The rat TSHβ gene contains distinct response elements for regulation by retinoids and thyroid hormone

Abstract We have previously shown that thyroid stimulating hormone- β (TSH β ) mRNA levels are modulated by vitamin A status in vivo and using transient transfection, that suppression of rat TSH β gene promoter activity by all- trans retinoic acid (RA) requires RA receptor (RAR) and retinoid X receptor (RXR). In this paper we have used deletion analysis to delineate the sequences of the rTSH β gene involved in RA regulation, their relationship to the rTSH β gene negative thyroid hormone response elements and the retinoid receptor species that interact with these sequences. Using transient transfection in CV-1 cells, we found that the −204/+9 region of the rat TSH β gene, when fused to a luciferase reporter, was sufficient for suppression by all- trans -RA in the presence of RAR/RXR. Thus, regulation by RA did not involve the major rTSH β negative TRE located between +15 and +43. Mutational analysis also showed that the minor rTSH β negative TRE between −11 and +5 was not required by suppression by RA. However, in a heterologous promoter this sequence element acted as a strong positive RARE. The combination of RA and T3 treatment caused synergistic inhibition of rat TSH β gene expression in the presence of RAR/RXR and TR. EMSA analysis demonstrated that the −204/−79 sequence binds RAR/RXR heterodimer. Therefore, we conclude that there are separate response elements for RA and T3 on the rat TSH β gene, that the RARE binds RAR/RXR heterodimer and that RA and T3 interact functionally via these elements in the negative regulation of rat TSH β gene expression.