THE CCAAT BOX-BINDING FACTOR, NF-Y, IS REQUIRED FOR THYROID-HORMONE REGULATION OF RAT-LIVER S14 GENE-TRANSCRIPTION

Triiodothyronine (T-3) activates rat liver S14 gene transcription thro ugh T-3 receptors (TR beta) binding distal thyroid hormone response el ements located between -2.8 and -2.5 kilobase pairs upstream from the transcription start site. Previous studies suggested that proximal pro moter elements located between -220 to -80 base pairs upstream from th e 5' end of the S14 gene were involve din hormone activation of the S1 4 gene. This report identifies an inverted CCAAT box (or Y box) at (-1 04)ATTGG(-100) as a core cis-regulatory element. Gel shift studies usi ng rat liver nuclear proteins show that at least three CCAAT-binding f actors interact with this region as follows: NF-Y and c/EBP-related pr oteins 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 b asal activity and a loss of T-3-mediated transactivation. Substitution of the S14 Y box in reporter genes with elements binding only NF-Y el evated basal activity and T-3-mediated transactivation, whereas substi tution with elements binding c/EBP-related proteins or SP1 displayed l ow basal activity and T-3-mediated transactivation. These studies indi cate that NF-Y and TR beta functionally interact to confer T-3 control to the S14 gene.