Poly-N-acetyllactosamine-specific tomato lectin interacts with gastric parietal cells. Identification of a tomato-lectin binding 60-90 X 10(3) Mr membrane glycoprotein of tubulovesicles.
The cytoplasmic tubulovesicular and canalicular membranes of gastric parietal cells are intimately involved in hydrochloric acid secretion. To characterise the glycoproteins of these membranes, we examined a panel of lectins for reactivity with parietal cells in paraffin sections of rat, dog and pig stomach. The poly-N-acetyllactosamine-specific lectin from Lycopersicon esculentum (tomato) and from Solanum tuberosum (potato), and the galactose-specific lectin Ricinus communis agglutinin (RCA120), showed strong cytoplasmic binding of parietal cells of all three species, with a pattern indicative of an intracellular membrane network. Binding to parietal cells was confirmed by double-labelling studies with parietal cell auto-antibodies from patients with autoimmune gastritis. Mucous cells and mucin also bound these lectins strongly. Other gastric cell types did not stain with either tomato or potato lectin, but stained weakly with RCA120. Electron-microscopic examination of lectin binding sites using biotinylated tomato lectin or RCA120 and streptavidin-gold, revealed specific binding to the luminal face of parietal cell tubulovesicular and canalicular membranes as well as the contents of mucous cell secretory granules. Tomato lectin and RCA120 reacted by lectin blotting with a major species of apparent molecular weight 60-90 X 10(3) Mr from rat, dog and pig gastric membranes. A tubulovesicular membrane fraction, enriched 10-fold for K(+)-dependent phosphatase activity, was also enriched three-fold for tomato lectin binding as assessed by a solid-phase lectin assay. The 60-90K (K = 10(3) Mr) component, in 125I-labelled detergent extracts of dog tubulovesicular membranes, bound to an affinity support of tomato lectin-Sepharose and was specifically eluted with N,N',N'-triacetylchitotriose. Digestion with N-glycanase collapsed the 60-90K component into a sharp 35K band. We conclude that: (1) a 60-90K membrane glycoprotein localised on the luminal face of tubulovesicles and canaliculi of parietal cells interacts strongly with tomato lectin and RCA120; and (2) the glycoprotein is composed of a 35K core protein glycosylated with N-glycans probably containing poly-N-acetyllactosamine sequences with terminal galactosyl residues. The properties of this 60-90K glycoprotein are identical to a major parietal cell autoantigen recognised by sera of patients with autoimmune gastritis.(ABSTRACT TRUNCATED AT 400 WORDS)
Callaghan
J M
JM
Department of Pathology and Immunology, Monash University Medical School, Melbourne, Victoria, Australia.
Toh
B H
BH
Pettitt
J M
JM
Humphris
D C
DC
Gleeson
P A
PA
D009597:4-Nitrophenylphosphatase / Q000378:metabolism; D000818:Animals; D001324:Autoantigens / Q000276:immunology; D002240:Carbohydrate Sequence; D002846:Chromatography, Affinity; D004285:Dogs; D004591:Electrophoresis, Polyacrylamide Gel; D005756:Gastritis / Q000276:immunology / Q000378:metabolism; D007124:Immunoenzyme Techniques; D037102:Lectins / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000302:isolation & purification* / Q000378:metabolism; D008969:Molecular Sequence Data; D009092:Mucous Membrane / Q000378:metabolism; D010295:Parietal Cells, Gastric / Q000201:enzymology / Q000378:metabolism* / Q000648:ultrastructure; D037121:Plant Lectins; D011134:Polysaccharides / Q000378:metabolism*; D006180:Proton-Translocating ATPases / Q000378:metabolism; D051381:Rats; D013552:Swine
D001324:Autoantigens; D037102:Lectins; D008562:Membrane Glycoproteins; D037121:Plant Lectins; D011134:Polysaccharides; C034981:tomato lectin; C037199:poly-N-acetyllactosamine; D009597:4-Nitrophenylphosphatase; D006180:Proton-Translocating ATPases