Regulation of Fibroblast Growth Factor Receptor-1 (Fgfr1) by Thyroid Hormone: Identification of a Thyroid Hormone Response Element in the Murine Fgfr1 Promoter

T3 is essential for normal skeletal development, acting mainly via the TRα1 nuclear receptor. Nevertheless, the mechanisms of T3 action in bone are poorly defined. Fibroblast growth factor receptor-1 (FGFR1) is also essential for bone formation. Fgfr1 expression and activity are positively regulated by T3 in osteoblasts, and in mice that harbor a dominant negative PV mutation targeted to TRα1 or TRβ, Fgfr1 expression is sensitive to skeletal thyroid status. To investigate mechanisms underlying T3 regulation of FGFR1, we obtained primary calvarial osteoblasts from wild-type and TRβPV/PV littermate mice. T3 treatment increased Fgfr1 expression 2-fold in wild-type cells, but 8-fold in TRβPV/PV osteoblasts. The 4-fold increased T3 sensitivity of TRβPV/PV osteoblasts was associated with a markedly increased ratio of TRα1:TRβ1 expression that resulted from reduced TRβ1 expression in TRβPV/PV osteoblasts compared with wild-type. Bioinformatics and gel shift studies, and mutational analysis, identified a specific TR binding site 279–264 nucleotides upstream of the murine Fgfr1 promoter transcription start site. Transient transfection analysis of a series of Fgfr1 promoter 5′-deletion constructs, of a mutant reporter construct, and a series of heterologous promoter constructs, confirmed that this region of the promoter mediates a TR-dependent transcriptional response to T3. Thus, in addition to indirect regulation of FGFR1 expression by T3 reported previously, T3 also activates the Fgfr1 promoter directly via a thyroid hormone response element located at positions −279/−264.

Avian MyoD and c-Jun Coordinately Induce Transcriptional Activity of the 3,5,3′-Triiodothyronine Nuclear Receptor c-ErbAα1 in Proliferating Myoblasts

Although physical interactions with other receptors have been reported, heterodimeric complexes of T3 nuclear receptors (TR) with retinoid X receptors (RXRs) are considered as major regulators of T3 target gene expression. However, despite the potent T3 influence in proliferating myoblasts, RXR isoforms are not expressed during proliferation, raising the question of the nature of the complex involved in TRα transcriptional activity. We have previously established that c-Jun induces TRα1 transcriptional activity in proliferating myoblasts not expressing RXR. This regulation is specific to the muscle lineage, suggesting the involvement of a muscle-specific factor. In this study, we found that MyoD expression in HeLa cells stimulates TRα1 activity, an influence potentiated by c-Jun coexpression. Similarly, in the absence of RXR, MyoD or c-Jun overexpression in myoblasts induces TRα1 transcriptional activity through a direct repeat 4 or an inverted palindrome 6 thyroid hormone response element. The highest rate of activity was recorded when c-Jun and MyoD were coexpressed. Using c-Jun-negative dominants, we established that MyoD influence on TRα1 activity needs c-Jun functionality. Furthermore, we demonstrated that TRα1 and MyoD physically interact in the hinge region of the receptor and the transactivation and basic helix loop helix domains of MyoD. RXR expression (spontaneously occurring at the onset of myoblast differentiation) in proliferating myoblasts abrogates these interactions. These data suggest that in the absence of RXR, TRα1 transcriptional activity in myoblasts is mediated through a complex including MyoD and c-Jun.