The long-term objective of this renewal application is to continue our investigations into the cellular and molecular mechanisms involved in the absorption of the water-soluble vitamin B1 (thiamin) in the human intestine and their regulation. Thiamin is essential for normal cellular functions and its deficiency (which represents a significant nutritional problem) leads to a variety of clinical abnormalities including cardiovascular and neurological disorders. Humans (and other mammals) cannot synthesize thiamin, and thus, must obtain the vitamin via intestinal absorption. Studies from our laboratory during the current funding period have characterized many aspects of the human intestinal thiamin uptake process including the demonstration that both of the thiamin transporters hTHTR-1 and hTHTR-2 are expressed in the human intestine and play a role in thiamin uptake. These studies have also shown that the hTHTR-1 is expressed at both the apical and basolateral membrane domains of intestinal epithelial cells, while the hTHTR-2 is mainly expressed at the apical membrane domain. In addition, we have cloned and performed initial characterization of the promoters of the genes that encode these transporters (SLC19A2 and SLC19A3, respectively). In new preliminary studies, we found the intestinal thiamin uptake process to be regulated by ontogeny and by dietary thiamin level, and that this regulation appears to involve transcriptional mechanism(s). In other preliminary studies, we have identified two sequences in the hTHTR-1 polypeptide that appear to be important for its targeting to intestinal cell membrane.
Our specific aims i n this application are: 1) To continue the characterization of the SLC19A2 and SLC19A3 promoters in vitro and in vivo in transgenic mice, 2) To determine the cellular and molecular mechanisms involved in the regulation of the intestinal thiamin uptake process by ontogeny and by dietary substrate levels, and 3) To study the mechanisms involved in membrane targeting and intracellular trafficking of the hTHTR-1 and the hTHTR-2 in human intestinal epithelial cells. Results of these studies should continue to provide novel and valuable information regarding the cellular and molecular mechanisms involved in thiamin uptake by the human intestine and their regulation. This should ultimately assist us in the designing of effective strategies to optimize thiamin body homeostasis in conditions associated with thiamin deficiency and sub-optimal levels.
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