The long-term objectives of these investigations are to understand the cellular mechanisms of how a single gene disorder (glycerol kinase deficiency) causes a complex phenotypic disease. Glycerol kinase (GK) catalyzes the phosphorylation of glycerol into glycerol 3-phosphate and is at the interface of glucose and fat metabolism. Glycerol kinase deficiency (GKD) is an X-linked disorder of metabolism that is due to mutations and/or deletions of the glycerol kinase gene (GK). Patients with GKD are phenotypically either symptomatic or asymptomatic. Our initial work on glycerol kinase deficiency has shown that there is no way to predict which patients will be symptomatic and which will be asymptomatic by GK activity or the location of the mutation in a model of the three dimensional structure of the protein. We hypothesize that it is the interaction of GK mutations with additional genetic and environmental influences on metabolic flux (other enzymes in related pathways and levels of critical intermediates) that are important to understand the pathogenesis of this disorder. The goals of this proposal are to understand better the complex interactions within the cell and how perturbations of an individual enzyme (GK) affects the other pathways and gene expression to result in the physiological changes seen in the whole animal. Our first Specific Aim (Task 1) is to characterize the metabolic pathways relating to GK and how they are changed in GKD using lymphoblastoid cells lines from the individuals with GKD as well as normal individuals. Metabolome analysis, flux analysis, and transcriptome analysis will allow us to investigate the effect of the mutations with varying levels of GK activity in the context of the individuals' genetic background. We will then investigate the effect of these mutations in liver and kidney cell lines as these tissues have the highest level of GK expression and will allow us to investigate the role of the GK mutations within the context of identical genetic background. Results of these studies will allow us to perform metabolome, flux, and transcriptome analysis in the whole animal model using the glycerol kinase knock-out (gyk k/o) mouse (Specific Aim/Task 2). These studies will provide a model system to understand the complex nature of genetic disorders and eventually help in treatment of such disorders.
