Cystic fibrosis (CF) is the most common severe autosomal recessive disorder in the Caucasian population. In North America, approximately 1 in 2,500 live-births is affected with CF, which is characterized by chronic obstructive lung disease, pancreatic insufficiency, abnormalities of electrolyte, fluid and macromolecule secretion of exocrine glands. The basic biochemical defect is unknown. The identification of the CF gene has provided an opportunity to understand the defect and the pathophysiology of the disease at the molecular level, which will allow the development of rational therapy. On the basis of DNA sequence analysis, the CF gene product (CFTR) is predicted to be a transmembrane protein with 2 ATP-binding domains. Genetic analysis shows that approximately 70% of the CF chromosomes suffer a 3 base pair deletion which corresponds to a single amino acid deletion at position 508 of CFTR.
Five specific aims are proposed in this application: (1) Additional mutations in the CF gene will be identified in order to map the functional domains of CFTR; simple detection procedures will be developed for each mutation; a number of mutations have already been identified for the remaining 30% of CF chromosomes. (2) Mutations and """"""""epitope tags"""""""" will be introduced into selected regions of CFTR to map its functional domains and topology in relation to the plasma membrane in mammalian cells. (3) The effect on the biosynthesis of CFTR of some of the naturally occurring mutations, especially the nonsense and frameshift mutations that have been detected, will be studied to investigate if true """"""""null"""""""" mutations exist in CF. (4) In order to understand the factors governing CF gene expression, the sequence elements responsible for basal promoter activity and tissue-specificity for CFTR will be determined by conventional techniques with the use of reporter gene constructs (CAT); transgenic mice carrying the CF gene promoter and a reporter gene (E. coli lacZ) will also be constructed to study developmental regulation of CFTR. (5) DNA sequences that have been detected and shown in preliminary studies, to be closely related to CFTR in the human genome will be isolated and characterized, with respect to their chromosomal location, expression pattern, and their structural and functional relationships with CFTR. The latter information may provide important insight into the evolution function of the CF gene and other CFTR-like genes. These studies, conducted in parallel with 2 other separately funded approaches (use of yeast and mouse models), constitute a comprehensive research program in elucidating the basic defect in CF.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
2R01DK034944-07
Application #
3233186
Study Section
Cellular Biology and Physiology Subcommittee 1 (CBY)
Project Start
1985-01-01
Project End
1995-12-31
Budget Start
1991-02-12
Budget End
1991-12-31
Support Year
7
Fiscal Year
1991
Total Cost
Indirect Cost
Name
Hospital for Sick Chldrn (Toronto)
Department
Type
DUNS #
208511808
City
Toronto
State
ON
Country
Canada
Zip Code
M5 1-X8
Browning, Kirsteen N; Mendelowitz, David (2003) Musings on the wanderer: what's new in our understanding of vago-vagal reflexes?: II. Integration of afferent signaling from the viscera by the nodose ganglia. Am J Physiol Gastrointest Liver Physiol 284:G8-14
Orozco, L; Zielenski, J; Markiewicz, D et al. (1997) Two novel frameshift deletions (1924del7, 2055del9-->A) in the CFTR gene in Mexican cystic fibrosis patients. Hum Mutat 10:239-40
Morral, N; Dork, T; Llevadot, R et al. (1996) Haplotype analysis of 94 cystic fibrosis mutations with seven polymorphic CFTR DNA markers. Hum Mutat 8:149-59
Zielenski, J; Markiewicz, D; Lin, S P et al. (1995) Skipping of exon 12 as a consequence of a point mutation (1898 + 5G-->T) in the cystic fibrosis transmembrane conductance regulator gene found in a consanguineous Chinese family. Clin Genet 47:125-32
Zielenski, J; Markiewicz, D; Chen, H S et al. (1995) Identification of six mutations (R31L, 441delA, 681delC, 1461ins4, W1089R, E1104X) in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Hum Mutat 5:43-7
Jarvi, K; Zielenski, J; Wilschanski, M et al. (1995) Cystic fibrosis transmembrane conductance regulator and obstructive azoospermia. Lancet 345:1578
Bozon, D; Zielenski, J; Rininsland, F et al. (1994) Identification of four new mutations in the cystic fibrosis transmembrane conductance regulator gene: I148T, L1077P, Y1092X, 2183AA-->G. Hum Mutat 3:330-2
Zielenski, J; Bozon, D; Markiewicz, D et al. (1993) Analysis of CFTR transcripts in nasal epithelial cells and lymphoblasts of a cystic fibrosis patient with 621 + 1G-->T and 711 + 1G-->T mutations. Hum Mol Genet 2:683-7
Tsui, L C (1992) Mutations and sequence variations detected in the cystic fibrosis transmembrane conductance regulator (CFTR) gene: a report from the Cystic Fibrosis Genetic Analysis Consortium. Hum Mutat 1:197-203
Mornet, E; Chateau, C; Simon-Bouy, B et al. (1992) Carrier detection and prenatal diagnosis of cystic fibrosis using an intragenic TA-repeat polymorphism. Hum Genet 88:479-81

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