Reconstitution of Triiodothyronine Inhibition in Non-Triiodothyronine-Responsive Thyrotropic Tumor Cells Using Transfected Thyroid Hormone Receptor Isoforms

Abstract

The triiodothyronine (T₃) inhibitory effect on the thyrotropin (TSH)-β-and a-subunit genes is believed to be mediated by binding of T₃ to specific nuclear receptors that are present in various isoforms. αTSH cells, which are derived from a pure α-subunit secreting thyrotropic tumor, contain the same nuclear factors that are important for α-subunit gene expression in TSHβ-expressing T₃-responsive thyrotropic cells (TtT97). However, as in the parent tumor, α-subunit expression in αTSH cells was not inhibited by T₃, despite the presence of high-affinity nuclear T₃ 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, TRα₁ was present, as well as the non-T₃-binding variant α₂, but transcripts encoding the opposite strand Rev-ErbAα were not detectable and neither TR/β₁ nor TRβ₂ mRNAs were detectable, whereas all these transcripts were detectable in TtT97 tumors. Similar findings were observed in αTSH cells, where TRβ₁ transcripts were barely detectable in Northern blots and TRβ₂ transcripts were detectable only by RT-PCR. The TRβ gene locus is present and unrearranged in the tumor genome. In transient transfection studies conducted in αTSH cells overexpression of either TRβ₁, TRβ₂, or TRα₁ reconstituted T₃-inhibition of the α-subunit promoter down to 40% to 50% of control. We conclude that the relative lack of TRβ gene expression correlates with unresponsiveness to T₃. The αTSH cell line represents a unique model in which to dissect the mechanism of T₃ inhibition.