Genetic Control of Phospholipid Biosynthesis and Muscular Dystrophy

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DESCRIPTION (provided by applicant): The long-term goals of this project are to determine how the dysregulation of organelle membrane lipid biosynthesis and maintenance leads to a congenital muscular dystrophy (CMD), how we can successfully design strategies to treat this disorder, and how this disease mechanism can inform upon other types of muscular dystrophy. We first identified PC homeostasis as critical for skeletal muscle maintenance in the rostrocaudal muscular dystrophy (rmd) mutant mouse, and later in human CMD patients with loss of function mutations in the choline kinase beta (CHKB) gene. CHKB is one of two mammalian enzymes catalyzing the phosphorylation of choline to phosphocholine in the Kennedy pathway. Loss of CHKB activity results in significantly reduced skeletal muscle PC levels and a progressive muscular dystrophy phenotype with nuclear membrane dysmorphology and distinctly enlarged mitochondria (megamitochondria) with reduced respiratory function. We hypothesize that alterations in membrane PC content directly affect the functional properties of skeletal muscle organelles (nuclei and mitochondria) and that a strategy to restore membrane PC levels will be therapeutically beneficial. In aim 1, we will define the mechanisms regulating mitochondrial and nuclear dysfunction. We propose to a) determine if PC deficiency disrupts mitochondrial fission at points of ER/mitochondrial contact, b) define mechanisms regulating mitochondrial fission/fusion using high-resolution FPALM microscopy to test the real-time dynamics of mitochondrial membrane curvature changes, and c) determine if nuclear membrane changes are functionally related to those seen in LMNA Emery-Dreifuss MD. In aim 2, we will test therapeutic strategies by a) determining if CHK-alpha (CHKA) can substitute for CHKB deficiency using a transgenic approach, b) testing if overexpression of mitochondrial fission proteins, or knockout of mitochondrial fusion proteins can alleviate the megamitochondrial disease phenotype, and c) testing if PC or an intermediate metabolite can be administered therapeutically to restore phospholipid homeostasis. PUBLIC HEALTH RELEVANCE: Alterations in cell and organelle membrane morphology and function have been shown to be key to a variety of human diseases, including muscular, neurological, and metabolic disorders. Our discovery of choline kinase beta (Chkb) mutations in the rmd mouse, and in a recently identified human congenital muscular dystrophy with null mutations in the CHKB gene, are the first indications that altered phospholipid synthesis can result in mitochondrial and nuclear membrane defects in muscle disease. The development of our proposed mitochondrial photoactivatable myoblast cell line, and our use of high-resolution FPALM microscopy to study the impact of membrane composition and architecture and muscular disease, will be a vital component for further understanding of the role of membrane phospholipids in a wide range of human illnesses.