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The molecular mechanism involved in the liganded thyroid hormone receptor suppression of the TSHbeta (thyroid-stimulating hormone beta, or thyrotropin beta) gene transcription is undetermined. One of the main reasons is the limitation of useful cell lines for the experiments. We have developed an assay system using non-pituitary CV1 cells and studied the negative regulation of the TSHbeta gene. In CV1 cells, the TSHbeta-CAT (chloramphenicol acetyltransferase) reporter was stimulated by Pit1 and GATA2 and suppressed by T3 (3,3',5-tri-iodothyronine)-bound thyroid hormone receptor. The suppression was dependent on the amounts of T3 and the receptor. Unliganded receptor did not stimulate TSHbeta activity, suggesting that the receptor itself is not an activator. Analyses using various receptor mutants revealed that the intact DNA-binding domain is crucial to the TSHbeta gene suppression. Co-activators and co-repressors are not necessarily essential, but are required for the full suppression of the TSHbeta gene. Among the three receptor isoforms, beta2 exhibited the strongest inhibition and its protein level was the most predominant in a thyrotroph cell line, TalphaT1, in Western blotting. The dominant-negative effects of various receptor mutants measured on the TSHbeta-CAT reporter were not simple mirror images of those in the positive regulation under physiological T3 concentration.

biochemical journal

Thyroid-hormone-dependent negative regulation of thyrotropin beta gene by thyroid hormone receptors: study with a new experimental system using CV1 cells