A c-erb-A binding site in rat growth hormone gene mediates trans-activation by thyroid hormone

Abstract

The substance 3,5,3-triiodothyronine (T₃) stimulates growth hormone gene transcription in rat pituitary tumour cells^1–4. This stimulation is thought to be mediated by the binding of nuclear T₃ receptors to regulatory elements 5' to the transcriptional start site^5–8. Understanding of the mechanism by which thyroid hormone activates gene transcription has been limited by failure to purify nuclear T₃ receptors because of their low abundance, and by the absence of defined T₃ receptor-DNA binding sites affecting T₃ regulation. Recently, human and avian c-erb-A gene products have been shown to bind thyroid hormone with high affinity^9,10 and to have a molecular weight and nuclear association characteristic of the thyroid hormone receptor. In the present report, we describe the development of an avidin–biotin complex DNA-binding assay which can detect specific, high-affinity binding of rat pituitary cell T₃ receptors to the sequence 5'CAGGGACGTGACCGCA3', located 164 base pairs 5' to the transcriptional start site of the rat growth hormone gene. An oligonucleotide containing this sequence transferred T₃ regulation to the herpes simplex virus thymidine kinase promoter in transfected rat pituitary GC2 cells, and specifically bound an in vitro translation product of the human placental c-erb-A gene. The data provide supporting evidence that the human c-erb-A gene product mediates the transcriptional effects of T₃ and also that GC2 cell nuclear extracts contain additional factors that modify the binding of pituitary T₃ receptors to the rat growth hormone gene T₃ response element.