心D的目标是为项目1、2、3和4提供电子显微镜超微结构支持 我们预计，对钙释放单位(CRU)内蛋白质处置的结构分析以及对 影响完整CRU组装的因素将显著提高我们对其功能的理解 以及RyR1中MH/Ccd突变引起的功能障碍。 A1.检测MH/CCDRyR1突变对骨骼超微结构的影响 肌肉。我们将确定任何由我的MH/CD突变引起的超微结构异常，特别是 关注RyR-DHPR三元关系和钙调素(支持项目1、2、3、4和# 与核心B和C的协作)。 A2.检测RyR构象变化和DHPR/RyR结构变化的证据 评估MH/CD引起的DHPR四分体参数变化的构象耦合 RyR1基因突变。我们将确定小鼠和人类MH/CCDRYR对 协调四分体的形成，并确定这些突变是否改变了四分体内或四分体间的间距。(支持 1和2与核心B和C协作)。 A3.明确线粒体和SR在发育/衰老过程中的结构关系 并确定wt和mh/ccd小鼠之间的这种关系是否存在差异。我们的 初步数据表明，线粒体逐渐获得与三联体相关的非常特定的位置 在出生后发育过程中。此外，我们将确定SR和 纯合子和杂合子MH小鼠的线粒体。(支持项目1、3、4)。 A4.从对照和纯合MH/CCD肌分离成熟树突状细胞中检测CRU 并定义它们的结构。我们将定义RyR1和CaV1.3所在位置的超微结构 共定位于树突状细胞(DC)，并检测MH/CD突变引起的任何变异 (支持项目2)

The goals of Core D are to provide electron microscopic ultrastructure support to Projects 1, 2, 3 and 4 We expect that a structural analysis of protein disposition within the calcium release units (CRUs) and of the factors that affect assembly of complete CRUs will significantly enhance to our understanding of their function and the dysfunction caused by MH/CCD mutations in RyR1. A1. To determine the effect of MH/CCD RyR1 mutations on the ultrastructure of affected skeletal muscle. We will define any ultrastructural abnormalities caused my MH/CCD mutations, paying particular attention to triadic RyR-DHPR relationships and calsequestrin (Supporting Projects 1, 2, 3, 4 and in collaboration with Cores B and C). A2. To detect evidence for RyR conformational changes and for structural changes in DHPR/RyR conformational coupling by assessing alterations in DHPR tetrad parameters resulting from MH/CCD mutations in RyR1. We will determine the relative effectiveness of mouse and human MH/CCD RyRs to coordinate tetrad formation and to determine if these mutations alter intra or inter tetrad spacing. (Supporting 1, and 2 in collaboration with Cores B and C). A3. To define the structural relationship between mitochondria and the SR during development/aging and determine if there are differences in this relationship between wt and MH/CCD mice. Our preliminary data indicate that mitochondria gradually acquire very specific locations relative to the triads during postnatal development. In addition we will determine the structural integrity of both the SR and the mitochondria in homozygous and heterozygous MH mice. (Supporting Projects 1, 3, and 4). A4. To detect CRUs in mature dendritic cells isolated from control and homozygous MH/CCD muscle and to define their structure. We will define the ultrastructure of sites where RyR1 and CaV1.3 are co-localized in dendritic cells (DC) and detect any variations introduced by the MH/CCD mutations (Supporting project 2)