Dominant-negative mutant thyroid hormone receptors prevent transcription from Xenopus thyroid hormone receptor beta gene promoter in response to thyroid hormone in Xenopus tadpoles in vivo.
We describe a dominant-negative approach in vivo to assess the strong, early upregulation of thyroid hormone receptor beta (TR beta) gene in response to thyroid hormone, characteristic of the onset of natural and thyroid hormone-induced amphibian metamorphosis, 3,3',5-Triiodo-thyronine (T3) treatment of organ cultures of premetamorphic Xenopus tadpole tails coinjected in vivo with the wild-type Xenopus TR beta (wt-xTR beta) and three different thyroid responsive element chloramphenicol acetyltransferase (TRE-CAT) reporter constructs, including a direct repeat +4 (DR +4) element in the -200/+87 fragment of the xTR beta promoter, resulted in a 4- to 8-fold enhancement of CAT activity. Two human C-terminal TR beta 1 mutants (delta-hTR beta 1 and Ts-hTR beta 1), an artificial Xenopus C-terminal deletion mutant (mt-xTR beta), and the oncogenic viral homology v-erbA, none of which binds T3, inhibited this T3 response of the endogenous wt-xTR in Xenopus XTC-2 cells cotransfected with the -1600/+87 xTR beta promoter-CAT construct, the potency of the dominant-negative effect of these mutant TRs being a function of the strength of their heterodimerization with Xenopus retinoid X receptor gamma. Coinjection of the dominant-negative Xenopus and human mutant TR beta s into Xenopus tadpole tails totally abolished the T3 responsiveness of the wt-xTR beta with different TREs, including the natural DR +4 TRE of the xTR beta promoter.
Ulisse
S
S
Division of Developmental Biochemistry, National Institute for Medical Research, London, United Kingdom.
Esslemont
G
G
Baker
B S
BS
Krishna
V
V
Chatterjee
K
K
Tata
J R
JR
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000096:biosynthesis; D005786:Gene Expression Regulation; D005799:Genes, Dominant; D006801:Humans; D046911:Macromolecular Substances; D008675:Metamorphosis, Biological / Q000187:drug effects*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009924:Organ Culture Techniques; D011401:Promoter Regions, Genetic; D018168:Receptors, Retinoic Acid / Q000096:biosynthesis / Q000502:physiology; D011988:Receptors, Thyroid Hormone / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D011994:Recombinant Proteins / Q000096:biosynthesis; D012091:Repetitive Sequences, Nucleic Acid; D047488:Retinoid X Receptors; D013623:Tail / Q000254:growth & development; D014157:Transcription Factors / Q000096:biosynthesis / Q000502:physiology; D014158:Transcription, Genetic / Q000187:drug effects / Q000502:physiology*; D014284:Triiodothyronine / Q000494:pharmacology*; D014981:Xenopus
D046911:Macromolecular Substances; D018168:Receptors, Retinoic Acid; D011988:Receptors, Thyroid Hormone; D011994:Recombinant Proteins; D047488:Retinoid X Receptors; D014157:Transcription Factors; D014284:Triiodothyronine; D015500:Chloramphenicol O-Acetyltransferase