TY - JOUR
T1 - Soluble adenylyl cyclase is localized to cilia and contributes to ciliary beat frequency regulation via production of cAMP
AU - Schmid, Andreas
AU - Sutto, Zoltan
AU - Nlend, Marie Christine
AU - Horvath, Gabor
AU - Schmid, Nathalie
AU - Buck, Jochen
AU - Levin, Lonny R.
AU - Conner, Gregory E.
AU - Fregien, Nevis
AU - Salathe, Matthias
PY - 2007/7
Y1 - 2007/7
N2 - Ciliated airway epithelial cells are subject to sustained changes in intracellular CO2/HCO3- during exacerbations of airway diseases, but the role of CO2/HCO3- -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 CO2/HCO3--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 CO2/HCO3 --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, CO2/HCO3 - increases indeed reversibly stimulate intracellular cAMP production by sAC in intact cells.
AB - Ciliated airway epithelial cells are subject to sustained changes in intracellular CO2/HCO3- during exacerbations of airway diseases, but the role of CO2/HCO3- -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 CO2/HCO3--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 CO2/HCO3 --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, CO2/HCO3 - increases indeed reversibly stimulate intracellular cAMP production by sAC in intact cells.
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U2 - 10.1085/jgp.200709784
DO - 10.1085/jgp.200709784
M3 - Article
C2 - 17591988
AN - SCOPUS:34347273088
VL - 130
SP - 99
EP - 109
JO - Journal of General Physiology
JF - Journal of General Physiology
SN - 0022-1295
IS - 1
ER -