The mechanism by which the loss of dystrophin leads to Duchenne Muscular Dystrophy is generally attributed to membrane fragility due to disruption of the linkage between the extracellular matrix and the cortical actin cytoskeleton. This structural role for dystrophin and its associated proteins is supported by substantial evidence. In addition, the dystrophin complex serves as a sarcolemmal scaffold for signaling proteins, including neuronal nitric oxide synthase (nNOS5, the muscle-specific isoform), several kinases, water and ion channels and other proteins. The demonstration that transgenic expression of nNOS in mdx mouse muscle ameliorates the dystrophic phenotype has focused attention on this signaling protein. Recently, we have found that a second isoform of this enzyme, nNOS2, is localized on the Golgi, and that its expression at this site is reduced in mdx muscle, even before the first phenotypic signs of dystrophy are evident. Furthermore, we find that soluble guanylate cyclase (sGC), which is activated by NO, and cGMP-activated protein kinase G (PKG) are colocalized with nNOS2 on the Golgi. Mice lacking both nNOS5 and nNOS2 (KN2) have numerous deficiencies, including a severe myopathic defect. Based on these preliminary results, we will characterize muscle from KN2 mice at the structural, biochemical and functional levels. In addition, we will test the hypothesis that transgenic expression of nNOS2 in skeletal muscle will rescue/ameliorate the KN2 and mdx phenotypes. The role of nNOS2 in blunting 1-adrenergic vasoconstriction in contracting muscle will provide information on the importance of this enzyme in alleviating functional ischemia, a component of muscular dystrophy. Finally, we will determine the Golgi substrates of the NO/cGMP/PKG pathway and of direct protein nitrosylation. Finally, we will identify the phosphodiesterase(s) (PDEs) that regulate(s) the NO/cGMP/PKG pathway. A complete understanding of this signaling pathway will enable a more rationale approach to selecting therapeutic targets, including PDE inhibitors, which slow the progression of muscle degeneration and ameliorate the dystrophic phenotype.
The research proposed here will provide new information about the mechanisms that lead to muscle degeneration in muscular dystrophies. The focus on a novel signaling mechanism at the Golgi, a cellular organelle that regulates protein modification and trafficking to the surface membrane, may reveal new therapeutic targets for slowing the progress of muscle degeneration. In general, the knowledge generated by this project will be relevant to many types of muscle diseases.
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|Allen, David G; Whitehead, Nicholas P; Froehner, Stanley C (2016) Absence of Dystrophin Disrupts Skeletal Muscle Signaling: Roles of Ca2+, Reactive Oxygen Species, and Nitric Oxide in the Development of Muscular Dystrophy. Physiol Rev 96:253-305|
|Froehner, Stanley C; Reed, Sarah M; Anderson, Kendra N et al. (2015) Loss of nNOS inhibits compensatory muscle hypertrophy and exacerbates inflammation and eccentric contraction-induced damage in mdx mice. Hum Mol Genet 24:492-505|
|Patrucco, Enrico; Domes, Katrin; Sbroggió, Mauro et al. (2014) Roles of cGMP-dependent protein kinase I (cGKI) and PDE5 in the regulation of Ang II-induced cardiac hypertrophy and fibrosis. Proc Natl Acad Sci U S A 111:12925-9|
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|Percival, Justin M; Siegel, Michael P; Knowels, Gary et al. (2013) Defects in mitochondrial localization and ATP synthesis in the mdx mouse model of Duchenne muscular dystrophy are not alleviated by PDE5 inhibition. Hum Mol Genet 22:153-67|
|Tsai, L-C L; Chan, G C-K; Nangle, S N et al. (2012) Inactivation of Pde8b enhances memory, motor performance, and protects against age-induced motor coordination decay. Genes Brain Behav 11:837-47|
|Percival, Justin M; Whitehead, Nicholas P; Adams, Marvin E et al. (2012) Sildenafil reduces respiratory muscle weakness and fibrosis in the mdx mouse model of Duchenne muscular dystrophy. J Pathol 228:77-87|
|Percival, Justin M; Adamo, Candace M; Beavo, Joseph A et al. (2011) Evaluation of the therapeutic utility of phosphodiesterase 5A inhibition in the mdx mouse model of duchenne muscular dystrophy. Handb Exp Pharmacol :323-44|
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