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Mutations of Thyroid Hormone Nuclear Receptors and Disease
Given the critical roles of TRs in cellular functions, it is reasonable to expect that mutations of TRs could have deleterious consequences. Indeed, shortly after the cloning of the TRβ gene, mutations of it were discovered to cause the genetic syndrome of resistance to thyroid hormone (RTH). However, whether mutations of the TRβ gene cause human diseases other than RTH has been unknown. Likewise, it has been unclear whether mutations of the TRα gene could also cause abnormalities. To address these questions, we used the powerful mouse genetic approach, introducing an identical mutation (PV mutation) into the TRβ and the TRα gene loci. The PV mutation was identified in an RTH patient at the NIH. It is a frame-shift mutation in the C-terminal 16 amino acids of TRβ1 (TRβ1PV) and TRα1 (TRα1PV) that leads to the complete loss of T3 binding activity and transcription capacity (Figure 1). Figure 1. The amino acid sequence of PV and its location in the carboxyl terminus of TRβ1 (A) and TRα1 (B). The PV mutation was identified in a patient with resistance to thyroid hormone. The mutation is from a C-insertion at codon 448 of TRβ1, resulting in a frame-shift mutation in the last 16 carboxyl terminal amino acids. The same PV mutation was targeted to the TRβ and TRα genes to create TRβPV and TRα1PV mice. The knockin mouse that harbors theTRβPV gene (TRβPV mouse; Figure 1, part A) recapitulates human RTH by exhibiting dysregulation of the pituitary-thyroid axis, reduced weight, abnormally accelerated bone development, hypercholesterolemia, and hyperactivity. This mouse model allowed an elucidation of the molecular basis of RTH unattainable otherwise. TRβPV manifests its dominant-negative activity in vivo via competition with wild-type TRs in binding to the promoters of T3-target genes. The variable phenotypic expression in RTH patients is dictated by the tissue-dependent abundance of TR isoforms and modulated by multiple combinatorial cellular factors. Remarkably, homozygous TRβPV/PV mice spontaneously develop follicular thyroid carcinoma, indicating that the deleterious effect of TRβ gene mutations is not limited to RTH. The pathologic progression from hyperplasia to capsular and vascular invasion, and eventually to distant metastasis in TRβPV/PV mice, is similar to human follicular thyroid cancer (Figure 2). Thyroid carcinomas are the most common endocrine neoplasms in humans, with a globally increasing incidence. However, little is known about the molecular genetic events underlying their development. This first mouse model of follicular thyroid carcinoma allows one to discern the genetic alterations contributing to thyroid carcinogenesis and to identify potential molecular targets for prevention and treatment. Indeed, analysis of altered gene expression profiles by cDNA microarray indicates a complex alteration of multiple signaling pathways is associated with thyroid carcinogenesis. One pathway, the peroxisome proliferator–activated receptor γ (PPARγ)–mediated signaling, was found to be repressed by PV during thyroid carcinogenesis. Further molecular study of this pathway led to the identification of PPARγ as a tumor suppressor, thus raising the possibility that PPARγ is a potential target for treatment. Indeed, activation of PPARγ-mediated signaling by treating TRβPV/PV mice with PPARγ agonists significantly delays the development and progression of thyroid carcinogenesis. These results suggest that PV could act as an oncogene by inhibiting the tumor suppressor functions of PPARγ in thyroid carcinogenesis. Figure 2. Morphological features in thyroid glands and metastasis of TRβPV/PV mice. Histological sections from tissues of TRβPV/PV mice stained with hematoxylin (blue) and eosin (pink) show evidence of capsular invasion (A) (arrows) and vascular invasion in thyroid (B) (arrow), spindle cell anaplasia within the thyroid shown at higher magnification (C) (arrow), and a cardiac metastasis (D) (arrow). Capsular and vascular invasion are the pathologic features used in the diagnosis of human neoplastic thyroid tumors. The pathologic progression of thyroid cancer in TRβPV/PV mice is similar to that in humans. The manifestation of the oncogenic actions of PV is not restricted to the thyroid. TRβPV/PV mice also spontaneously develop thyroid stimulating hormone (TSH)–secreting pituitary tumors (TSH-omas). TSH-omas represent about 2% of all pituitary adenomas in humans, affecting vision and causing headaches and other endocrine disorders. The molecular genetics underlying their pathogenesis is largely unknown. Using TRβPV/PV mice as a model, we uncovered a novel mechanism by which PV could function as an oncogene in TSH-omas. PV acts as a constitutive activator of the expression of cyclin D1, a well-known tumor promoter, by tethering to the cyclic AMP response element binding protein (CREB) on the cyclin D1 promoter. These findings suggest that mutation of TRβ is one of the genetic events underlying the pathogenesis of TSH-omas. The knockin mice harboring the PV mutation in the TRα gene (TRα1PV mice; Figure 1, part B) exhibit a phenotype distinct from that of TRβPV mice. Homozygous TRα1PV/PV mice die very shortly after birth. The heterozygous mice (TRα1PV/+) display reduced fertility, increased mortality, delayed bone development, dwarfism, and metabolic disorder, indicating that mutations of TRα lead to severe consequences. The contrasting phenotypes of TRα1PV and TRβPV mice reveal that the actions of TR mutants in vivo are isoform dependent. Analysis of transcription regulation of T3-response genes in several target tissues suggests that distinct phenotypic expression is mediated, in part, by the differentially dominant activity of TR isoform mutants in vivo. The TRα1PV and TRβPV mice have provided a powerful tool to uncover the novel functions of TR mutants and to elucidate their mechanisms of molecular actions in vivo. Accumulated evidence from the study of TRβPV/PV mice suggests that PV, a TRβ mutant, not only causes RTH, but could also function as a new oncogene to contribute to thyroid carcinogenesis and pathogenesis of TSH-omas. Importantly, TRβPV/PV mice could be used as a preclinical model to develop new treatment strategies. Considering that human diseases harboring TRα mutations are yet to be discovered, TRα1PV mice could be used as a model to search for and to identify those diseases whose phenotypic manifestation resembles that of TRα1PV mice.
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hyroid hormone nuclear receptors (TRs) are ligand-dependent transcription factors that mediate the biological activities of thyroid hormone (T3) in growth, development, differentiation, and the maintenance of metabolic homeostasis. Two TR genes, TRα and TRβ, located on chromosomes 17 and 3 respectively, encode four major T3-binding TR isoforms (α1, β1, β2, and β3). The TRs consist of modular functional structures with the N-terminal A/B, central DNA-binding, and the C-terminal ligand-binding domains. TRα1 and TRβ1 share high sequence homology in the DNA- and ligand-binding domains, but the sequence differs significantly in the A/B domain. The C-terminal region and the A/B domain contain the transcription-activation functions.
