Lactase and sucrase-isomaltase gene expression during Caco-2 cell differentiation

• Published in: Biochemical journal 1995-06, Vol.308 ( Pt 3) (3), p.769-775
• Main Authors: Van Beers, E H, Al, R H, Rings, E H, Einerhand, A W, Dekker, J, Büller, H A
• Format: Article
• Language: English
• Subjects: beta-Galactosidase - genetics; beta-Galactosidase - metabolism; Cell Count; Cell Differentiation - genetics; Colonic Neoplasms - metabolism; Electrophoresis, Polyacrylamide Gel; Electrophysiology; Gene Expression Regulation, Neoplastic - genetics; Humans; Immunohistochemistry; Lactase; Precipitin Tests; Proteins - metabolism; Ribonucleases - metabolism; RNA - metabolism; RNA Probes - genetics; RNA, Messenger - genetics; RNA, Messenger - metabolism; Sucrase-Isomaltase Complex - genetics; Sucrase-Isomaltase Complex - metabolism; Transcription, Genetic - genetics; Tumor Cells, Cultured
• ISSN: 0264-6021; 1470-8728
• DOI: 10.1042/bj3080769

The Caco-2 cell line is derived from a human colon adenocarcinoma and differentiates in vitro into small-intestinal enterocyte-like cells, expressing the hydrolases lactase and sucrase-isomaltase. We cultured Caco-2 cells on permeable supports from 0 to 37 days after plating to study endogenous lactase and sucrase-isomaltase gene expression in relation to cell differentiation. Profiles of lactase and sucrase-isomaltase mRNA, protein and enzyme activity were analysed on a per-cell basis, using immunocytochemistry, RNase protection assays, metabolic polypeptide labelling and enzyme activity assays. Tight-junction formation was complete 6 days after plating. Immunocytochemistry of Caco-2 cross-sections showed lactase and sucrase-isomaltase predominantly in the microvillar membrane of polarized cells. mRNA, protein and enzyme activity of lactase appeared consecutively, reaching maximum levels 8-11 days after plating. Whereas lactase mRNA and protein biosynthesis showed a sharp decline after peak levels, lactase activity remained high until 37 days after plating. In contrast, mRNA and protein biosynthesis and activity of sucrase-isomaltase peaked successively 11-21 days after plating, and exhibited comparable levels throughout the entire experiment. The following conclusions were reached. (1) In Caco-2 cells, biosynthesis of lactase and sucrase-isomaltase is regulated by the amount of their mRNAs, indicating transcriptional control. (2) Sucrase-isomaltase activity is most probably transcriptionally controlled at all time 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 indicate different mechanisms of transcriptional regulation of these genes.