Intestinal anion exchanger down-regulated in adenoma (DRA) is inhibited by intracellular calcium.
The Na/H exchanger 3 (NHE3) and the Cl/HCO(3) exchanger down-regulated in adenoma (DRA) together facilitate intestinal electroneutral NaCl absorption. Elevated Ca(2+)(i) inhibits NHE3 through mechanisms involving the PDZ domain proteins NHE3 kinase A regulatory protein (E3KARP) or PDZ kidney 1 (PDZK1). DRA also possesses a PDZ-binding motif, but the roles of interactions with E3KARP or PDZK1 and Ca(2+)(i) in DRA regulation are unknown. Wild type DRA and a mutant lacking the PDZ interaction motif (DRA-ETKFminus) were expressed constitutively in human embryonic kidney (HEK) and inducibly in Caco-2/BBE cells. DRA-mediated Cl/HCO(3) exchange was measured as intracellular pH changes. Ca(2+)(i) was assessed fluorometrically. DRA was induced 8-16-fold and was delivered to the apical surface of polarized Caco-2 cells. Putative anion transporter 1 and cystic fibrosis transmembrane regulator did not contribute to Cl/HCO(3) exchange in transfected Caco-2 cells. The calcium ionophore 4Br-A23187 inhibited DRA and DRA-ETKFminus in HEK cells, but only full-length DRA was inhibited in Caco-2 cells. In contrast, 100 microm UTP, which increased Ca(2+)(i), inhibited full-length DRA but not DRA-ETKFminus in Caco-2 and HEK cells. In HEK cells, which express little PDZK1, additional transfection of PDZK1 was required for UTP to inhibit DRA. As HEK cells do not express cystic fibrosis transmembrane regulator or NHE3, the data indicate that Ca(2+)(i)-dependent DRA inhibition is not because of modulation of other transport activities. In polarized epithelium, this inhibition requires interaction of DRA with PDZK1. Together with data from PDZK1(-/-) mice, these data underscore the prominent role of PDZK1 in Ca(2+)(i)-mediated inhibition of colonic NaCl absorption.
Lamprecht
Georg
G
1st Medical Department, University of Tübingen, 72076 Tübingen, Germany. hans-georg.lamprecht@uni-tuebingen.de
Hsieh
Chih-Jen
CJ
Lissner
Simone
S
Nold
Lilia
L
Heil
Andreas
A
Gaco
Veronika
V
Schäfer
Julia
J
Turner
Jerrold R
JR
Gregor
Michael
M
D017920:Antiporters / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites / Q000235:genetics; D018938:Caco-2 Cells; D000001:Calcimycin / Q000031:analogs & derivatives / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D027963:Chloride-Bicarbonate Antiporters; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D015151:Immunoblotting; D007413:Intestinal Mucosa / Q000378:metabolism; D042541:Intracellular Space / Q000378:metabolism; D007476:Ionophores / Q000494:pharmacology; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D054731:PDZ Domains / Q000235:genetics; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers / Q000235:genetics / Q000378:metabolism; D000076988:Sulfate Transporters; D014162:Transfection; D014544:Uridine Triphosphate / Q000494:pharmacology
D017920:Antiporters; C505032:CFTR protein, human; D027963:Chloride-Bicarbonate Antiporters; D007476:Ionophores; D008565:Membrane Proteins; C509768:PDZK1IP1 protein, human; D010750:Phosphoproteins; D011993:Recombinant Fusion Proteins; C081011:SLC26A3 protein, human; C000613385:SLC9A3 protein, human; C000613386:Slc9a3 protein, mouse; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers; D000076988:Sulfate Transporters; C093376:sodium-hydrogen exchanger regulatory factor; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D049452:Green Fluorescent Proteins; D000001:Calcimycin; C033170:4-bromo-A-23187; D002118:Calcium; D014544:Uridine Triphosphate