Identification of calbindin D-9k mRNA and its regulation by 1,25-dihydroxyvitamin D3 in Caco-2 cells.
We have identified and quantified specific mRNAs in the human colonic carcinoma cell line Caco-2 by reverse transcriptase-polymerase chain reaction. Initial examination revealed that like rat duodenal mucosa, Caco-2 cells possessed mRNA for the vitamin D receptor. Using primers for human calbindin we found a 237-bp PCR product in Caco-2 cell RNA, but not from rat duodenal RNA. Primers for rat calbindin did not amplify calbindin mRNA in Caco-2 RNA, confirming a high degree of mismatch between rat and human sequences. 1,25(OH)2 vitamin D3 treatment (10 nM) significantly elevated calbindin mRNA levels 50% by 12 h, with maximal levels occurring by 48 h (fivefold elevation). Increasing concentrations of 1,25(OH)2 vitamin D3 (from 15 pM to 100 nM) caused progressive increases in calbindin mRNA levels following 48 h of treatment. Elevated calbindin mRNA levels were associated with enhancement of transcellular calcium transport. Our results are the first demonstration of vitamin D-regulated calbindin mRNA in a human intestinal cell line.
Fleet
J C
JC
Mineral Bioavailability Laboratory, USDA Human Nutrition Research Center on Aging, Tufts University, Boston, Massachusetts 02111.
Wood
R J
RJ
D000818:Animals; D001483:Base Sequence; D064026:Calbindins; D002117:Calcitriol / Q000494:pharmacology*; D002460:Cell Line; D003110:Colonic Neoplasms; D004386:Duodenum; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism*; D007700:Kinetics; D016133:Polymerase Chain Reaction / Q000379:methods; D012333:RNA, Messenger / Q000096:biosynthesis / Q000378:metabolism*; D012334:RNA, Neoplasm / Q000096:biosynthesis / Q000378:metabolism*; D051381:Rats; D064030:S100 Calcium Binding Protein G / Q000096:biosynthesis*; D012689:Sequence Homology, Nucleic Acid; D014407:Tumor Cells, Cultured
D064026:Calbindins; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D064030:S100 Calcium Binding Protein G; D002117:Calcitriol