The rat TSHbeta gene contains distinct response elements for regulation by retinoids and thyroid hormone.
We have previously shown that thyroid stimulating hormone-beta (TSHbeta) mRNA levels are modulated by vitamin A status in vivo and using transient transfection, that suppression of rat TSHbeta 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 rTSHbeta gene involved in RA regulation, their relationship to the rTSHbeta 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 TSHbeta 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 rTSHbeta negative TRE located between +15 and +43. Mutational analysis also showed that the minor rTSHbeta 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 TSHbeta 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 TSHbeta gene, that the RARE binds RAR/RXR heterodimer and that RA and T3 interact functionally via these elements in the negative regulation of rat TSHbeta gene expression.
Breen
J J
JJ
Department of Biochemistry, Temple University School of Medicine, Philadelphia, PA 19140, USA. jb253x@nih.gov
Hickok
N J
NJ
Gurr
J A
JA
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D019281:Dimerization; D004347:Drug Interactions; D005786:Gene Expression Regulation / Q000187:drug effects*; D008156:Luciferases / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism*; D051381:Rats; D018168:Receptors, Retinoic Acid / Q000235:genetics / Q000502:physiology; D011993:Recombinant Fusion Proteins; D047488:Retinoid X Receptors; D012176:Retinoids / Q000494:pharmacology*; D013963:Thyroid Hormones / Q000494:pharmacology*; D013972:Thyrotropin / Q000235:genetics*; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology; D014162:Transfection; D014212:Tretinoin / Q000494:pharmacology; D014284:Triiodothyronine / Q000494:pharmacology
D012333:RNA, Messenger; D018168:Receptors, Retinoic Acid; D011993:Recombinant Fusion Proteins; D047488:Retinoid X Receptors; D012176:Retinoids; D013963:Thyroid Hormones; D014157:Transcription Factors; D014284:Triiodothyronine; D014212:Tretinoin; D013972:Thyrotropin; D008156:Luciferases