Dominant negative activity of thyroid hormone receptor variant α2 and interaction with nuclear corepressors

The splicing variant of the thyroid hormone receptor alpha (TR alpha) gene, TR variant alpha 2 (TRv alpha 2), lacks the second half of the ninth heptad, a domain thought to be important for heterodimerization with retinoid X receptors (RXRs). In transient transfection studies, TRv alpha 2 exhibits weak dominant negative inhibition of TR alpha 1-mediated transcription. In contrast, a TRv alpha 2 mutant in which the ninth heptad was restored (alpha 2 + 9H), exhibits very strong dominant negative activity. We have examined the role of nuclear corepressors (CoRs) in the dominant negative activity of TRv alpha 2 and alpha 2 + 9H. Glutathione S-transferase pull down experiments revealed that TRv alpha 2 barely interacts with CoRs, whereas alpha 2 f 9H interaction with CoRs is as strong as that of TR alpha 1. A P160R CoR box mutation was introduced in the context of TRv alpha 2 and alpha 2 + 9H, which nearly abolishes the ability of these receptors to interact with CoRs. In transient transfection the dominant negative activity of TRv alpha 2 was only marginally impaired by the P160R mutation. In contrast, alpha 2 + 9H-P160R had approximate to 66% less dominant negative activity than alpha 2 + 9H. These results suggest that the weak dominant negative activity of TRv alpha 2 is due in part to its lack of interaction with CoRs, and that restoration of the ninth heptad restores CoR interaction and strong dominant negative activity. Further, the data reveal aspects of the dominant negative action that are dependent on the orientation of the TRE. (C) 1999 Elsevier Science Ireland Ltd. All rights reserved.