IDENTIFICATION OF A CIS-REGULATORY ELEMENT AND A THYROID-SPECIFIC NUCLEAR FACTOR MEDIATING THE HORMONAL-REGULATION OF RAT-THYROID PEROXIDASE PROMOTER ACTIVITY

The mechanism for hormonal regulation of rat thyroperoxidase (rTPO) ge ne transcription in rat FRTL-5 thyroid cells has been investigated. Tr ansient transfection experiments demonstrate that the minimal rTPO pro moter that confers thyroid-specific expression also confers responsive ness to TSH and insulin. TSH induces a 7-fold increase in promoter act ivity, and the induction is detected almost immediately after the addi tion of the hormone. Insulin also stimulates TPO promoter activity, bu t the effect of this hormone is weaker and slower than that of TSH. Th e effect of TSH in increasing TPO promoter activity is mimicked by the cAMP agonist forskolin. The calcium-protein kinase C pathway is also involved in the regulation of the rTPO promoter activity, since a calc ium ionophore (A23187) and phorbol esters [12-O-tetradecanoyl-phorbol- 13-acetate (TPA)] inhibit it quickly. These data indicate that the reg ion of the rTPO promoter used here contains the DNA signals necessary for its hormonal regulation. Protein-DNA binding studies show that the thyroid-specific nuclear protein TTF-2, which binds to the rTPO promo ter, is induced by TSH and forskolin, and this effect is clearly obser vable as early as 5 h post induction. Moreover, the DNA binding activi ty of TTF-2 is inhibited by both A23187 and TPA. Heterologous promoter constructs containing four, eight, or 12 tandem repeats of an oligonu cleotide that includes the TTF-2 binding site increase their activity in response to TSH, forskolin, and insulin, while the the presence of A23187 or TPA inhibits their activity. These data indicate thata the T TF-2 protein plays an important role in the hormonal control of thyroi d-specific transcription.