1.6.1 Structure and Function of the CFTR: The human CFTR (hCFTR) is a 1480 amino 
acid polypeptide (Mr= 168,021) with distinct structural domains characterized by distinct 
hydrophobic regions each corresponding to transmembrane segments (Kerem et.al., 1989). A 
putative regulatory, or R domain, and two nucleotide binding folds in which resides the A508 
mutation (fold 1) are characteristic of the CFTR polypeptide in all species studied to date. The 
CFTR protein is membrane bound, binds to and hydrolyses ATP, and is both a phosphoprotein 
and a glycoprotein (Kerem et.al., 1989, Riordan et.al., 1989, Cheng et.al., 1991, Gregory 
et.al., 1990). Transfer of the full-length cDNA encoding CFTR confers cAMP-dependent 
chloride pump activity that will complement the chloride transport defect characteristic of cells 
derived from cystic fibrosis patients (Rich et.al., 1990, Drumm et.al., 1990). Evidence to date 
strongly supports the role of the CFTR as a chloride transporting protein. Site-specific 
mutations of the CFTR transmembrane region are associated with alterations in the selectivity 
of halide ions (Anderson et.al., 1990). However, the strong structural relationship of CFTR to 
multiple drug resistance proteins (MDR) and other members of the ABC co-transport protein 
family supports its role as an energy-dependent transporter as well. Abnormalities of routing 
and processing of CFTR mutant protein are important components of the cellular disorder 
(Cheng SH, et.al., 1990). The hCFTR cDNA has been transferred to a variety of cystic fibrosis 
and non-cystic fibrosis cell types wherein the CFTR protein was identified immunohistochemical- 
ly and associated with the generation of cAMP-dependent chloride transport activity. This 
activity is distinct from calcium-dependent chloride channels which appear to be present in most 
cells and distinct from the cystic fibrosis defect. These calcium-dependent regulated chloride 
channels appear to be intact in cells from cystic fibrosis patients. While the precise manner by 
which abnormalities of the CFTR cause pulmonary disease and infection are not known, the 
rapid expansion of our understanding of the molecular aspects of CFTR has provided reagents 
required for transfer of the CFTR gene to cells, animals and humans for somatic cell gene 
therapy of CF. 
Cystic fibrosis is a disease primarily affecting electrolyte transport in secretory epithelia 
(Davis, P. 1991). Pulmonary complications of cystic fibrosis represent the major life-threatening 
abnormality in cystic fibrosis, ^/hile growth and function of the lungs in cystic fibrosis patients 
are generally normal at birth, pulmonary complications begin as early as the first year of life and 
generally are characterized by thick pulmonary secretions, mucous plugging and the inability to 
clear respiratory pathogens from the lung, particularly bacterial organisms such as Pseudomonas 
aerugenosa and Staphylococcus aureus . Chronic pulmonary infection, atelectasis and pneumonia 
lead to bronchiectasis, and fibrosis, ultimately leading to cor pulmonale and respiratory failure. 
The importance of the lung complications in cystic fibrosis has focused attention to the CFTR 
defect in the respiratory epithelium, rather than other organs, for therapy of cystic fibrosis. 
1.7 Cells of the Respiratory Epithelium: The respiratory tract begins as an outpouching of 
the foregut endoderm early in the first trimester of human development and on day 9.6 in the 
mouse embryo. Initially, the respiratory epithelium consists of undifferentiated columnar 
epithelial cells which form a dichotomously-branching respiratory tract during the glandular and 
pseudoglandular period of development. After 4-5 months gestation in the human, during the 
canalicular period, the epithelial elements of the proximal respiratory tract interact more closely 
with complex mesenchymal components including blood vessels, smooth muscle and endothelial 
cells, and begin generating a highly differentiated pattern of cells in the tracheal, bronchial and 
bronchiolar region of the lung. The differentiation of the proximal respiratory epithelium 
proceeds until a variety of cell types are produced in the respiratory tract, including squamous, 
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Recombinant DNA Research, Volume 17 
