A: TAF-Iα and Set/TAF-Iβ repress GRα-induced transcriptional activity of the MMTV promoter stably incorporated in host cell chromatin in HCT116/MMTV cells.
HCT116/MMTV cells were transfected with the indicated amounts of the TAF-Iα-or Set/TAF-Iβ-expressing plasmid, together with pRShGRα and pSV40-β-Gal.
Left panel: Dose effects of TAF-Iα and Set/TAF-Iβ are shown. Bars represent mean ± S.E. values of luciferase activity normalized for β-galactosidase activity in the absence or presence of 10−6 M of dexamethasone (Dex). *: p<0.01 (ANOVA, followed by Bonferroni correction), compared to the baseline (the value obtained in the absence of TAF-Iα and Set/TAF-Iβ, but in the presence of dexamethasone).
Right panel: Set/TAF-Iβ shifts downward the dexamethasone titration curve of luciferase activity from the chromatin-integrated MMTV promoter. Circles represent mean ± S.E. values of luciferase activity normalized for β-galactosidase activity in the indicated concentrations of dexamethasone (Dex). *: p<0.01 (ANOVA, followed by Bonferroni correction), obtained by comparing the results in the absence and presence of Set/TAF-Iβ transfection.
B: Endogenous Set/TAF-Iβ is a negative regulator of GR-induced transcriptional activity in HTC cells.
HTC cells were transfected with Set/TAF-Iβ siRNA or control siRNA. The cells were stimulated with 10−6M of dexamethasone (Dex). Bars indicate the mean ± S.E. of the tyrosine aminotransferase (TAT) activity, mRNA levels of TAT or those of Set/TAF-Iβ. *: p<0.01 (unpaired two-tailed Student t test), by comparing the two values indicated.
C: Set/TAF-Iβ represses the transcriptional activity of the mineralocorticoid (MR) and progesterone receptor (PR)-A in addition to that of the GR.
HCT116 cells were transfected with Set/TAF-Iβ-expressing plasmid together with GRα-, MR- or PR-A-expressing plasmid, pMMTV-Luc or (κB)3-Luc, and pSV40-β-Gal. Bars represent mean ± S.E. values of luciferase activity normalized for β-galactosidase activity in the absence or presence of 10−6 M of dexamethasone (Dex) or progesterone (Prog), 10−8 M of aldosterone (Aldo), or p65/p50 transfection. *: p<0.01, n.s.: not significant (unpaired two-tailed Student t test), compared to the baseline (“(-)” in the presence of ligand or p65/p50).