Figure 7

NPDC-1 selectively represses nuclear receptor-mediated transcription. A. rNPDC-1 was analyzed for its ability to modulate the transcription mediated by endogenous E2F transcription factors. A pBL-E2FRE-tk-luc (E2FRE) reporter plasmid and a mammalian expression β-galactosidase construct were transfected into HepG2 cells with and without a mammalian expression construct for rNPDC-1. Luciferase data was normalized as described above and data was reported as a % of the constitutive activation of the E2F reporter plasmid. B. Mammalian expression plasmids for pRS-hRXRα, pRS-hRARβ, pRS-rPPARα, pRS-hERα and pRS-hVDR were transiently transfected into HEK 293 cells along with their respective luciferase reporter constructs {hRXRα: pBL-CRBP2, hRARβ: pBL-βRE(2), rPPARα: pBL-AOXRE, hERα: pBL-ERE, and hVDR: pBL-VDRE(3)} and with or without a mammalian expression construct for rNPDC-1. Transfected cells were incubated with appropriate ligands at indicated doses. A mammalian expression β-galactosidase construct was included in all transfections to normalize for transfection efficiency. Multiple data points were collected and calculated as average induced luciferase activity normalized to constitutive βGal activity {normalized response = average full integral luciferase activity/ [average (βGal activity/min)]}. Data is expressed here as a percentage of the normalized response for the ligand induced receptor-reporter constructs. Error bars represent the value of standard deviations n = 4.