Advances in understanding of the molecular basis of cystic fibrosis, although still far from complete, have enabled the initiation of new approaches to therapy. Several strategies are feasible, but it is not yet possible to predict which will be successful. Gene therapy is the most direct and obvious approach and is appropriately being intensively pursued. Additionally, new knowledge of the steps between mutations in the CFTR gene and pathological changes in the patient provide other potential sites of intervention. This SCOR will focus on four of these. Project I will further elucidate the molecular events involved in the biosynthetic arrest and subcellular mislocalization of deltaF508 CFTR which is present in more than 90% of patients. Very recent evidence suggests that the formation of a complex between the mutant molecule and a chaperone which is resident in the endoplasmic reticulum may contribute to the intracellular retention. Perturbation of the tight complex which the mutant molecule forms with the chaperone may be possible with inhibitors of glycosylation because oligosaccarides are required for complex formation. Project II aims to enhance the functional capacity of CFTR by genetically changing it or by altering the phosphorylation- dephosphorylation cycle involved in its regulation, a maneuver which can activate several mutant forms of CFTR when certain phosphatase inhibitors are employed. Project III will pursue exciting new findings on a molecular network which mediates osmotic regulation of cellular chloride permeability. One member of this group, IClnm, has been cloned and characterized and now provides a means of access to other molecules involved, including a novel chloride channel with properties distinct from those previously described. It may be possible to circumvent the defective CFTR chloride channel by harnessing this pathway. Project IV approaches the CF problem still further downstream from the CFTR at the level of mucin production. Using recently-generated DNA and antibody probes for different members of the mucin super family, the basis of mucin overexpression in CF will be explored at the level of the mucus cells themselves. The ultimate objective is to learn enough about the mechanism of up-regulation to be able to prevent or reduce it. These four projects will rely on four scientific core resources, including cellular and transgenic expression, sophisticated multi-label subcellular imaging and access to large CF clinic facilities.