Stimulation of β2-adrenergic receptor increases cystic fibrosis transmembrane conductance regulator expression in human airway epithelial cells through a cAMP/protein kinase A-independent pathway

Abstract

PSD-95/Dlg-A/ZO-1 (PDZ) domains play an essential role in determining cell polarity. The Na+/H+ exchanger regulatory factor (NHERF), also known as EBP50, contains two PDZ domains that mediate the assembly of transmembrane and cytosolic proteins into functional signal transduction complexes. Moreover, it has been shown that cystic fibrosis transmembrane conductance regulator (CFTR) and β2-adrenergic receptor (β2AR) bind equally well to the PDZ1 domain of EBP50. We hypothesized that β2AR activation may regulate CFTR protein expression. To verify this, we evaluated the effects of a pharmacologically relevant concentration of salmeterol (2.10-7 M), a long acting β2AR agonist, on CFTR expression in primary human airway epithelial cells (HAEC). β2AR stimulation induced a time-dependent increase in apical CFTR protein expression, with a maximal response reached after treatment for 24 h. This effect was post-transcriptional, dependent upon the β2AR agonist binding to β2AR and independent of the known β2AR agonist-mediated cAMP/PKA pathway. We demonstrated by immunohistochemistry that CFTR, β2AR, and EBP50 localize to the apical membrane of HAEC. Analyses of anti-EBP50 protein immunoprecipitate showed that salmeterol induced an increase in the amount of CFTR that binds to EBP50. These data suggest that β2AR activation regulates the association of CFTR with EBP50 in polarized HAEC.