title: Soluble adenylyl cyclase is localized to cilia and contributes to ciliary beat frequency regulation via production of cAMP.
abstract: Ciliated airway epithelial cells are subject to sustained changes in intracellular CO(2)/HCO(3)(-) during exacerbations of airway diseases, but the role of CO(2)/HCO(3)(-)-sensitive soluble adenylyl cyclase (sAC) in ciliary beat regulation is unknown. We now show not only sAC expression in human airway epithelia (by RT-PCR, Western blotting, and immunofluorescence) but also its specific localization to the axoneme (Western blotting and immunofluorescence). Real time estimations of [cAMP] changes in ciliated cells, using FRET between fluorescently tagged PKA subunits (expressed under the foxj1 promoter solely in ciliated cells), revealed CO(2)/HCO(3)(-)-mediated cAMP production. This cAMP production was specifically blocked by sAC inhibitors but not by transmembrane adenylyl cyclase (tmAC) inhibitors. In addition, this cAMP production stimulated ciliary beat frequency (CBF) independently of intracellular pH because PKA and sAC inhibitors were uniquely able to block CO(2)/HCO(3)(-)-mediated changes in CBF (while tmAC inhibitors had no effect). Thus, sAC is localized to motile airway cilia and it contributes to the regulation of human airway CBF. In addition, CO(2)/HCO(3)(-) increases indeed reversibly stimulate intracellular cAMP production by sAC in intact cells.
authors.affiliation: Division of Pulmonary and Critical Care, University of Miami, Miami, FL 33136, USA.
authors.forename: Andreas
authors.initials: A
authors.lastname: Schmid
authors.forename: Zoltan
authors.initials: Z
authors.lastname: Sutto
authors.forename: Marie-Christine
authors.initials: MC
authors.lastname: Nlend
authors.forename: Gabor
authors.initials: G
authors.lastname: Horvath
authors.forename: Nathalie
authors.initials: N
authors.lastname: Schmid
authors.forename: Jochen
authors.initials: J
authors.lastname: Buck
authors.forename: Lonny R
authors.initials: LR
authors.lastname: Levin
authors.forename: Gregory E
authors.initials: GE
authors.lastname: Conner
authors.forename: Nevis
authors.initials: N
authors.lastname: Fregien
authors.forename: Matthias
authors.initials: M
authors.lastname: Salathe
mesh_terms: D000262:Adenylyl Cyclases / Q000378:metabolism*; D002478:Cells, Cultured; D002923:Cilia / Q000201:enzymology* / Q000502:physiology*; D000242:Cyclic AMP / Q000378:metabolism*; D004847:Epithelial Cells / Q000378:metabolism; D006801:Humans; D008168:Lung / Q000166:cytology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012995:Solubility
chemical_list: D000242:Cyclic AMP; D000262:Adenylyl Cyclases