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Figure 4


Fig. 4. Overexpression of GNL3L inhibits transcriptional activity of ERR proteins independently of nucleolar distribution. (A1) Estrogen response element (ERE)-specific transcriptional activities were measured in CV-1 cells by the ratio between the ERE-driven firefly luciferase activity and the Renilla-null luciferase activity. ERR{gamma} elicits a six-fold increase in the ERE-specific transcriptional activity. Coexpression of wild-type GNL3L (WT) leads to a 50% reduction in the ERR{gamma}-mediated transcriptional activity. This decrease is reversed by a deletion of the ERR{gamma}-binding I-domain of GNL3L (dI). Coexpression of either the nucleolar form (NoG3l) or the nucleoplasmic form (dBC) of GNL3L suppresses the ERR{gamma} transcriptional activity more than or to the same extent as the wild-type GNL3L protein. (B1,C1) Using the same approach, we show that this inhibitory activity of GNL3L can also work on (B1) ERRbeta and (C1) ERR{alpha} with the exception that the dBC mutant has little effect on the ERR{alpha}-mediated transactivation. Error bars represent the standard error of the mean (± s.e.m.). ***P<0.0001. (A2,B2,C2) Expression levels of wild-type and mutant GNL3L proteins and ERR proteins in the experimental samples are compared in western blots side-by-side using anti-HA and anti-Myc antibodies, respectively; {alpha}-tubulin ({alpha}-Tub) was used as a loading control. (D) GNL3L fails to suppress the estradiol (E2)-induced transcriptional activity of ER{alpha} on the ERE-driven promoter in the same cell-based reporter system.





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