Novel Role for Pendrin in Orchestrating Bicarbonate Secretion in Cystic Fibrosis Transmembrane Conductance Regulator (CFTR)-expressing Airway Serous Cells

Rachel Burrows,Alan W. Cuthbert,Michael A. Gray,Emma Hickman,Péter Hegyi,Peying Fong,James P. Garnett,László Tiszlavicz

doi:10.1074/jbc.m111.266734

Abstract

In most HCO(3)(-)-secreting epithelial tissues, SLC26 Cl(-)/HCO(3)(-) transporters work in concert with the cystic fibrosis transmembrane conductance regulator (CFTR) to regulate the magnitude and composition of the secreted fluid, a process that is vital for normal tissue function. By contrast, CFTR is regarded as the only exit pathway for HCO(3)(-) in the airways. Here we show that Cl(-)/HCO(3)(-) anion exchange makes a major contribution to transcellular HCO(3)(-) transport in airway serous cells. Real-time measurement of intracellular pH from polarized cultures of human Calu-3 cells demonstrated cAMP/PKA-activated Cl(-)-dependent HCO(3)(-) transport across the luminal membrane via CFTR-dependent coupled Cl(-)/HCO(3)(-) anion exchange. The pharmacological and functional profile of the luminal anion exchanger was consistent with SLC26A4 (pendrin), which was shown to be expressed by quantitative RT-PCR, Western blot, and immunofluorescence. Pendrin-mediated anion exchange activity was confirmed by shRNA pendrin knockdown (KD), which markedly reduced cAMP-activated Cl(-)/HCO(3)(-) exchange. To establish the relative roles of CFTR and pendrin in net HCO(3)(-) secretion, transepithelial liquid secretion rate and liquid pH were measured in wild type, pendrin KD, and CFTR KD cells. cAMP/PKA increased the rate and pH of the secreted fluid. Inhibiting CFTR reduced the rate of liquid secretion but not the pH, whereas decreasing pendrin activity lowered pH with little effect on volume. These results establish that CFTR predominately controls the rate of liquid secretion, whereas pendrin regulates the composition of the secreted fluid and identifies a critical role for this anion exchanger in transcellular HCO(3)(-) secretion in airway serous cells.

Highlights

Impede bacterial killing by phagocytic cells [10, 11]
Our results show for the first time that human airway serous cells possess a luminal cAMP/PKA-activated ClϪ/HCO3Ϫ exchanger that exhibits functional properties consistent with those of SLC26A4
Our results agree with data from Xenopus oocytes expressing human SLC26A4, which show that pendrin can function as a ClϪ/HCO3Ϫ exchanger as well a ClϪ/IϪ exchanger [39]

Summary

Introduction

Impede bacterial killing by phagocytic cells [10, 11]. Collectively, these consequences of inadequate HCO3Ϫ transport predispose the lungs to mucus blockage, bacterial infection, and disease, all hallmarks of the CF lung. RNA analysis has shown that human lungs express SLC26A9 [26] which appears to act as a constitutively active CFTR-regulated ClϪ channel in cultured human bronchial epithelial cells [27]. To date, it is unknown whether SLC26 proteins are involved in HCO3Ϫ secretion in the airways. Short hairpin RNA (shRNA)-mediated knockdown of pendrin expression significantly reduced ClϪ/HCO3Ϫ exchange activity and markedly lowered the HCO3Ϫ content of the secreted fluid These studies identify for the first time a critical role for pendrin in transcellular HCO3Ϫ secretion by airway serous cells

Objectives

Results

Conclusion