NAD+ Biosynthesis Ameliorates a Zebrafish Model of Muscular Dystrophy

Abstract

Muscular dystrophies are common, currently incurable diseases. A subset of dystrophies result from genetic disruptions in complexes that attach muscle fibers to their surrounding extracellular matrix microenvironment. Cell-matrix adhesions are exquisite sensors of physiological conditions and mediate responses that allow cells to adapt to changing conditions. Thus, one approach towards finding targets for future therapeutic applications is to identify cell adhesion pathways that mediate these dynamic, adaptive responses in vivo. We find that nicotinamide riboside kinase 2b-mediated NAD+ biosynthesis, which functions as a small molecule agonist of muscle fiber-extracellular matrix adhesion, corrects dystrophic phenotypes in zebrafish lacking either a primary component of the dystrophin-glycoprotein complex or integrin alpha7. Exogenous NAD+ or a vitamin precursor to NAD+ reduces muscle fiber degeneration and results in significantly faster escape responses in dystrophic embryos. Overexpression of paxillin, a cell adhesion protein downstream of NAD+ in this novel cell adhesion pathway, reduces muscle degeneration in zebrafish with intact integrin receptors but does not improve motility. Activation of this pathway significantly increases organization of laminin, a major component of the extracellular matrix basement membrane. Our results indicate that the primary protective effects of NAD+ result from changes to the basement membrane, as a wild-type basement membrane is sufficient to increase resilience of dystrophic muscle fibers to damage. The surprising result that NAD+ supplementation ameliorates dystrophy in dystrophin-glycoprotein complex– or integrin alpha7–deficient zebrafish suggests the existence of an additional laminin receptor complex that anchors muscle fibers to the basement membrane. We find that integrin alpha6 participates in this pathway, but either integrin alpha7 or the dystrophin-glycoprotein complex is required in conjunction with integrin alpha6 to reduce muscle degeneration. Taken together, these results define a novel cell adhesion pathway that may have future therapeutic relevance for a broad spectrum of muscular dystrophies. A variety of diseases, both inherited and acquired, affect muscle tissues in humans. Critical to muscle homeostasis is the anchoring of muscle fibers to their surrounding microenvironment through cell adhesion complexes that help to resist the repeated stress experienced during muscle contraction. Genetic mutations in these complexes weaken this mechanical attachment, making fibers more susceptible to damage and death. The resulting increased fiber degeneration can eventually lead to progressive muscle-wasting diseases, known collectively as muscular dystrophies. Although clinical trials are ongoing, there is presently no way to cure the loss of muscle structure and function associated with these diseases. We identified a novel cell adhesion pathway involving integrin alpha6 that promotes adhesion of muscle cells to their microenvironment. Here, we show that activation of this pathway not only significantly reduces muscle degeneration but also improves the swimming ability of dystrophic zebrafish. We explore the likely benefits and limitations of this pathway in treating symptoms of congenital muscular dystrophies. Our findings suggest that activation of this pathway (for example, by boosting levels of NAD+) has the potential to ameliorate loss of muscle structure and function in multiple muscular dystrophies.