Differential regulation of Cl− transport proteins by PKC in Calu-3 cells

Abstract

Cl− transport proteins expressed in a Calu-3 airway epithelial cell line were differentiated by function and regulation by protein kinase C (PKC) isotypes. mRNA expression of Cl− transporters was semiquantitated by RT-PCR after transfection with a sense or antisense oligonucleotide to the PKC isotypes that modulate the activity of the cystic fibrosis transmembrane conductance regulator [CFTR (PKC-ε)] or of the Na/K/2Cl (NKCC1) cotransporter (PKC-δ). Expression of NKCC1 and CFTR mRNAs and proteins was independent of antisense oligonucleotide treatment. Transport function was measured in cell monolayers grown on a plastic surface or on filter inserts. With both culture methods, the antisense oligonucleotide to PKC-ε decreased the amount of PKC-ε and reduced cAMP-dependent activation of CFTR but not α1-adrenergic activation of NKCC1. The antisense oligonucleotide to PKC-δ did not affect CFTR function but did block α1-adrenergic activation of NKCC1 and reduce PKC-δ mass. These results provide the first evidence for mRNA and protein expression of NKCC1 in Calu-3 cells and establish the differential regulation of CFTR and NKCC1 function by specific PKC isotypes at a site distal to mRNA expression and translation in airway epithelial cells.