A bimodal switch model is widely used to describe transcriptional regulation by the thyroid hormone receptor (TR). In this model, the unliganded TR forms stable, chromatin-bound complexes with transcriptional co-repressors to repress transcription. Binding of hormone dissociates co-repressors and facilitates recruitment of co-activators to activate transcription. Here we show that in addition to hormone-independent TR occupancy, ChIP-seq against endogenous TR in mouse liver tissue demonstrates considerable hormone-induced TR recruitment to chromatin associated with chromatin remodelling and activated gene transcription. Genome-wide footprinting analysis using DNase-seq provides little evidence for TR footprints both in the absence and presence of hormone, suggesting that unliganded TR engagement with repressive complexes on chromatin is, similar to activating receptor complexes, a highly dynamic process. This dynamic and ligand-dependent interaction with chromatin is likely shared by all steroid hormone receptors regardless of their capacity to repress transcription in the absence of ligand.

Activation of the phosphatidylinositol 3-kinase (PI3K)–AKT/protein kinase B signaling pathway has been associated with multiple human cancers. Recently we showed that AKT is activated in both the thyroid and metastatic lesions of a mouse model of follicular thyroid carcinoma [thyroid hormone β receptor (TRβ)^PV/PV mice]. This TRβ^PV/PV mouse harbors a knock-in mutant TRβ gene (TRβPV mutant) that spontaneously develops thyroid cancer and distant metastasis similar to human follicular thyroid cancer. Here we show that in thyroid tumors, PV mutant bound significantly more to the PI3K-regulatory subunit p85α, resulting in a greater increase in the kinase activity than did TRβ1 in wild-type mice. By GST pull-down assays, the ligand-binding domain of TR was identified as the interaction site with p85α. By confocal fluorescence microscopy, p85α was shown to colocalize with TRβ1 or PV mainly in the nuclear compartment of cultured tumor cells from TRβ^PV/PV mice, but cytoplasmic p85α/PV or p85α/TRβ1 complexes were also detectable. Further biochemical analysis revealed that the activation of the PI3K–AKT–mammalian target of the rapamycin–p70^S6K pathway was observed in both the cytoplasmic and nuclear compartments, whereas the activation of the PI3K–integrin-linked kinase–matrix metalloproteinase 2 pathway was detected mainly in the extranuclear compartments. These results suggest that PV, via the activation of p85α, could act to affect PI3K downstream signaling in both the nuclear and extranuclear compartments, thereby contributing to thyroid carcinogenesis. Importantly, the present study unveils a mechanism by which a mutant TR acts to activate PI3K activity via protein–protein interactions.

Thyroid hormone receptors (TRs) play critical roles in energy homeostasis. To understand the role of TRs in lipid homeostasis in vivo, we adopted the loss-of-function approach by creating knock-in mutant mice with targeted mutation in the TRα gene (TRα1PV mouse) or TRβ gene (TRβPV mouse). The PV mutation, identified in a patient with resistance to thyroid hormone, exhibits potent dominant-negative activity. Here we show that in contrast to TRα1PV mouse, TRβPV mice exhibited no significant reduction in WAT but had significant increases in serum free fatty acids and total triglycerides. Moreover, the liver of TRβPV mice was markedly increased (33%) with excess lipid accumulation, but the liver mass of TRα1PV mouse was decreased (23%) with paucity of lipids. These results indicate that apo-TRβ and apo-TRα1 exerted distinct abnormalities in lipid metabolism. Further biochemical analyses indicate that increased lipogenic enzyme expression, activated peroxisome proliferator-activated receptor γ (Pparγ) signaling, and decreased fatty acid β-oxidation activity contributed to the adipogenic steatosis and lipid accumulation in the liver of TRβPV mice. In contrast, the expression of lipogenic enzymes and Pparγ was decreased in the liver of TRα1PV mice. These results suggest that the regulation of genes critical for lipid metabolism by TRs in the liver is isoform dependent. These results indicate that apo-TRβ and apo-TRα1 had different effects on lipid metabolism and that both TR isoforms contribute to the pathogenesis of lipid metabolism in hypothyroidism.

Aberrant activation of the phosphatidylinositol 3-kinase (PI3K)–AKT/protein kinase B-signaling pathway has been associated with multiple human cancers, including thyroid cancer. Recently, we showed that, similar to human thyroid cancer, the PI3K–AKT pathway is overactivated in both the thyroid and metastatic lesions of a mouse model of follicular thyroid carcinoma (TRβ^PV/PV mice). This TRβ^PV/PV mouse harbors a knockin mutant thyroid hormone receptor β gene (TRβPV mutant) that spontaneously develops thyroid cancer and distant metastasis similar to human follicular thyroid cancer. That the activation of the PI3K–AKT signaling contributes to thyroid carcinogenesis raised the possibility that this pathway could be a potential therapeutic target in follicular thyroid carcinoma. The present study tested this possibility by treating TRβ^PV/PV mice with LY294002 (LY), a potent and specific PI3K inhibitor, and evaluating the effect of LY on the spontaneous development of thyroid cancer. LY treatment inhibited the AKT–mammalian target of rapamycin (mTOR)–p70^S6K signaling, and it decreased cyclin D1 and increased p27^Kip1 expression to inhibit thyroid tumor growth and reduce tumor cell proliferation. LY treatment increased caspase 3 and decreased phosphorylated-BAD to induce apoptosis. In addition, LY treatment reduced the AKT–matrix metalloproteinase 2 signaling to decrease cell motility to block metastatic spread of thyroid tumors. Thus, these altered signaling pathways converged effectively to prolong survival of TRβ^PV/PV mice treated with LY. No significant adverse effects were observed for wild-type mice treated similarly with LY. The present study provides the first preclinical evidence for the in vivo efficacy for LY in the treatment of follicular thyroid cancer.