Transcobalamin II expression is regulated by transcription factor(s) binding to a hexameric sequence (TGGTCC) in the promoter region of the gene

Transcobalamin II (TCII) is a plasma protein that transports cobalamin to tissues for cellular uptake by receptor-mediated endocytosis. Human umbilical vein endothelial cells (HUVEC) in culture constitutively express TCII. However, in other cell lines, TCII expression is dependent on high cell density. ECV304, a cell line with some properties of HUVEC, expresses TCII only when seeded at high density. An electrophoretic mobility-shift assay using nuclear extract from such high-density-seeded ECV 304 cells shifted a 24-bp oligonucleotide probe to generate an unique slow moving band that was competed out by unlabeled probe. This unique band was not observed with nuclear extract from low-density-seeded ECV304 cells. A 3′ sequence, 5′-TGGTCC-3′, in the 24-bp oligonucleotide was identified as the binding site for the nuclear protein(s) because this band was not competed out when the hexameric sequence was scrambled to 5′-CTTCTT-3′. Binding of a transcription factor(s) to this hexamer, that is located 121 bp upstream of the transcription start site, appears to be essential for the regulated or constitutive expression of TCII.