Genetic Control of Phospholipid Biosynthesis and Muscular Dystrophy

描述(申请人提供)：肌营养不良症包括一组不同的遗传异质性疾病，影响全球1：2000出生。这些疾病的特点是进行性肌肉无力和消瘦，导致严重残疾，通常还会过早死亡。轮尾型肌营养不良症(RMD)是一种新的小鼠隐性突变，导致进行性肌营养不良症，其严重程度为从吻端到尾端的梯度。我们已经确定RMD突变是胆碱激酶β(Chkb)基因内1.6kb的缺失，导致CHKB蛋白和酶活性的丧失。CHKB是哺乳动物胆碱激酶(ChK)的两种酶之一，在主要的膜磷脂磷脂酰胆碱(PC)的生物合成过程中催化胆碱磷酸化为磷胆碱。PC是真核细胞中含量最丰富的磷脂，它不仅是膜双层的结构基础，也是多种脂质信使的前体。虽然突变的RMD小鼠在所有被检查的组织中都表现出显著的CHK活性降低，但这种疾病只在骨骼肌组织中明显存在，这些组织的总PC含量减少了30%。已经在肌膜或核膜上起作用的膜相关蛋白中发现了几个肌肉营养不良突变。然而，RMD突变小鼠是第一个证明膜磷脂组成的变化可以导致进行性肌营养不良表型的。我们建议的目的是：1)验证RMD肌营养不良症是由于骨骼肌中CHKB的细胞自主丢失所致的假设。2)验证线粒体功能障碍是小鼠成肌细胞原代培养和成人骨骼肌进行性退变的基础假说。3)对巴西大量先天性肌营养不良症(CMD)和四肢带状肌营养不良症(LGMD)患者进行CHKB基因突变筛查。与Mayana Zatz博士合作，我们将检测来自患者的DNA样本中的人类CHKB突变，这些患者之前已被筛查并消除了肌肉营养不良的已知分子原因。这项建议的主要目标是使用新的小鼠RMD突变来探索骨骼肌中这种缺陷的细胞自主性质，剖析肌肉退化的分子和遗传机制，并确定CHKB缺陷的范围，这些缺陷是以前未知的人类肌肉营养不良的基础。RMD小鼠首次展示了导致肌肉营养不良的磷脂生物合成酶缺陷，并可能揭示干预疾病过程的独特切入点。

DESCRIPTION (provided by applicant): Muscular dystrophies include a diverse group of genetically heterogeneous disorders that affect 1:2000 births worldwide. The diseases are characterized by progressive muscle weakness and wasting that lead to severe disability and often premature death. Rostrocaudal muscular dystrophy (rmd) is a new recessive mouse mutation causing a progressive muscular dystrophy with a rostral-to-caudal gradient of severity. We have identified the rmd mutation as a 1.6 kb deletion within the choline kinase beta (Chkb) gene, resulting in loss of CHKB protein and enzymatic activity. CHKB is one of two mammalian choline kinase (CHK) enzymes catalyzing the phosphorylation of choline to phosphocholine in the biosynthesis of the major membrane phospholipid phosphatidylcholine (PC). PC is the most abundant phospholipid in eukaryotic cells and it serves not only as a structural foundation for membrane bilayers, but also as a precursor for several lipid messengers. While mutant rmd mice show dramatically decreased CHK activity in all tissues examined, the disease is only evident in skeletal muscle tissues which display a 30% reduction in total PC content. Several muscular dystrophy mutations have been identified in membrane-associated proteins that function at the sarcolemma or nuclear envelope. However, the rmd mutant mouse is the first to demonstrate that alterations in membrane phospholipid composition can result in a progressive muscular dystrophy phenotype. The aims of our proposal are: 1) To test the hypothesis that rmd muscular dystrophy is due to a cell autonomous loss of CHKB in skeletal muscles. 2) To test the hypothesis that mitochondrial dysfunction underlies the progressive skeletal muscle degeneration in primary mouse myoblast cultures and adult skeletal muscles. 3) To screen a large Brazilian population of congenital muscular dystrophy (CMD) and limb-girdle muscular dystrophy (LGMD) patients for mutations in the orthologous human CHKB gene. In collaboration with Dr. Mayana Zatz, we will test for human CHKB mutations in DNA samples from patients that have previously been screened and eliminated for known molecular causes of muscular dystrophy. The primary goals of this proposal are to use the novel mouse rmd mutation to explore the cell-autonomous nature of this defect in skeletal muscles, to dissect the molecular and genetic mechanisms of muscular degeneration and to determine the range of defects in CHKB that underlie previously unidentified human muscular dystrophies. The rmd mouse offers the first demonstration of a defect in a phospholipid biosynthetic enzyme causing muscular dystrophy and may reveal unique entry points in which to intervene in the disease process.