title: Nonsynonymous single nucleotide polymorphisms of NHE3 differentially decrease NHE3 transporter activity.
abstract: Genetic determinants appear to play a role in susceptibility to chronic diarrhea, but the genetic abnormalities involved have only been identified in a few conditions. The Na⁺/H⁺ exchanger 3 (NHE3) accounts for a large fraction of physiologic intestinal Na⁺ absorption. It is highly regulated through effects on its intracellular COOH-terminal regulatory domain. The impact of genetic variation in the NHE3 gene, such as single nucleotide polymorphisms (SNPs), on transporter activity remains unexplored. From a total of 458 SNPs identified in the entire NHE3 gene, we identified three nonsynonymous mutations (R474Q, V567M, and R799C), which were all in the protein's intracellular COOH-terminal domain. Here we evaluated whether these SNPs affect NHE3 activity by expressing them in a mammalian cell line that is null for all plasma membrane NHEs. These variants significantly reduced basal NHE3 transporter activity through a reduction in intrinsic NHE3 function in variant R474Q, abnormal trafficking in variant V567M, or defects in both intrinsic NHE3 function and trafficking in variant R799C. In addition, variants NHE3 R474Q and R799C failed to respond to acute dexamethasone stimulation, suggesting cells with these mutant proteins might be defective in NHE3 function during postprandial stimulation and perhaps under stressful conditions. Finally, variant R474Q was shown to exhibit an aberrant interaction with calcineurin B homologous protein (CHP), an NHE3 regulatory protein required for basal NHE3 activity. Taken together, these results demonstrate decreased transport activity in three SNPs of NHE3 and provide mechanistic insight into how these SNPs impact NHE3 function.
authors.affiliation: Department of Medicine, Division of Gastroenterology and Hepatology, Center of Cardiovascular Sciences, Albany Medical Center, Albany, New York; Departments of Physiology and Medicine, Division of Gastroenterology, Johns Hopkins University School of Medicine, Baltimore, Maryland; and.
authors.forename: Xinjun Cindy
authors.initials: XC
authors.lastname: Zhu
authors.affiliation: Departments of Physiology and Medicine, Division of Gastroenterology, Johns Hopkins University School of Medicine, Baltimore, Maryland; and.
authors.forename: Rafiquel
authors.initials: R
authors.lastname: Sarker
authors.affiliation: Department of Biochemistry, Emory University School of Medicine, Atlanta, Georgia.
authors.forename: John R
authors.initials: JR
authors.lastname: Horton
authors.affiliation: Departments of Physiology and Medicine, Division of Gastroenterology, Johns Hopkins University School of Medicine, Baltimore, Maryland; and.
authors.forename: Molee
authors.initials: M
authors.lastname: Chakraborty
authors.affiliation: Departments of Physiology and Medicine, Division of Gastroenterology, Johns Hopkins University School of Medicine, Baltimore, Maryland; and.
authors.forename: Tian-E
authors.initials: TE
authors.lastname: Chen
authors.affiliation: Departments of Physiology and Medicine, Division of Gastroenterology, Johns Hopkins University School of Medicine, Baltimore, Maryland; and.
authors.forename: C Ming
authors.initials: CM
authors.lastname: Tse
authors.affiliation: Departments of Physiology and Medicine, Division of Gastroenterology, Johns Hopkins University School of Medicine, Baltimore, Maryland; and.
authors.forename: Boyoung
authors.initials: B
authors.lastname: Cha
authors.affiliation: Departments of Physiology and Medicine, Division of Gastroenterology, Johns Hopkins University School of Medicine, Baltimore, Maryland; and mdonowit@JHMI.edu.
authors.forename: Mark
authors.initials: M
authors.lastname: Donowitz
mesh_terms: D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D000818:Animals; D001692:Biological Transport; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D003907:Dexamethasone / Q000494:pharmacology; D015536:Down-Regulation; D005838:Genotype; D006801:Humans; D009154:Mutation; D010641:Phenotype; D020641:Polymorphism, Single Nucleotide; D011485:Protein Binding; D021381:Protein Transport; D011817:Rabbits; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism*; D014162:Transfection
chemical_list: C469743:CHP1 protein, human; D002135:Calcium-Binding Proteins; C000613385:SLC9A3 protein, human; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers; D015124:8-Bromo Cyclic Adenosine Monophosphate; D003907:Dexamethasone
keywords: CHP; NHE3; SNP; dexamethasone