LACTASE AND SUCRASE-ISOMALTASE GENE-EXPRESSION DURING CACO-2 CELL-DIFFERENTIATION

The Caco-2 cell line is derived from a human colon adenocarcinoma and differentiates in vitro into small-intestinal enterocyte-like cells, e xpressing the hydrolases lactase and sucrase-isomaltase. We cultured C aco-2 cells on permeable supports from 0 to 37 days after plating to s tudy endogenous lactase and sucrase-isomaltase gene expression in rela tion to cell differentiation. Profiles of lactase and sucrase-isomalta se mRNA, protein and enzyme activity were analysed on a per-cell basis , using immunocytochemistry, RNase protection assays, metabolic polype ptide labelling and enzyme activity assays. Tight-junction formation w as complete 6 days after plating. Immuno-cytochemistry of Caco-2 cross -sections showed lactase and sucrase-isomaltase predominantly in the m icrovillar membrane of polarized cells. mRNA, protein and enzyme activ ity of lactase appeared consecutively, reaching maximum levels 8-11 da ys after plating. Whereas lactase mRNA and protein biosynthesis showed a sharp decline after peak levels, lactase activity remained high unt il 37 days after plating. In contrast, mRNA and protein biosynthesis a nd activity of sucrase-isomaltase peaked successively 11-21 days after plating, and exhibited comparable levels throughout the entire experi ment. The following conclusions were reached. (1) In Caco-2 cells, bio synthesis of lactase and sucrase-isomaltase is regulated by the amount of their mRNAs, indicating transcriptional control. (2) Sucrase-isoma ltase activity is most probably transcriptionally controlled at all ti me points. (3) In contrast, lactase activity is initially regulated by its level of biosynthesis. After its peak at 8 days, the slow decline in activity compared with its biosynthesis indicates high stability. (4) Different mRNA profiles for lactase and sucrase-isomaltase indicat e different mechanisms of transcriptional regulation of these genes.