FIG. 4.

LMP1^TRADD fails to suppress the expression of p16^Ink4a. (A) Cell lysates were prepared from MEF infected with various retroviruses at the indicated passages and examined for the expression of LMP1, p16^Ink4a, and p21^Waf1 by immunoblotting with specific antibodies as previously described (39). (B) p16 promoter reporter assay. REF52 rat embryonic fibroblasts were cotransfected with 5 ng of pRL-SV40 reporter, 100 ng of pGL2-p16 promoter reporter (−1214 to −1), and 1,000 ng of the indicated LMP1 constructs by FuGENE 6 (GIBCO). pGL2-basic plasmid was used as a control in the reporter assay. Cells were harvested 48 h after transfection, and a dual luciferase assay was performed (Promega). The results shown here are representative of at least three separate experiments performed in triplicate. The error bars represent calculated standard deviations.