Rescue of epithelial HCO3− secretion in murine intestine by apical membrane expression of the cystic fibrosis transmembrane conductance regulator mutant F508del
Authors: Xiao Fang , Li Junhua , Singh Anurag Kumar , Riederer Brigitte , Wang Jiang , Sultan Ayesha , Park Henry , Lee Min Goo , Lamprecht Georg , Scholte Bob J. , De Jonge Hugo R. , Seidler Ursula 
Published: July 2012

Key points

Cystic fibrosis (CF) is a lethal disease characterized by low rates of epithelial Cl− and HCO3− secretion and obstruction of the airways and gastrointestinal and reproductive organs by sticky mucus. HCO3− secretion has recently been demonstrated to be necessary for mucus hydration.

The most frequent CF mutation is F508del. This mutant protein is usually degraded in the proteasome. New therapeutic strategies have been developed which deliver F508del to the plasma membrane.

Utilizing transgenic F508del mutant and cystic fibrosis transmembrane conductance regulator (CFTR) knockout mice, apical membrane expression of F508del protein was found to be associated with enhanced stimulation of intestinal HCO3− secretion.

The predominant molecular mechanism for enhanced F508del HCO3− stimulation appeared to be the activation of a Cl− recycling pathway, with Cl− exit via membrane‐resident F508del protein and Cl− entry in exchange for HCO3− by apical Cl−/HCO3− exchange. In contrast, the predominant molecular mechanism for cAMP‐activated HCO3− secretion in WT intestine appears to be HCO3− exit via CFTR itself.

Abstract  This study investigated whether expression of the common cystic fibrosis transmembrane conductance regulator (CFTR) mutant F508del in the apical membrane of enterocytes confers increased bicarbonate secretory capacity on the intestinal epithelium of F508del mutant mice compared to that of CFTR knockout (KO) mice. CFTR KO mice, F508del mutant mice (F508del) and wild‐type (WT) littermates were bred on the FVB/N background. F508del isolated brush border membrane (BBM) contained approximately 5–10% fully glycosylated band C protein compared to WT BBM. Similarly, the forskolin (FSK)‐induced, CFTR‐dependent short‐circuit current (ΔIsc) of F508del mucosa was approximately 5–10% of WT, whereas the HCO3− secretory response () was almost half that of WT in both duodenum and mid‐colon studied in vitro and in vivo. While WT intestine retained full FSK‐induced in the absence of luminal Cl−, the markedly higher than ΔIsc in F508del intestine was dependent on the presence of luminal Cl−, and was blocked by CFTR inhibitors. The Ste20‐related proline–alanine‐rich kinases (SPAK/OSR1), which are downstream of the with‐no‐lysine (K) protein kinases (WNK), were rapidly phosphorylated by FSK in WT and F508del, but significantly more slowly in CFTR KO intestine. In conclusion, the data demonstrate that low levels of F508del membrane expression in the intestine of F508del mice significantly increased FSK‐induced HCO3− secretion mediated by Cl−/HCO3− exchange. However, in WT mucosa FSK elicited strong SPAK/OSR1 phosphorylation and Cl−‐independent HCO3− efflux. This suggests that therapeutic strategies which deliver F508del to the apical membrane have the potential to significantly enhance epithelial HCO3− secretion.