(Na+ + K+)-ATPase and plasma membrane polarity of intestinal epithelial cells: presence of a brush border antigen in the distal large intestine that is immunologically related to beta subunit.
The previously produced monoclonal antibody IEC 1/48 against cultured rat intestinal crypt cells (Quaroni, A., and K. J. Isselbacher. 1981. J. Natl. Cancer Inst. 67:1353-1362) was extensively characterized and found to be directed against the beta subunit of (Na+ + K+)-ATPase as assessed by immunological and enzymatic criteria. Under nondenaturing conditions the antibody precipitated the alpha-beta enzyme complex (98,000 and 48,000 Mr). This probe, together with the monoclonal antibody C 62.4 against the alpha subunit (Kashgarian, M., D. Biemesderfer, M. Caplan, and B. Forbush. 1985. Kidney Int. 28:899-913), was used to localize (Na+ + K+)-ATPase in epithelial cells along the rat intestinal tract by immunofluorescence and immunoelectron microscopy. Both antibodies exclusively labeled the basolateral membrane of small intestine and proximal colon epithelial cells. However, in the distal colon, IEC 1/48, but not C 62.4, also labeled the brush border membrane. The cross-reacting beta-subunit-like antigen on the apical cell pole was tightly associated with isolated brush borders but was apparently devoid of (Na+ + K+)-ATPase activity. Subcellular fractionation of colonocytes in conjunction with limited proteolysis and surface radioiodination of intestinal segments suggested that the cross-reacting antigen in the brush border may be very similar to the beta subunit. The results support the notion that in the small intestine and proximal colon the enzyme subunits are exclusively targeted to the basolateral membrane while in the distal colon nonassembled beta subunit or a beta-subunit-like protein is also transported to the apical cell pole.
Marxer
A
A
Department of Pharmacology, University of Basel, Switzerland.
Stieger
B
B
Quaroni
A
A
Kashgarian
M
M
Hauri
H P
HP
D000818:Animals; D000911:Antibodies, Monoclonal; D000954:Antigens, Surface / Q000032:analysis*; D002458:Cell Fractionation / Q000379:methods; D002462:Cell Membrane / Q000201:enzymology / Q000502:physiology / Q000648:ultrastructure*; D003106:Colon / Q000201:enzymology* / Q000648:ultrastructure; D004591:Electrophoresis, Polyacrylamide Gel; D004848:Epithelium / Q000201:enzymology / Q000648:ultrastructure; D005455:Fluorescent Antibody Technique; D005643:Fucose / Q000378:metabolism; D007421:Intestine, Small / Q000201:enzymology / Q000648:ultrastructure; D046911:Macromolecular Substances; D008715:Methionine / Q000378:metabolism; D008854:Microscopy, Electron; D008871:Microvilli / Q000201:enzymology / Q000648:ultrastructure*; D010449:Peptide Mapping; D051381:Rats; D011919:Rats, Inbred Strains; D000254:Sodium-Potassium-Exchanging ATPase / Q000032:analysis*; D013462:Sulfur Radioisotopes; D014316:Tritium
D000911:Antibodies, Monoclonal; D000954:Antigens, Surface; D046911:Macromolecular Substances; D013462:Sulfur Radioisotopes; D014316:Tritium; D005643:Fucose; D008715:Methionine; D000254:Sodium-Potassium-Exchanging ATPase