title: CFTR Cl- channel functional regulation by phosphorylation of focal adhesion kinase at tyrosine 407 in osmosensitive ion transporting mitochondria rich cells of euryhaline killifish.
abstract: Cystic fibrosis transmembrane conductance regulator (CFTR) anion channels are the regulated exit pathway in Cl(-) secretion by teleost mitochondria rich salt secreting (MR) cells of the gill and opercular epithelia of euryhaline teleosts. By confocal light immunocytochemistry, immunogold transmission electron microscopy (TEM), and co-immunoprecipitation, using regular and phospho-antibodies directed against conserved sites, we found that killifish CFTR (kfCFTR) and the tyrosine kinase focal adhesion kinase (FAK) phosphorylated at Y407 (FAK pY407) are colocalized in the apical membrane and in subjacent membrane vesicles of MR cells. We showed previously that basolateral FAK pY407, unlike other FAK phosphorylation sites, is osmosensitive and dephosphorylates during hypotonic shock of epithelial cells (Marshall et al., 2008). In the present study, we found that hypotonic shock and the alpha(2)-adrenergic agonist clonidine (neither of which affects cAMP levels) rapidly and reversibly inhibit Cl(-) secretion by isolated opercular membranes, simultaneous with dephosphorylation of FAK pY407, located in the apical membrane. FAK pY407 is rephosphorylated and Cl(-) secretion rapidly restored by hypertonic shock as well as by forskolin and isoproterenol, which operate via cAMP and protein kinase A. We conclude that hormone mediated, cAMP dependent and osmotically mediated, cAMP independent pathways converge on a mechanism to activate CFTR and Cl(-) secretion, possibly through tyrosine phosphorylation of CFTR by FAK.
authors.affiliation: Department of Biology, St Francis Xavier University, PO Box 5000 Antigonish, Nova Scotia, Canada B2G 2W5. bmarshal@stfx.ca
authors.forename: William S
authors.initials: WS
authors.lastname: Marshall
authors.forename: Kaitlyn D
authors.initials: KD
authors.lastname: Watters
authors.forename: Leah R
authors.initials: LR
authors.lastname: Hovdestad
authors.forename: Regina R F
authors.initials: RR
authors.lastname: Cozzi
authors.forename: Fumi
authors.initials: F
authors.lastname: Katoh
mesh_terms: D000818:Animals; D003000:Clonidine / Q000494:pharmacology; D005576:Colforsin / Q000494:pharmacology; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000032:analysis / Q000378:metabolism*; D004594:Electrophysiology; D051416:Focal Adhesion Protein-Tyrosine Kinases / Q000032:analysis / Q000378:metabolism*; D023781:Fundulidae / Q000378:metabolism*; D006982:Hypertonic Solutions / Q000494:pharmacology; D007038:Hypotonic Solutions / Q000494:pharmacology; D017136:Ion Transport; D007545:Isoproterenol / Q000494:pharmacology; D046529:Microscopy, Electron, Transmission; D016253:Microscopy, Immunoelectron; D008928:Mitochondria / Q000378:metabolism*; D010766:Phosphorylation; D021381:Protein Transport / Q000187:drug effects; D013565:Sympatholytics / Q000494:pharmacology; D013566:Sympathomimetics / Q000494:pharmacology; D014443:Tyrosine / Q000378:metabolism
chemical_list: D006982:Hypertonic Solutions; D007038:Hypotonic Solutions; D013565:Sympatholytics; D013566:Sympathomimetics; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D005576:Colforsin; D014443:Tyrosine; D051416:Focal Adhesion Protein-Tyrosine Kinases; D007545:Isoproterenol; D003000:Clonidine