Defining the Role of Laforin in Glycogen Metabolism and Lafora Disease

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Project Summary/Abstract The goal of this proposal is to use the unique window offered by Lafora disease (LD) to understand how laforin regulates glycogen metabolism and define the molecular perturbation of LD patient mutations. Lafora disease (LD), one of the five major progressive myoclonus epilepsies, is a fatal, genetic, neurological disorder that manifests during adolescence and invariably leads to neurodegeneration and death. Patients with LD display accumulations of hyperphosphorylated, aberrantly branched, glycogen-like particles called Lafora bodies (LBs). LBs are found in most tissues but neurological symptoms predominate in LD patients. Mutations in the gene encoding the glycogen phosphatase laforin cause approximately 70% of LD cases. LD largely centers on glycogen metabolism. Glycogen is the most important carbohydrate storage molecule in mammals, and misregulation of glycogen is implicated in many diseases. Recent studies show that glycogen plays a dynamic role in brain function and that neurons are highly sensitive to glycogen perturbations. Although glycogen has been studied for many years, gaps in our understanding of its regulation remain. The Gentry lab established laforin as the founding member of the glucan phosphatase family (i.e. enzymes that release phosphate from carbohydrates), and studies on LD show that the absence of laforin triggers the transformation of glycogen into the neurotoxic carbohydrate that makes up LBs. As the only glucan phosphatase in humans, laforin is the lynchpin of glycogen regulation by phosphorylation. We recently determined the crystal structure of laforin. This proposal will couple insights from the structure with biochemical and biophysical tools to define how laforin functions as a glucan phosphatase and regulates glycogen metabolism. Further, the 33 LD-associated missense mutations are scattered throughout the structure and have different effects on laforin activity. A goal in our lab is to define the effect of all of these mutations. Aim 1A will address the nature of the dimer interface, which has been controversial in the field, and the molecular effect of LD mutations in this region. Aim 1B will define how laforin cooperatively binds the glucan chains in glycogen and the effects of length and branching. Aim 2 will establish the role of the interdomain region of laforin (i.e. between the phosphatase and carbohydrate-binding domains) in maintaining C3- and C6-specific dephosphorylation of glycogen. A number of LD mutations fall in this region and this aim will demonstrate their functional effects. In summary, this proposal utilizes cross-disciplinary techniques to study glycogen homeostasis and the mechanisms of LD, studies that will provide insights into brain metabolism and pave the way for patient-specific diagnoses and treatments.