TR3 orphan nuclear receptor mediates apoptosis through up-regulating E2F1 in human prostate cancer LNCaP cells

Early studies suggested both TR3 orphan receptor (TR3) and apoptosis mediator E2F1 might play an important role in mediating prostate cancer cell apoptosis. Their linkage and relationship, however, remain unclear. Here we found that 12-O-tetradecanoylphorbol-13-acetate (TPA) could induce cell apoptosis via induction of TR3 and E2F1 expression in LNCaP prostate cancer cells. Addition of antisense E2F1 could partially rescue the TR3-mediated cell apoptosis, and transfection of the TR3 dominant-negative plasmid could block the TR3-induced E2F1 expression. These data suggest that TPA is able to induce LNCaP cell apoptosis via induction of TR3 resulting in the induction of E2F1. Promoter reporter assays show that TR3 can induce E2F1 expression via binding to the TR3 response element (TR3RE) in the E2F1 promoter -316 to -324 bp region. TR3 can bind specifically to this TR3RE with a Kd of 6.29 nm, and mutations of this E2F1-TR3RE can partially block the TR3-mediated E2F1 expression. Taken together, these data suggest that TPA is able to induce cell apoptosis via a TPA --> TR3 --> E2F1 --> apoptosis pathway in LNCaP cells. Further studies of how to modulate this pathway may allow us to better understand how to control the prostate cancer growth.     

E2F and p53 induce apoptosis independently during Drosophila development but intersect in the context of DNA damage

In mammalian cells, RB/E2F and p53 are intimately connected, and crosstalk between these pathways is critical for the induction of cell cycle arrest or cell death in response to cellular stresses. Here we have investigated the genetic interactions between RBF/E2F and p53 pathways during Drosophila development. Unexpectedly, we find that the pro-apoptotic activities of E2F and p53 are independent of one another when examined in the context of Drosophila development: apoptosis induced by the deregulation of dE2F1, or by the overexpression of dE2F1, is unaffected by the elimination of dp53; conversely, dp53-induced phenotypes are unaffected by the elimination of dE2F activity. However, dE2F and dp53 converge in the context of a DNA damage response. Both dE2F1/dDP and dp53 are required for DNA damage-induced cell death, and the analysis of rbf1 mutant eye discs indicates that dE2F1/dDP and dp53 cooperatively promote cell death in irradiated discs. In this context, the further deregulation in the expression of pro-apoptotic genes generates an additional sensitivity to apoptosis that requires both dE2F/dDP and dp53 activity. This sensitivity differs from DNA damage-induced apoptosis in wild-type discs (and from dE2F/dDP-induced apoptosis in un-irradiated rbf1 mutant eye discs) by being dependent on both hid and reaper. These results show that pro-apoptotic activities of dE2F1 and dp53 are surprisingly separable: dp53 is required for dE2F-dependent apoptosis in the response to DNA damage, but it is not required for dE2F-dependent apoptosis caused simply by the inactivation of rbf1.     

Methylation-mediated regulation of E2F1 in DNA damage-induced cell death

E2F1 promotes DNA damage-induced apoptosis and the post-translational modifications of E2F1 play an important role in the regulation of E2F1-mediated cell death. Here, we found that Set9 and LSD1 regulate E2F1-mediated apoptosis upon DNA damage. Set9 methylates E2F1 at lysine 185, a conserved residue in the DNA-binding domain of E2F family proteins. The methylation of E2F1 by Set9 leads to the stabilization of E2F1 and up-regulation of its proapoptotic target genes p73 and Bim, and thereby induces E2F1-mediated apoptosis in response to genotoxic agents. We also found that LSD1 demethylates E2F1 at lysine 185 and reduces E2F1-mediated cell death. The identification of the methylation/demethylation of E2F1 by Set9/LSD1 suggests that E2F1 is dynamically regulated by epigenetic enzymes in response to DNA damage.     

Methylation-mediated regulation of E2F1 in DNA damage-induced cell death

E2F1 promotes DNA damage-induced apoptosis and the post-translational modifications of E2F1 play an important role in the regulation of E2F1-mediated cell death. Here, we found that Set9 and LSD1 regulate E2F1-mediated apoptosis upon DNA damage. Set9 methylates E2F1 at lysine 185, a conserved residue in the DNA-binding domain of E2F family proteins. The methylation of E2F1 by Set9 leads to the stabilization of E2F1 and up-regulation of its proapoptotic target genes p73 and Bim, and thereby induces E2F1-mediated apoptosis in response to genotoxic agents. We also found that LSD1 demethylates E2F1 at lysine 185 and reduces E2F1-mediated cell death. The identification of the methylation/demethylation of E2F1 by Set9/LSD1 suggests that E2F1 is dynamically regulated by epigenetic enzymes in response to DNA damage.