CELL TYPE-DEPENDENT MODULATION OF THE DOMINANT-NEGATIVE ACTION OF HUMAN MUTANT THYROID-HORMONE BETA-1 RECEPTORS

Background: Mutations in the ligand-binding domain of the thyroid horm one receptor beta (TR beta) gene cause the syndrome of resistance to t hyroid hormone (RTH). The clinical phenotype results from the antagoni sm of the normal TR alpha and the non-mutated TR beta alleles by the T R beta 1 mutants, via a dominant negative effect. There is, however, m arked heterogeneity of organ resistance within and among kindreds with RTH. This study examines the potential role of cell type in modulatin g the dominant negative potency of human TR beta 1 (h-TR beta 1) mutan ts. Materials and Methods: Transient transfections were performed in H eLa and NIH3T3 cells, using a wild type (WT) and three naturally occur ring mutant h-TR beta 1 constructs, and three natural thyroid hormone response elements (TREs). Immunocytochemistry was performed to detect levels of TR beta 1 expression in these two cell types. In order to de termine how TR beta 1 interacts with other cellular partners, gel-shif t analyses using HeLa and NM3T3 nuclear extracts were performed. Resul ts: Transfection studies using WT h-TR beta 1 in HeLa and NM3T3 cells, showed that the 3,3',5-triiodothyronine (T-3)-induced transactivation of the different TREs varied between cell. types. Unlike the non-T-3- binding h-TR beta 1 mutant, PV, mutants ED and OK displayed the expect ed T-3-induced dose responsiveness in these two cell types. For each T RE examined, the magnitude of the dominant negative effect varied betw een the cell types. The levels of receptor expression in HeLa and NIH3 T3 cells were identical, as determined by immunocytochemistry. Gel-shi ft analyses showed differences in the formation of hetero- and homodim ers depending on both the cell type and TRE motif. Conclusions: The ce ll type in which a mutant receptor operates affects the relative amoun ts of hetero- and homodimers. Together with the nature of the mutation and the TRE-motif, this could modulate the dominant negative action o f mutant receptors in different tissues, which, in turn, could contrib ute to the variable phenotypic characteristics of RTH.