The goals of this proposal are to use metabolomics analysis to understand the mechanisms of neuromuscular diseases including a novel mouse mutation in Nadkd1, an uncharacterized NAD Kinase family member that localizes to mitochondria, and to develop informatics tools for the analysis of metabolomics data obtained from the laboratory mouse. The use of mass spectroscopy-based analysis of low molecular weight metabolites (metabolomics) has the potential to identify disease mechanisms based on changes in metabolites that are directly impacted by the disease process. Such an unbiased, broad-scope approach is particularly valuable when novel genes with unknown functions are mutated. This is the case for mutations in Nadkd1, which cause neuromuscular degeneration in mice, with overt onset of neuromuscular disease symptoms at approximately eight weeks of age and death by approximately 5 months of age. Neuromuscular junctions appear to progressively denervate in these mice, leading to loss of motor function resembling a distal spinal muscular atrophy. The NADKD1 protein is in the NAD Kinase family of small molecule kinases, and localizes to mitochondria, but the substrate and function of NDAKD1 is unknown. Given the putative small molecule kinase activity and the mitochondrial localization, metabolite profiling will be used to determine the substrate and biochemical pathway for this protein by identifying accumulated substrate and reduced phosphorylated product. We have a preliminary metabolomics data set from a mouse model of Charcot-Marie-Tooth 2D peripheral neuropathy that will be used for comparison. However, this preliminary study also highlighted a number of challenges in the analysis and interpretation of metabolomics data. We are therefore proposing two specific aims. First, we will perform additional metabolomic analyses to compare the Nadkd1 mutant mice to other relevant models and appropriate controls. These comparisons will include 1) pre- and 2) post-onset Nadkd1 mutant mice, and 3) Pla2g6 mutant mice, a model of Infantile Neuroaxonal Dystrophy (INAD), which also results in denervation and peripheral axon degeneration. Each comparative study has merit on its own, and in aggregate, we will be able to compare Nadkd1-associated profiles to other genetic neuromuscular models to determine which changes are specifically found in the Nadkd1 mutant mouse and therefore may be directly related to the gene function and disease mechanism. Second, we will develop a web-based bioinformatics resource specifically designed for the analysis of data from metabolomics data generated for the laboratory mouse. This resource, called the Mouse Metabolic Analysis Platform (mMAP) will facilitate the integration of metabolomics data with other genomics data and with information from existing knowledgebases of biochemical pathways and biological annotations for mouse genes and proteins. The informatics tools developed for this exploratory R21 will be generally applicable to future metabolomics studies of mouse models of human disease.
Neuromuscular diseases are a major clinical and economic issue in health care, but current therapies are limited in part because of poor understanding of disease mechanisms. We have identified a novel mouse mutation that causes neuromuscular disease. We are proposing to use a large-scale, unbiased analysis of metabolites to understand the disease mechanism in this mouse. This work will improve our understanding of neuromuscular disease, identify new avenues for therapy, and create the bioinformatics tools necessary for analyzing metabolite profiles, which are currently lacking despite a rapidly increasing number of studies using this approach.
|Bais, Preeti; Beebe, Kirk; Morelli, Kathryn H et al. (2016) Metabolite profile of a mouse model of Charcot-Marie-Tooth type 2D neuropathy: implications for disease mechanisms and interventions. Biol Open 5:908-20|