FIGURE 5.

Phosphorylation by UL97 relieves Rb-mediated repression of E2F-responsive promoters. A, Saos-2 cells were transfected with a luciferase reporter driven by the E2F1 promoter together with an empty vector (−) or an expression plasmid for Rb and the indicated allele of UL97. Lysates harvested 48 h after transfection were analyzed for luciferase activity (top) and protein expression with the indicated antibodies (bottom). Luciferase activity was normalized to total protein concentration and is presented relative to the activity of the reporter without Rb or UL97 (set at 100%). Error bars denote the S.D. *, p < 0.05; **, p < 0.01; n.s., not significant. B, luciferase and Western blot analyses were performed as in A except an Rb allele in which CDK consensus phosphorylation residues were replaced with alanines (RbΔCDK;Δ) was also included. C, luciferase and Western blot analyses were performed as in A except the reporter contained an E2F1 promoter in which the E2F binding sites were mutated. D, luciferase and Western blot analyses were performed as in A except the reporter contained the Orc1 promoter. E, luciferase and Western blot analyses were performed as in A except the reporter contained the cyclin A promoter. F, luciferase and Western blot analyses were performed as in A except an Rb allele with an N757F CM was also included. KD, kinase-deficient.