title: Congenital chloride-losing diarrhea causing mutations in the STAS domain result in misfolding and mistrafficking of SLC26A3.
abstract: Congenital chloride-losing diarrhea (CLD) is a genetic disorder causing watery stool and dehydration. Mutations in SLC26A3 (solute carrier 26 family member 3), which functions as a coupled Cl(-)/HCO(3)(-) exchanger, cause CLD. SLC26A3 is a membrane protein predicted to contain 12 transmembrane-spanning alpha-helices and a C-terminal STAS (sulfate transporters and anti-sigma-factor) domain homologous to the bacterial anti-sigma-factor antagonists. The STAS domain is required for SLC26A3 Cl(-)/HCO(3)(-) exchange function and for the activation of cystic fibrosis transmembrane conductance regulator by SLC26A3. Here we investigate the molecular mechanism(s) by which four CLD-causing mutations (DeltaY526/7, I544N, I675/6ins, and G702Tins) in the STAS domain lead to disease. In a heterologous mammalian expression system biochemical, immunohistochemical, and ion transport experiments suggest that the four CLD mutations cause SLC26A3 transporter misfolding and/or mistrafficking. Expression studies with the isolated STAS domain suggest that the I675/6ins and G702Tins mutations disrupt the STAS domain directly, whereas limited proteolysis experiments suggest that the DeltaY526/7 and I544N mutations affect a later step in the folding and/or trafficking pathway. The data suggest that these CLD-causing mutations cause disease by at least two distinct molecular mechanisms, both ultimately leading to loss of functional protein at the plasma membrane.
authors.affiliation: Department of Physiology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
authors.forename: Michael R
authors.initials: MR
authors.lastname: Dorwart
authors.forename: Nikolay
authors.initials: N
authors.lastname: Shcheynikov
authors.forename: Jennifer M R
authors.initials: JM
authors.lastname: Baker
authors.forename: Julie D
authors.initials: JD
authors.lastname: Forman-Kay
authors.forename: Shmuel
authors.initials: S
authors.lastname: Muallem
authors.forename: Philip J
authors.initials: PJ
authors.lastname: Thomas
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D017920:Antiporters / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D027963:Chloride-Bicarbonate Antiporters; D002712:Chlorides / Q000378:metabolism*; D002522:Chlorocebus aethiops; D002942:Circular Dichroism; D017124:Conserved Sequence; D003967:Diarrhea / Q000151:congenital* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D021381:Protein Transport; D016415:Sequence Alignment; D000076988:Sulfate Transporters; D013696:Temperature
chemical_list: D017920:Antiporters; D027963:Chloride-Bicarbonate Antiporters; D002712:Chlorides; C081011:SLC26A3 protein, human; D000076988:Sulfate Transporters