Reconstitution of triiodothyronine inhibition in non-triiodothyronine-responsive thyrotropic tumor cells using transfected thyroid hormone receptor isoforms

The triiodothyronine (T-3) inhibitory effect on the thyrotropin (TSH)beta- and alpha-subunit genes is believed to be mediated by binding of T-3 to specific nuclear receptors that are present in various isoforms. alpha TSH cells, which are derived from a pure alpha-subunit secreting thyrotropic tumor, contain the same nuclear factors that are important for alpha-subunit gene expression in TSH beta-expressing T-3-responsive thyrotropic cells (TtT97). However, as in the parent tumor, alpha-subunit expression in alpha TSH cells was not inhibited by T-3, despite the presence of high-affinity nuclear T-3 receptors (TRs) with a similar number of sites per cell as in TtT97. When transcripts coding for the different TR isoforms from the MGH101A tumor were analyzed by Northern blot, TR alpha(1) was present, as well as the non-T-3-binding variant alpha(2), but transcripts encoding the opposite strand Rev-ErbA alpha were not detectable and neither TR beta(1) nor TR beta(2) mRNAs were detectable, whereas all these transcripts were detectable in TtT97 tumors. Similar findings were observed in alpha TSH cells, where TR beta(1) transcripts were barely detectable in Northern blots and TR beta(2) transcripts were detectable only by RT-PCR. The TRP gene locus is present and unrearranged in the tumor genome. In transient transfection studies conducted in alpha TSH cells overexpression of either TR beta(1), TR beta(2), or TR alpha(1) reconstituted T-3-inhibition of the alpha-subunit promoter down to 40% to 50% of control. We conclude that the relative lack of TRP gene expression correlates with unresponsiveness to T-3. The alpha TSH cell line represents a unique model in which to dissect the mechanism of T-3 inhibition.