FIG. 7.
Effect of AMPK-expressing adenoviruses on the association of HuR with target transcripts. EMSA were performed using radiolabeled RNAs encoding the 3′ UTRs of p21, cyclin A, and cyclin B1 genes (see Materials and Methods) and proteins present in cytoplasmic lysates of RKO cells that were infected with either AdGFP, Ad(CA)AMPK, or Ad(DN)AMPK and then were either exposed to 20-J/m2 UVC or left unirradiated and collected 3 h later. The presence of HuR in RNA-protein complexes was assayed by monitoring the formation of supershifted bands in the presence of anti-HuR antibodies (+HuR ab) or control antibodies (+p38 ab). Arrowheads, supershifted complexes. Fold, difference in total signals of radiolabeled complexes between indicated cells and AdGFP-infected, unirradiated cells. Fold differences were not calculated for supershift lanes. f, free probe, not incubated with cytoplasmic lysate; high intensity, supershifted bands developed at greater intensity.