The overall objective of the proposed research is to delineate the molecular bases of the marked phenotypic variability in Gaucher disease (GD), a prototype inborn error of metabolism. The proposed studies address the hypotheses: 1) The mutations that predispose to GD are associated with disruptions of acid B-glucosidase [glucosylceramide (GC) glucohydrolase; UCase: GBA locus] structure and function leading to differential threshold levels of enzymatic activity in various patient tissues. Although this is a major basis of the phenotypic spectrum, intra- and inter- locus sequence variants and polymorphisms may provide for additional contexts for phenotypic expression of disease-related mutations, e.g., altered substrate flux. To understand the interplay of such sequence variations, the determinants for the control of normal and mutant GCase enzyme activities will be evaluated with liposomal (in vitro) and lysosomal (ex vivo) membrane binding systems. 2) The in vivo levels of specific substrate synthesis and degradation, e.g., GC synthase (GCS) expression, and hydrolase activity, in selected inborn errors of glycosphingolipid (GSL) metabolism are primary determinants of their regional, tissue or cellular pathophysiology. The functional polymorphic variation at the GCS and GCase loci and the relationships of these variations to defined phenotypic parameters will be determined in patients with GD type 1. As a corollary, effective enzyme or gene therapy in GD, as a prototype, requires specific levels of enzyme in various organs. 3) The lack of adequate mouse models for GD have been a major impediment to continuing progress in pathophysiologic understanding and therapeutic developments, and their creation and characterization is a major focus of this proposal. We have developed conditional (tetracycline-on) and fixed (five specific point mutations) GCase expressing mice to simulate human GD variants. These alternative systems, together with the GCS KO heterozgyote mice will be used to address GC flux in relation to phenotype in vivo. These studies should provide insights into the pathophysiology and therapy of GD, and to over 20 glycolipid storage diseases that depend on the GCS synthetic and GCase degradative pathwavs.
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