p53 Represses Transcription of RING Finger LIM Domain-Binding Protein RLIM through Sp1
Figure 4
p53 inhibits Sp1-mediated activation of the RLIM promoter.
(A) Sp1 activated the RLIM promoter activity. H1299 cells were cotransfected with 100 ng of NP500-Luc with either 160 ng pCMV empty vector as control or increasing amounts (0, 10, 20, 40, 80 and 160 ng) of pCMV-Myc-Sp1 plasmid. The total amount of plasmid in each transfection was adjusted to be the same using the empty vector. Cells were harvested 30 h after transfection and lysed for measuring luciferase activity. (B) p53 inhibits Sp1-stimulated activity of the RLIM luciferase reporter. H1299 cells were cotransfected with 100 ng of NP500-Luc and increasing amounts (0, 1, 2.5, 5 and 10 ng) of pCMV-HA-p53 plasmid in the presence of a fixed amount (160 ng) of pCMV-Myc-Sp1 plasmid. H1299 cells were transfected with 100 ng NP500-Luc alone as control. Cell lysates were used for luciferase assays as described above. (C) p53 mutants can not inhibit Sp1-stimulated activity of the RLIM luciferase reporter. The indicated p53 wild type or mutant constructs were cotransfected with 100 ng of RLIM promoter reporter construct NP500-Luc and 160 ng pCMV-Myc-Sp1 plasmid. Cells were harvested 30 h after transfection and the cell lysates were prepared and used for luciferase assays as described above.