Evidence for a membrane carbonic anhydrase IV anchored by its C-terminal peptide in normal human pancreatic ductal cells.
The high concentration of HCO(3)(-) ions (150 mM) in the human pancreatic ducts raises the question of the membrane proteins responsible for their secretion in addition to the Cl(-)/HCO(3)(-) exchanger. In this study, we investigated the expression of carbonic anhydrase IV (CA IV), a possible candidate. Experiments were carried out on specimens of normal human pancreas obtained from brain-dead donors ( n=9) as well as on isolated human ductal cells. Two antibodies were generated: CA IV NH(2) antibody directed against the NH(2) terminal of human glycosyl phosphatidylinositol (GPI)-anchored CA IV and CA IV COOH antibody directed against the COOH terminal of the same protein before its association with a GPI in the rough endoplasmic reticulum. A 35-kDa CA IV was detected in the homogenates of human pancreas. Immunocytochemistry demonstrated the expression of CA IV in centroacinar cells and in intercalated, intralobular, and interlobular ductal cells. The immunoreactivity observed with the CA IV COOH antibody was mainly localized on luminal membranes of ductal cells. Treatment of purified plasma membranes with phosphatidylinositol-phospholipase C indicated that the CA IV expressed in pancreatic ducts was not GPI-anchored. Its detection in the same extracts by the CA IV COOH antibody indicated that it was anchored by a hydrophobic segment at the carboxy terminal. Taken together, these results suggest that normal human pancreatic ductal cells express a 35-kDa CA IV anchored in their luminal plasma membrane by a hydrophobic segment of the COOH terminus. In view of its localization and its mode of anchorage in luminal plasma membranes, this CA IV may participate in the maintenance of luminal pH.
Fanjul
Marjorie
M
Laboratoire de Biologie Cellulaire et Moléculaire des Epithéliums (EA 3032), Université Paul Sabatier, 38 rue des 36 Ponts, 31400 Toulouse, France.
Alvarez
Laetitia
L
Salvador
Christel
C
Gmyr
Valéry
V
Kerr-Conte
Julie
J
Pattou
François
F
Carter
Nicholas
N
Hollande
Etienne
E
D001639:Bicarbonates / Q000378:metabolism; D030741:Carbonic Anhydrase IV / Q000737:chemistry / Q000378:metabolism*; D002462:Cell Membrane / Q000201:enzymology; D002469:Cell Separation; D002478:Cells, Cultured; D006801:Humans; D007124:Immunoenzyme Techniques; D010183:Pancreatic Ducts / Q000166:cytology / Q000201:enzymology*; D010446:Peptide Fragments / Q000032:analysis / Q000378:metabolism; D017434:Protein Structure, Tertiary
D001639:Bicarbonates; D010446:Peptide Fragments; D030741:Carbonic Anhydrase IV