Bicarbonate Secretion, Cystic Fibrosis and Congenital Chloride Diarrhea: Molecular Mechanisms in Ion Transport and Disease

Cystic Fibrosis (CF) and Congenital Chloride Diarrhea (CLD) are two genetic diseases which without treatment can be fatal. Cystic Fibrosis is a disease which disrupts normal fluid secretion in multiple organs and tissues, and is caused by mutations in the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) gene. Congenital Chloride Diarrhea is a disease caused by mutations in the SLC26A3 gene, which cause patients to suffer from watery stool and dehydration. The proteins encoded by the CFTR and SLC26A3 genes have been demonstrated to be a chloride channel and a chloride-bicarbonate exchanger respectively. They have also been shown to reciprocally activate one another's transport activity. Understanding how these proteins transport chloride and bicarbonate across the apical membrane of epithelial cells, and how they mutually activate one another's transport activity is critical for our understanding of CF and CLD. The SLC26A3 protein is a membrane protein predicted to contain 12 transmembrane spanning alpha -helices, and a C-terminal STAS domain which is homologous to the bacterial anti-sigma factor antagonists. The STAS domain is required for proper SLC26A3 chloride-bicarbonate exchange function, and it is also required for the reciprocal activation of SLC26A3 and CFTR activities. Here we investigate the molecular interaction between the STAS domain of SLC26A3 and the R-domain of CFTR, as well as the molecular mechanism(s) by which four CLD causing mutations (delta Y526/7, I544N, I675/6ins and G702Tins) residing in the SLC26A3 STAS domain lead to disease. The STAS domain and R-domain have been demonstrated to directly bind to one another, and this interaction is modulated by phosphorylation of the R-domain. Functional, biochemical and cell biological experiments performed on wild type and mutant SLC26A3 proteins suggest that the CLD mutations cause transporter misfolding and/or mistrafficking. Biochemical and biophysical studies performed on the purified STAS domains suggest that these CLD causing mutations have differential effects on the STAS domain's structure. Our data taken together suggest that the CLD causing mutations cause disease by at least two distinct molecular mechanisms, ultimately leading to loss of functional protein at the plasma membrane.