Use of monoclonal antibodies to investigate the intracellular transport and biogenesis of intestinal brush-border proteins.

The differentiated enterocyte of the mammalian small intestine is an excellent model to study the polarized expression of surface membrane proteins in epithelial cells. A number of digestive hydrolases, most of which are major integral membrane proteins, are specifically expressed in the brush-border membrane of these cells and they can serve as molecular markers for the apical membrane domain (Semenza, 1985). Previous studies on the biogenesis of the brush-border membrane have been carried out with subcellular fractions of either scraped mucosa, isolated villus cells or organ-cultured mucosa, and have given important clues on precursor forms and the post-translational processing of these hydrolases (Hauri, 1983, Quaroni, 1984, Danielsen t't al., 1984). I t would be highly desirable to have an intestinal cell culture system in which biosynthetic processes can be manipulated experimentally in a much easier way than in intact tissues. Although the establishment of permanent intestinal cell cultures has been achieved it has become apparent that these cells are of crypt origin and it has not been possible to allow them to differentiate into villus cells in culture (Quaroni et ul., 1979). Recently, a highly differentiated colonic-adenocarcinoma cell line, Caco 2 , was described. which expresses small-intestine brushborder enzyme activities (Pinto ~t ul., 1983). We have prepared a panel of monoclonal antibodies against human intestinal brush-border enzymes and have used these probes to study the transport of four hydrolases to the luminal cell surface in Caco 2 cells.