Folate uptake in the human intestine: promoter activity and effect of folate deficiency

The intestinal folate absorption process occurs via a specialized mechanism that involves the reduced folate carrier (RFC). In humans, multiple variants of the hRFC (driven by multiple promoters) have been identified with variant I being the prominent form expressed in the intestine. While it is known that promoter B (pB) of hRFC drives the expression of this variant, little is known about the minimal region required for basal activity of this promoter in human intestinal epithelial cells. Also not known is whether folate absorption in the human intestine is up-regulated during folate deficiency (as occur in animal studies), and if so, whether transcriptional mechanisms via activation of hRFC pB are involved in such regulation. To address these issues, we have used deletion constructs of the hRFC pB and determined their activity in two human intestinal epithelial cell lines: the colon-derived Caco-2 cells, and the duodenum-derived HuTu-80 cells. Our results showed that activity of hRFC pB to be significantly higher in Caco-2 cells compared to HuTu-80 cells, a finding that corresponds with a higher level of folate uptake and endogenous hRFC mRNA levels in the former compared to the latter cell type. The minimal region required for basal activity of hRFC pB in Caco-2 cells was found to be encoded in a sequence between -1088 and -1043, while in HuTu-80 cells it was encoded in a sequence between -1431 and -1088. Growing Caco-2 cells in a folate deficient medium led to a significant and specific up-regulation in folate uptake. This up-regulation was associated with a parallel increase in hRFC protein and mRNA levels, and in the activity of hRFC pB. The most responsive sequence of pB to the effect of folate deficiency was found to be encoded in a sequence between -2016 and -1431, i.e., outside the minimal region of the pB. These results show that different minimal regions for hRFC pB are utilized by different intestinal epithelial cells. In addition, folate-deficiency was found to up-regulate folate uptake by human intestinal epithelial cells and that this regulation involves activation of hRFC pB.