title: The CFTR associated protein CAP70 interacts with the apical Cl-/HCO3- exchanger DRA in rabbit small intestinal mucosa.
abstract: DRA (down regulated in adenoma) is an intestinal anion exchanger, acting in parallel with NHE3 to facilitate ileal and colonic NaCl absorption. Furthermore it is involved in small intestinal bicarbonate secretion. Because DRA has a PDZ interaction motif, which may influence its properties, we searched for DRA-interacting PDZ adapter proteins in the small intestine. Using an overlay assay with the recombinant DRA C-terminus as a ligand, a 70 kDa protein was labeled, which was restricted to the brush border membrane in rabbit duodenal and ileal mucosa and was not detected in the colon. Destruction of the C-terminal PDZ interaction motif abolished this band, suggesting a specific protein-protein interaction. The 70 kDa protein was identified as CAP70 (CFTR associated protein of 70 kDa) by an anti-CAP70 antibody and by two in vitro binding assays after cloning CAP70 from rabbit duodenum and ileum. The interaction was recapitulated in HEK cells transfected with DRA and PDZK1, the human orthologue of CAP70. Corresponding to the overlay assay, no CAP70 mRNA or protein was detected in the colon. In vitro protein-protein interaction studies revealed specific binding of DRA to the 2nd and 3rd PDZ domain, while CFTR is known to interact with PDZ1, PDZ3, and PDZ4. The composition of macromolecular complexes assembled by CAP70 in the distal small bowel is unknown. Its restricted expression shows that it cannot be involved in NaCl absorption in the proximal colon. We suggest that CAP70 mediates regulatory functions specific to the small intestine.
authors.affiliation: Department of Clinical Chemistry and Laboratory Medicine, University of Mainz, Mainz, Germany.
authors.forename: Heidi
authors.initials: H
authors.lastname: Rossmann
authors.forename: Petra
authors.initials: P
authors.lastname: Jacob
authors.forename: Susannah
authors.initials: S
authors.lastname: Baisch
authors.forename: Racha
authors.initials: R
authors.lastname: Hassoun
authors.forename: Julia
authors.initials: J
authors.lastname: Meier
authors.forename: Dorgam
authors.initials: D
authors.lastname: Natour
authors.forename: Karim
authors.initials: K
authors.lastname: Yahya
authors.forename: Chris
authors.initials: C
authors.lastname: Yun
authors.forename: Jürg
authors.initials: J
authors.lastname: Biber
authors.forename: Karl J
authors.initials: KJ
authors.lastname: Lackner
authors.forename: Walter
authors.initials: W
authors.lastname: Fiehn
authors.forename: Michael
authors.initials: M
authors.lastname: Gregor
authors.forename: Ursula
authors.initials: U
authors.lastname: Seidler
authors.forename: Georg
authors.initials: G
authors.lastname: Lamprecht
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D017920:Antiporters / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D002352:Carrier Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D002460:Cell Line; D027963:Chloride-Bicarbonate Antiporters / Q000378:metabolism*; D003106:Colon / Q000378:metabolism; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000378:metabolism*; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism*; D007421:Intestine, Small / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism; D012333:RNA, Messenger; D011817:Rabbits; D000076988:Sulfate Transporters
chemical_list: D017920:Antiporters; C505032:CFTR protein, human; D002352:Carrier Proteins; D027963:Chloride-Bicarbonate Antiporters; D008565:Membrane Proteins; C484659:PDZK1 protein, human; D010446:Peptide Fragments; D012333:RNA, Messenger; C081011:SLC26A3 protein, human; D000076988:Sulfate Transporters; D019005:Cystic Fibrosis Transmembrane Conductance Regulator