FIG. 8.
HIF antagonizes c-Myc transcriptional activity. (A) Representative quantitative real-time PCR analysis for Pgk, Vegf, c-Myc, p21, and p27 mRNA levels. Left, higher normoxic expression levels of Pgk, Vegf, p21, and p27 in Vhl−/− MEFs compared to that of wild-type controls. Right, hypoxia greatly induced the expression of Pgk, Vegf, p21, and p27 in Vhl+/+ MEFs, while no induction of the genes was observed in Vhl−/− MEFs. Numbers indicate induction (n-fold) under hypoxia. Reactions were performed in triplicate and normalized to endogenous 18S RNA expression. (B) Myc transcriptional activity was reduced in Vhl−/− MEFs as determined by c-Myc reporter assays. Assays were performed in triplicate. (C) HIF transcriptional activity was elevated under conditions that repressed c-Myc activity. Hypoxia increased luciferase expression in Vhl+/+ MEFs. However, luciferase expression was elevated in Vhl−/− MEFs under normoxia and not further induced by hypoxia. Regulation was restored by cotransfection of a plasmid encoding hemagglutinin-VHL. Cells were exposed to hypoxia as defined as 0.5% O2 for 48 h.