The overall goal of this project is to understand the role of gallbladder mucin in the pathogenesis of cholesterol gallstones. Medical or surgical treatment for symptomatic gallstones is a frequent cause for hospitalization in the United States and consumes more than 8 billion dollars in health care costs per year. Gallbladder mucin has been shown to have a central role in gallstone pathogenesis. Mucin hypersecretion occurs prior to stone formation in both experimental animals and man. Mucin also promotes cholesterol crystal nucleation, a critical early step in stone formation, and the mucin gel lining. The gallbladder provides an ideal environment for growth of cholesterol crystals into mature stones. Human gallbladder expresses genes for five different mucins and two of these, MUC5B and MUC3 have been identified as the major human gallbladder mucins. Nothing is known about the structural features of these proteins which interact with biliary lipids and lead to the lithogenic state. In this project, a novel recombinant approach will be used to identify the structural domains of MUC5B and MUC3 which bind biliary lipids.
The specific aims of this proposal are to: (l) characterize the major human gallbladder mucins MUC5B and MUC3 by determining the nucleotide and deduced amino acid sequences of the poorly glycosylated amino- and carboxyl-terminal regions, (2) determine the genomic organization of these mucins and (3) to identify functional domains in MUC5B and MUC3. Information obtained in specific aims 1 and 2 will be used to design constructs containing individual mucin domains. Recombinant mucin polypeptides will be expressed in bacteria and examined in lipid binding, cholesterol crystal nucleation and vesicle fusion assays. The results of these studies will provide new information about the relationship between the structure of the major human gallbladder mucins and their function in normal and pathologic gallbladder. This information is necessary for the rational design of therapies for the prevention and treatment of gallstone disease.
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