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Volume 272, Number 44, Issue of October 31, 1997 pp. 27778-27786
©1997 by The American Society for Biochemistry and Molecular Biology, Inc.

The CCAAT Box Binding Factor, NF-Y, Is Required for Thyroid Hormone Regulation of Rat Liver S14 Gene Transcription

(Received for publication, June 23, 1997, and in revised form, August 6, 1997)

From the Departments of Physiology and Biochemistry, Michigan State University, East Lansing, Michigan 48824

Triiodothyronine (T₃) activates rat liver S14 gene transcription through T₃ receptors (TR) 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 ¹⁰⁴ATTGG¹⁰⁰ 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 TR functionally interact to confer T₃ control to the S14 gene.

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