Naturally Occurring Amino Acids in Helix 10 of the Thyroid Hormone Receptor Mediate Isoform-Specific TH Gene Regulation

Abstract

Thyroid hormone (TH) action is mediated by the products of two genes, TH receptor (THR)α (THRA) and THRβ (THRB) that encode several closely related receptor isoforms with differing tissue distributions. The vast majority of THR isoform-specific effects are thought to be due to tissue-specific differences in THR isoform expression levels. We investigated the alternative hypothesis that intrinsic functional differences among THR isoforms mediate these tissue-specific effects. To achieve the same level of expression of each isoform, we created tagged THR isoforms and tested their DNA and functional properties in vitro. We found significant homodimerization and functional differences among the THR isoforms. THRA1 was unable to form homodimers on direct repeat separated by 4 bp DNA elements and was also defective in TH-dependent repression of Tshb and Rxrg in a thyrotroph cell line, TαT1.1. In contrast, THRB2 was both homodimer sufficient and fully functional on these negatively regulated genes. Using domain exchanges and individual amino acid switches between THRA1 and THRB2, we identified three amino acids in helix 10 of the THRB2 ligand-binding domain that are required for negative regulation and are absent in THRA1.

Regulation of hypothalamic reactive oxygen species and feeding behavior by phosphorylation of the beta 2 thyroid hormone receptor isoform

“…To test the direct role of THRB2 phosphorylation in ROS metabolism, we employed two neuroblastoma cell lines, Neuro-2a and N1E-115 (commercially available, see Methods). Both lines express Thra , but have extremely low or no Thrb1/Thrb2 expression, allowing us to develop a neuronal cell model expressing similar levels of HA tagged THRB1, THRB2, and THRB2 S101A 50 . Cells expressing incrementally higher levels of THRs were subjected to oxidative stress using tBHP (Fig.…”

“…Cells were maintained at 37 °C, 5% CO 2 . Cells were transduced with replication incompetent adenoviral constructs (expressing GFP, HA-THRB2 WT , HA-THRB2 S101A , HA-THRB1, and HA-THRA1 ) as described previously 12 , 50 , 54 . Treatments with T 3 , tert-Butyl Hydroperoxide (tBHP, Thermo Fisher Scientific, PA, USA), were performed in DMEM high glucose containing 6% charcoal:dextran stripped FBS (Gemini bio-products, CA, USA) 24 h after transduction.…”

Daily exposure to low concentrations Tetrabromobisphenol A interferes with the thyroid hormone pathway in HepG2 cells

“…After exposure to low-concentration TBBPA for five generations, the genes related to the thyroid hormone signaling pathway were upregulated 1.31−7.37 folds in HepG2 cells. These genes were mainly associated with the Ras homolog (RHEB2) [35] , solute carrier family 16 member 2 (SLC16A2) [36] , thyroid hormone receptor beta (THRB) [37] , type Ⅱ thyroxine 5′-deiodinase (DIO2) [38] , Ras-related protein M-Ras (MR) [39] , thyroid hormone receptor alphaⅠ (THRA1) [40] , thyroid hormone receptor alphaⅠ type 2 (THRA2) [41] , Ras-related protein H-Ras (HR), [42] and protein kinase A (PKA) [43] . In addition, the expression levels of genes encoding PKA and parathyroid hormone receptor (PTHR1) were upregulated 2.11−4.46 folds in HepG2 cells after exposure to TBBPA, probably related to the parathyroid hormone signaling pathway [44] .…”