Differentiation-dependent up-regulation of intestinal thiamin uptake: cellular and molecular mechanisms

Differentiation of intestinal epithelial cells is associated with up-and-down regulation of expression of a variety of genes including those involved in nutrient uptake. Nothing is known about possible differentiation-dependent regulation of the intestinal thiamin uptake process and the cellular and molecular mechanisms involved in such regulation. Using as models human-derived intestinal epithelial Caco-2 cells and crypt/villus epithelial cells isolated from wild-type and transgenic mice carrying promoters for human thiamin transporter-1 and -2 (hTHTR-1 and hTHTR-2), we addressed this issue. Our results showed that differentiation of Caco-2 cells is associated with a significant up-regulation in carrier-mediated thiamin uptake. Up-regulation was associated with a significant increase in the level of expression of hTHTR-1 and hTHTR-2 protein and mRNA as well as in activity of the corresponding transfected human thiamin transporter-1 (SLC19A2) and -2 (SLC19A3) promoters. Deletion analysis identified the differentiation-responsive region to be at position -356 to -275 bp for the SLC19A2 promoter and at position -77 to -13 bp for the SLC19A3 promoter. In addition, a critical and specific role in the differentiation-mediated effects for an NF1 binding site (-348 to -345 bp) in the SLC19A2 promoter and a SP1/GC-box binding site (-48 to -45 bp) in the SLC19A3 promoter were established using mutational analysis. The physiological relevance of in vitro findings with Caco-2 cells was confirmed in wild-type and transgenic mice by demonstrating that thiamin uptake and mRNA levels of the mouse THTR-1 and THTR-2, as well as activity of human SLC19A2 and SLC19A3 promoters expressed in transgenic mice, were all significantly higher in intestinal villus compared with crypt epithelial cells. These studies demonstrate for the first time that differentiation of intestinal epithelial cells is associated with an up-regulation in thiamin uptake process and that this up-regulation appears to be mediated via transcriptional regulatory mechanisms that involve the SLC19A2 and SLC19A3 genes.