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

• Published in: American journal of physiology. Lung cellular and molecular physiology 2001-04, Vol.280 (4), p.739-L747
• Main Authors: Liedtke, Carole M, Cody, Derek, Cole, Thomas S
• Format: Article
• Language: English
• Subjects: Carrier Proteins - genetics; Carrier Proteins - metabolism; Carrier Proteins - physiology; Chlorides - metabolism; Cystic Fibrosis Transmembrane Conductance Regulator - genetics; Cystic Fibrosis Transmembrane Conductance Regulator - physiology; Humans; Oligonucleotides - pharmacology; Protein Kinase C - physiology; RNA, Messenger - metabolism; Sodium-Potassium-Chloride Symporters; Tumor Cells, Cultured
• ISSN: 1040-0605; 1522-1504
• DOI: 10.1152/ajplung.2001.280.4.l739

The Cystic Fibrosis Center, Departments of Pediatrics and Physiology and Biophysics, Case Western Reserve University, Cleveland, Ohio 44106-4948 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- )] or of the Na/K/2Cl (NKCC1) cotransporter (PKC- ). 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- decreased the amount of PKC- and reduced cAMP-dependent activation of CFTR but not 1 -adrenergic activation of NKCC1. The antisense oligonucleotide to PKC- did not affect CFTR function but did block 1 -adrenergic activation of NKCC1 and reduce PKC- 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. sodium-potassium-2chloride cotransport; antisense oligonucleotide; permeabilized monolayer; reverse transcriptase-polymerase chain reaction; actin; -adrenergic activation; methoxamine; phorbol ester; cystic fibrosis transmembrane conductance regulator; protein kinase C