Difference in dominant negative activities between mutant thyroid hormone receptors α1 and β1 with an identical truncation in the extreme carboxyl-terminal tau4 domain

Nishiyama, Kozo; Andoh, Shinichiro; Kitahara, Akira; Natsume, Hiroko; Mikami, Takuma; Genma, Rieko; Nakamura, Hirotoshi

doi:10.1016/s0303-7207(98)00014-8

Cited by 7 publications

(4 citation statements)

References 34 publications

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“…aF397X has the equivalent C-terminal truncation to bF451X identified in a patient with severe RTH [20]. In a transient transfection assay in CV-1 cells, aF397X showed strong dominant negative effects comparable to bF451X through several positive TREs [15]. We carried out microinjection of the transgene into fertilized eggs once for the EF1a-bF451X and eight times for the EF1a-aF397X.…”

Section: Resultsmentioning

confidence: 99%

“…The EF1a-bF451X and EF1a-aF397X transgenes were constructed as follows. A 1.5 kbp HindIII/BamHI fragment containing bF451X cDNA and a 1.9 kbp HindIII/BamHI fragment containing aF397X cDNA were derived from pCMX-bF451X and pCMX-aF397X, respectively [15]. Both fragments were filled in using the Klenow enzyme and inserted into the blunt-ended XbaI site of pEF-BOS, which was kindly provided by Dr. S. Nagata [16].…”

Section: Methodsmentioning

confidence: 99%

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Embryonic Lethal Effect of Expressing a Dominant Negative Mutant Human Thyroid Hormone Receptor .ALPHA.1 in Mice

et al. 2003

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Abstract. Resistance to thyroid hormone (RTH) is caused mainly by mutations of the thyroid hormone receptor (TR) b gene. Although, in vitro, TRa1 and TRb1 mutants exhibit similar dominant negative effects against wild-type TR, no TRa mutants have ever been identified in RTH patients. It has been postulated that mutations in TRa gene may be lethal, compensated completely by intact TRb or associated with phenotypic manifestations different from RTH. To investigate the consequences of mutant TRa1 expression in vivo, we tried to generate two different lines of transgenic mice which express a strong or a weak dominant negative mutant TRa1, respectively. First, we expressed bF451X identified in a patient with severe RTH and aF397X, which has an identical C-terminal truncation and a similarly strong dominant negative potency to bF451X, under the control of human polypeptide chain elongation factor 1a promoter. Six bF451X-transgenic mice were born from 223 transferred embryos, giving a transgenic frequency of 2.7%. By contrast, expression of aF397X resulted in quite a low transgenic frequency with 0.39% of the transferred embryos bearing the transgene. Only three transgenic mice were born with no apparently overt abnormalities, of which one male produced F1 offspring. The transgenic progeny expressed aF397X in the testis but we did not succeed in generating transgenic mice expressing aF397X in multiple organs. To avoid toxic effects mediated by a strong dominant negative activity of mutant TRa1, we exchanged aF397X for aK389E, which has an identical missense mutation and a relatively weak transdominant potency as bK443E identified in a patient with mild RTH. When expressed by cytomegalovirus immediate early enhancer-chicken b-actin promoter, we did not succeed in creating aK389E-transgenic mice despite three independent transgene-injections. These findings define crucial in vivo functions of mutant TRa1s during mouse fetal development and suggest the possibility that the expression of a dominant negative mutant TRa1 in extensive tissues from the early embryonal stages might be lethal.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Embryonic Lethal Effect of Expressing a Dominant Negative Mutant Human Thyroid Hormone Receptor .ALPHA.1 in Mice

et al. 2003

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“…The base substitutions in the recombinant expression vectors were confirmed by DNA sequencing. Expression vectors (pCMX) encoding human TR 1, TR 1, F451X, F397X and human RXR were the same as those used in the reference reported by Nishiyama et al (1998). Reporter plasmids containing TRE-DR4, TREpal2 and the chicken lysozyme silencer F2-TRE (F2-TRE) were described previously (Nishiyama et al 1998).…”

Section: Plasmidsmentioning

confidence: 99%

“…Although F451X and E449X have the same functional disruption of the autonomous activation function domain (AF2) and negligible T 3 -binding activity, the silencing potency of F451X was stronger than that of E449X and wild-type TR 1 (Miyoshi et al 1998). We also analysed the DNA-binding and dimerization properties of wild-type TR 1 and F451X by a gel-shift assay, but no apparent difference was observed in DNA binding between wild-type TR 1 and F451X (Nishiyama et al 1998).…”

Section: Introductionmentioning

confidence: 99%

Differences between the silencing-related properties of the extreme carboxyl-terminal regions of thyroid hormone receptors alpha 1 and beta 1

Nishiyama

Matsushita²,

Natsume³

et al. 2000

Journal of Endocrinology

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Human thyroid hormone receptor (TR) is encoded by two distinct genes, TR and TR . TR heterodimerizes with retinoid X receptor (RXR) and binds efficiently to the thyroid hormone (T 3 ) response element (TRE) of target genes. In the absence of T 3 , unliganded TR suppresses the basal promoter activity of positively regulated genes (silencing). Silencing mediator for retinoid and thyroid hormone receptors (SMRT) and nuclear receptor co-repressor (N-CoR) interact with unliganded TR and function as corepressor proteins. Previously, we found F451X with carboxyl (C)-terminal 11-amino acid deletion had stronger silencing potency than wild-type TR 1 and E449X with C-terminal 13-amino acid deletion on a subset of TREs. In the present study, to assess the isoform-specific effects of the C-terminal truncations on TR silencing, we constructed two mutant TR 1s ( F397X and E395X) with the same respective C-terminal truncations as F451X and E449X and analysed their silencing activities. Unlike F451X and E449X, F397X and E395X showed similarly stronger silencing potency than wild-type TR 1. We further studied the abilities of wild-type and the mutant TR 1s and 1s on RXR and co-repressor binding by a twohybrid interference assay. F451X had significantly stronger abilities to bind to RXR and SMRT than did wild-type TR 1 and E449X. In contrast, wild-type TR 1, F397X and E395X showed similar abilities to bind to RXR and SMRT. E449X and E395X, which have identical C-terminal truncation, showed less ability to bind to N-CoR than did wild-type TR 1 and F451X and wild-type TR 1 and F397X respectively. These results indicate that an identical C-terminal truncation gives rise to different effects on TR 1 and 1 with respect to silencing potency, RXR binding and SMRT binding. The difference in the silencing potency among wild-type TR 1, F451X and E449X correlated well with the difference in the ability to bind co-repressor SMRT.

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Learning from Nature’s Experiments on the Thyroid Hormone Receptor; X-Ray Structures of RTH Mutant Ligand-Binding Domains

Sandler

Baxter

Fletterick

2004

Syndromes of Hormone Resistance on the Hypothalamic-Pituitary-Thyroid Axis

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Difference in dominant negative activities between mutant thyroid hormone receptors α1 and β1 with an identical truncation in the extreme carboxyl-terminal tau4 domain

Cited by 7 publications

References 34 publications

Embryonic Lethal Effect of Expressing a Dominant Negative Mutant Human Thyroid Hormone Receptor .ALPHA.1 in Mice

Embryonic Lethal Effect of Expressing a Dominant Negative Mutant Human Thyroid Hormone Receptor .ALPHA.1 in Mice

Differences between the silencing-related properties of the extreme carboxyl-terminal regions of thyroid hormone receptors alpha 1 and beta 1

Learning from Nature’s Experiments on the Thyroid Hormone Receptor; X-Ray Structures of RTH Mutant Ligand-Binding Domains

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