title: Role of Exchange Protein Activated by cAMP 1 in Regulating Rates of Microtubule Formation in Cystic Fibrosis Epithelial Cells.
abstract: The regulation of microtubule dynamics in cystic fibrosis (CF) epithelial cells and the consequences of reduced rates of microtubule polymerization on downstream CF cellular events, such as cholesterol accumulation, a marker of impaired intracellular transport, are explored here. It is identified that microtubules in both CF cell models and in primary CF nasal epithelial cells repolymerize at a slower rate compared with respective controls. Previous studies suggest a role for cAMP in modulating organelle transport in CF cells, implicating a role for exchange protein activated by cAMP (EPAC) 1, a regulator of microtubule elongation, as a potential mechanism. EPAC1 activity is reduced in CF cell models and in Cftr(-/-) mouse lung compared with respective non-CF controls. Stimulation of EPAC1 activity with the selective EPAC1 agonist, 8-cpt-2-O-Me-cAMP, stimulates microtubule repolymerization to wild-type rates in CF cells. EPAC1 activation also alleviates cholesterol accumulation in CF cells, suggesting a direct link between microtubule regulation and intracellular transport. To verify the relationship between transport and microtubule regulation, expression of the protein, tubulin polymerization-promoting protein, was knocked down in non-CF human tracheal (9/HTEo(-)) cells to mimic the microtubule dysregulation in CF cells. Transduced cells with short hairpin RNA targeting tubulin polymerization-promoting protein exhibit CF-like perinuclear cholesterol accumulation and other cellular manifestations of CF cells, thus supporting a role for microtubule regulation as a mechanism linking CFTR function to downstream cellular manifestation.
authors.affiliation: Departments of 1 Pharmacology.|2 Pediatrics, and.
authors.forename: Sharon M
authors.initials: SM
authors.lastname: Rymut
authors.affiliation: 2 Pediatrics, and.
authors.forename: Tracy
authors.initials: T
authors.lastname: Ivy
authors.affiliation: 2 Pediatrics, and.
authors.forename: Deborah A
authors.initials: DA
authors.lastname: Corey
authors.affiliation: 2 Pediatrics, and.
authors.forename: Calvin U
authors.initials: CU
authors.lastname: Cotton
authors.affiliation: 3 Chemistry, Case Western Reserve University, Cleveland, Ohio.
authors.forename: James D
authors.initials: JD
authors.lastname: Burgess
authors.affiliation: Departments of 1 Pharmacology.|2 Pediatrics, and.|3 Chemistry, Case Western Reserve University, Cleveland, Ohio.
authors.forename: Thomas J
authors.initials: TJ
authors.lastname: Kelley
mesh_terms: D000818:Animals; D002460:Cell Line; D000242:Cyclic AMP; D003550:Cystic Fibrosis / Q000378:metabolism* / Q000473:pathology; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000502:physiology; D004847:Epithelial Cells / Q000378:metabolism*; D020662:Guanine Nucleotide Exchange Factors / Q000502:physiology*; D006801:Humans; D007700:Kinetics; D018345:Mice, Knockout; D008870:Microtubules / Q000378:metabolism* / Q000473:pathology; D055503:Protein Multimerization; D020545:Respiratory Mucosa / Q000378:metabolism / Q000473:pathology
chemical_list: C505032:CFTR protein, human; D020662:Guanine Nucleotide Exchange Factors; C116122:RAPGEF3 protein, human; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D000242:Cyclic AMP
keywords: Rap1; cholesterol; cystic fibrosis; exchange protein activated by cAMP 1; microtubules