College of Health

Triiodothyronine (T3) activates rat liver S14 gene transcription through T3 receptors (TRbeta) binding distal thyroid hormone response elements located between -2.8 and -2.5 kilobase pairs upstream from the transcription start site. Previous studies suggested that proximal promoter elements located between -220 to -80 base pairs upstream from the 5' end of the S14 gene were involved in hormone activation of the S14 gene. This report identifies an inverted CCAAT box (or Y box) at -104ATTGG-100 as a core cis-regulatory element. Gel shift studies using rat liver nuclear proteins show that at least three CCAAT-binding factors interact with this region as follows: NF-Y and c/EBP-related proteins formed major complexes, whereas NF-1/CTF forms a minor complex in gel shift assay. Mutation of the Y box indicated that loss of NF-Y binding, but not c/EBP or NF-1, correlated closely with a decline in basal activity and a loss of T3-mediated transactivation. Substitution of the S14 Y box in reporter genes with elements binding only NF-Y elevated basal activity and T3-mediated transactivation, whereas substitution with elements binding c/EBP-related proteins or SP1 displayed low basal activity and T3-mediated transactivation. These studies indicate that NF-Y and TRbeta functionally interact to confer T3 control to the S14 gene.