Research Paper Volume 12, Issue 8 pp 6644—6666

Figure 7. miR-22/miR-142 targets the key autophagy proteins ATG5, ATG4A and ULK1. (A) Predicted miR-142 binding sites in the ULK1, ATG4A, and ATG5 3'UTR. Wild-type and mutant-type ULK1, ATG4A, and ATG5 3'UTR reporter vectors containing wild-type or mutant-type miR-142 binding sites were constructed. (B–D) These vectors were co-transfected into HEK293 cells with miR-142 mimics or inhibitor, and the luciferase activity was determined. (E) Predicted miR-22 binding sites in ULK1. Wild-type and mutant-type ULK1 3'UTR reporter vectors containing wild- or mutant-type miR-22 binding sites were constructed. (F) These vectors were cotransfected into HEK293 cells with miR-22 mimics or inhibitor, and the luciferase activity was determined. (G, H) U2OS cells were transfected with miR-142 mimics in the presence or absence of Dox and examined for the protein levels of ULK1, ATG4A, and ATG5 were examined. (I, J) U2OS cells were transfected with miR-22 mimics in the presence or absence of Dox (5 μM), and the protein levels of ULK1 were examined. The data are presented as the mean ± SD of three independent experiments. *P<0.05, **P<0.01, compared to the control group; ##P<0.01, compared to the Dox group.