A Dominant-Negative Thyroid Hormone Receptor Blocks Amphibian Metamorphosis by Retaining Corepressors at Target Genes

Abstract

The total dependence of amphibian metamorphosis on thyroid hormone (T 3 ) provides a unique vertebrate model for studying the molecular mechanism of T 3 receptor (TR) function in vivo. In vitro transcription and developmental expression studies have led to a dual function model for TR in amphibian development, i.e., TRs act as transcriptional repressors in premetamorphic tadpoles and as activators during metamorphosis. We examined molecular mechanisms of TR action in T3-induced metamorphosis by using dominant-negative receptors (dnTR) ubiquitously expressed in transgenic Xenopus laevis. We showed that T 3 -induced activation of T 3 target genes and morphological changes are blocked in dnTR transgenic animals. By using chromatin immunoprecipitation, we show that dnTR bound to target promoters, which led to retention of corepressors and continued histone deacetylation in the presence of T 3 . These results thus provide direct in vivo evidence for the first time for a molecular mechanism of altering gene expression by a dnTR. The correlation between dnTR-mediated gene repression and inhibition of metamorphosis also supports a key aspect of the dual function model for TR in development: during T 3 -induced metamorphosis, TR functions as an activator via release of corepressors and promotion of histone acetylation and gene activation.

Gene-specific Changes in Promoter Occupancy by Thyroid Hormone Receptor during Frog Metamorphosis

“…TRs bind TREs in the absence and presence of ligand (1). Most interestingly, in our studies on TR binding to TREs in vivo during metamorphosis by ChIP, we often observed variable amounts of TR binding to TRE of the TH-inducible TH/bZIP promoter, depending on the presence of ligand (15,25). To investigate the TH-dependent phenomenon, we treated premetamorphic tadpoles with or without 3,5,3Ј-triiodothyronine (T 3 ), a potent form of TH, and analyzed TR binding to two TH-inducible promoters, TR␤ and TH/bZIP, in the intestine and tail with anti-TR(PB) antibody (Fig.…”

“…The destruction of larval organs and the formation of adult organs during frog metamorphosis is totally dependent upon TH (13,14). Transgenic overexpression of mutant TRs and cofactors showed gene activation by TR is necessary and sufficient to initiate TH-dependent developmental transitions in frogs (15,16). In knock-out mice lacking TRs, developmental defects are evident in brain, heart, and intestine, among other organs (17).…”

A Causative Role of Stromelysin-3 in Extracellular Matrix Remodeling and Epithelial Apoptosis during Intestinal Metamorphosis in Xenopus laevis

“…Northern blot hybridization was done as previously described with a 32 P-labeled ST3 probe (43). RT-PCR was performed by using Superscript One-Step RT-PCR (Invitrogen) from 0.1 g of total RNA as described (44). RT-PCR for the transgene was done with the primers 5Ј-GGT ATC CTC ACC TGA TCA GTC AAG-3Ј (in the ST3 coding region) and 5Ј-CTT CAG CAC GTG TCT TGT AGT TCC-3Ј (in the GFP coding region).…”

Genistein prevents thyroid hormone‐dependent tail regression of Rana catesbeiana tadpoles by targetting protein kinase C and thyroid hormone receptor α

“…Key players in this transition are the TH receptors, TR␣ and TR␤ (Schreiber et al, 2001;Buchholz et al, 2003Buchholz et al, , 2004. To investigate whether genistein affects TR␣ and TR␤ expression, tadpoles were injected with vehicle alone (DMSO) or T 3 and tail tips were cultured after 24 hr (24-hr injection) or 48 hr (48-hr injection).…”

“…TR␣ and TR␤ are the two major TR isoforms that possess dual functions as transcription repressors and activators in the absence and presence of ligand (primarily T 3 ), respectively (Sachs et al, 2000). Use of dominantnegative TR␣ (Schreiber et al, 2001;Buchholz et al, 2004) and dominantpositive TR␣ genes (Buchholz et al, 2003) in transgenic Xenopus laevis has shown that TR␣ is critical for T 3 -induced frog metamorphosis and for the regulation of known T 3 -response genes, such as TR␤, at the transcriptional level in the tail. TR␣ is present in the premetamorphic tadpole tail whereas TR␤ levels are extremely low until TH induction (Eliceiri and Brown, 1994;Veldhoen et al, 2002), suggesting that TR␣ is critical for the competence to respond to TH while TR␤ is required for establishment of the metamorphic genetic program.…”