Sub-Project #3

骨骼肌钙释放通道，也被称为ryanodine受体(RYR1)，调节 肌浆网(SR)释放的钙离子在人体中央核储存并发生突变 疾病(CCD)和药物遗传综合征中的恶性高热(MH)。这个 突变集中在RYR1的三个区域：区域1从氨基酸35到614，区域2从 2163到2459和区域3从4647到4914。MH突变主要位于细胞质 区域1和区域2，而大部分的Cd突变发生在跨膜区3。 本项目中要测试的工作假设是：a.位于夹具中的区域1和区域2 结构域相互作用，以变构方式稳定通道的关闭状态，b.MH突变，通过 破坏这种相互作用，增强通道(从而肌肉)对激活剂的反应 温度升高，c.运动中RyR1活性增强会提高内源性 作为热原的细胞因子(如IL-6)和随后的体温升高 增加MH反应的概率。为了检验这些假设，有以下具体目标 建议：A1.定义MH/CD突变对构象变化的影响 RyR1A2的钳制结构域。评价MH/CD基因突变对细胞结构和功能的影响 肌肉的收缩特性。A3.确定运动性MH的概率 反应和/或横纹肌溶解会因体温升高和/或增加而增加 内源性IL-6浓度。这些研究将使用具有MH/CD突变的小鼠 已在项目1和3中生成。

The skeletal muscle Ca2+ release channel, also known as the ryanodine receptor (RYR1), regulates the release of Ca2+ from sarcoplasmic reticulum (SR) stores and is mutated in human central core disease (CCD) and in the pharmacogenetic syndrome, malignant hyperthermia (MH). The mutations cluster in three regions of RYR1: region 1 from amino acids 35 to 614, region 2 from 2163 to 2459 and region 3 from 4647-4914. The MH mutations are located primarily in cytoplasmic regionsl and 2 while most of the CCD mutations are found in the transmembrane region 3. The working hypotheses to be tested in this project are: a. Regions 1 and 2, located in the clamp domain, interact to allosterically stabilize a closed state of the channel, b. The MH mutations, by destabilizing this interaction, enhance the response of the channel (and hence muscle) to activators and elevated temperature, c. Enhanced RyR1 activity during exercise elevates endogenous cytokines that act as pyrogens (such as IL-6) and the subsequent increase in body temperature increases the probability of an MH response. To test these hypotheses, the following specific aims are proposed: A1. Define the effects of the MH/CCD mutations on the conformational changes in the clamp domain of RyR1 A2. Evaluate the effects of the MH/CCD mutations on the structural and contractile properties of the muscle. A3. Determine if the probability of exercise-induced MH responses and/or rhabdomyolysis is increased by elevated body temperature and/or increased endogenous IL-6 concentrations. These studies will use mice with MH/CCD mutations that have been generated in Projects 1 and 3.