Negative regulation of thyroid stimulating hormone α and β subunit genes by thyroid hormone and its receptor

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The negative regulation of thyrotropin (thyroid stimulating hormone, TSH,) production by the thyroid hormone (T3) and its receptor (TR) is the central mechanism of the pituitary-thyroid axis. We previously reported that such negative regulation is able to be observed in kidney derived CV1 cells in the presence of GATA2 and Pit1, which are the thyrotroph specific transcription factors. Using CAT-based reporter assays in CV1 cells, we have analyzed the function of the putative negative regulatory element (NRE) which has been reported to mediate the negative regulation of TSHβ gene by T3-bound TR (T3/TR). Unexpectedly, the negative regulation was maintained even in the deletion or the mutation of NRE. Further analysis revealed that the negative regulation was detected in the construct, TSHβ-D4-CAT, which possesses only Pit1 and GATA2 binding sites. Interestingly, TSHβ-D4-CAT is activated by the presence of GATA2 only and such a transactivation is specifically inhibited by T3/TR. We found that the Zn-finger of GATA2 interacts with the DNA binding domain (DBD) of TR in a T3 independent fashion, suggesting that the negative regulation of TSHβ promoter is mediated by the cross-talk between IR and GATA2. The suppression by T3/TR was recovered by the overexpression of dominant negative-type TR associated protein (TRAP) 220 indicating an essential participation of TRAP220 in the negative regulation of TSHβ gene. Chromatin immunoprecipitation assay with thyrotroph cell line, TαT1, showed that the acetylation of histone H4 is reduced by the addition of T3. In conclusion, the present study suggests (1) GATA2 is the major transcriptional activator for TSHβ promoter (2) T3/TR interacts and interferes with GATA2 (3) TRAP220 that lacks intrinsic histone acetyltransferase activity and the alteration of histone acetylation are involved in the mechanism of the negative regulation of TSHβ gene.