Cystic fibrosis (CF) is the most prevalent autosomal recessive lethal genetic disease in the U.S. with 5% of the population carrying a mutant CFTR gene. The long term goals of this proposal are to understand how mutant CFTRs fail to function correctly in CF patients and how these failures might be repaired. The deltaF508 mutant of CFTR is retained in the endoplasmic reticulum (ER) and thereby fails to traffic properly to the apical membrane of epithelial cells. While the mechanism of this trafficking failure may involve recognition of improperly folded CFTR by protein chaperones, preliminary data indicate that phospholipid interactions with the first nucleotide binding fold domain (NBF-1) might also play a role in CFTR trafficking. Dr. Pollard and his colleagues have therefore hypothesized that aberrant interactions between the mutant NBF-1 domain and specific phospholipids might contribute to the ER retention and degradation, and thus cause the disease. To test this hypothesis, the investigators propose: 1) to determine the specificity of phospholipid interaction with NBF-1 and the alteration of this specificity by the deltaF508 mutation by measuring lipid interaction with wild type or mutant NBF-1 using biophysical assays; and 2) to show that NBF-1 induces permeability changes in membranes, and that deltaF508 mutation changes the lipid specificity. This property will be used to develop a screening assay for identifying drugs that might affect mutant CFTR trafficking in CF patients; 3) to show that changes in the phospholipid specificity of NBF-1 have direct consequences for CFTR trafficking in vivo.
This aim will be approached by analyzing wild type and deltaF508-CFTR trafficking in a cell line which is temperature sensitive for lipid biosynthesis. The significance of this proposal is that it suggests a novel approach to the mechanism of CF, involving a trafficking defect affected by phospholipid interactions with the mutant protein. Identifying this defect can provide targets for repair. If successful, this approach will set the stage for development of therapeutic means for correcting the aberrant interactions between mutant CFTR and specific phospholipids.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
1R01DK053051-01A1
Application #
2628917
Study Section
Medical Biochemistry Study Section (MEDB)
Program Officer
Mckeon, Catherine T
Project Start
1998-06-15
Project End
2001-05-31
Budget Start
1998-06-15
Budget End
1999-05-31
Support Year
1
Fiscal Year
1998
Total Cost
Indirect Cost
Name
Henry M. Jackson Fdn for the Adv Mil/Med
Department
Type
DUNS #
City
Rockville
State
MD
Country
United States
Zip Code
20817
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