THE UNIQUE C-TERMINI OF THE THYROID-HORMONE RECEPTOR VARIANT, C-ERBA-ALPHA-2, AND THYROID-HORMONE RECEPTOR ALPHA-1 MEDIATE DIFFERENT DNA-BINDING AND HETERODIMERIZATION PROPERTIES

Thyroid hormone receptors (TRs) mediate the regulation of gene transcription by thyroid hormone (T3) by binding to T3-responsive elements (TREs) in target genes. c-erbA-alpha-2 is a C-terminal TR variant which does not bind T3 and is a dominant inhibitor of T3 action. When synthesized in Escherichia Coli, alpha-2 formed two TRE-binding complexes similar to the monomeric and homodimeric forms of TR-alpha-1. However, alpha-2 did not bind nearly as well as TR-alpha-1. Furthermore, alpha-2 failed to bind DNA with proteins that heterodimerized with TR-alpha-1. TR-alpha-1 and alpha-2 also did not bind DNA as heterodimers with one another. The differences between TR-alpha-1 and alpha-2 were further analyzed by studying a variety of C-terminal mutants synthesized in reticulocyte lysates. Deletion of the last 20 of the 122 unique amino acids (aa) of alpha-2 increased its DNA binding to approximately the level of TR-alpha-1, indicating that the C-terminus of alpha-2 is an inhibitory domain. This alpha-2 mutant (alpha-2-DELTA-C) was still unable to heterodimerize with nuclear proteins, as were C-terminal deletion mutants of TR-alpha-1. We hypothesized that fusion of TR-alpha-1-specific sequences to the C-terminus of alpha-2-DELTA-C would transfer the property of heterodimerization. Indeed, although alpha-2/alpha-1 chimeras containing the last 40 and 70 aa of TR-alpha-1 failed to heterodimerize with nuclear proteins, addition of the last 100 or 150 aa of TR-alpha-1 did render alpha-2-DELTA-C heterodimerization competent. Thus, TR-alpha-1 contains a C-terminal structure which is necessary for heterodimerization and can confer this property on alpha-2, which lacks this domain. The effects of the unique C-termini of TR-alpha-1 and alpha-2 on their in vitro DNA binding have important implications for their mechanisms of action in vivo.