Fig. 4

bta-miR-221 Targets and Suppresses ATG7 Expression. A, bioinformatics tools were used to predict the interaction site between bta-miR-221 and the ATG7 3’UTR. B, Effect of bta-miR-221 on ATG7 expression. HEK-293T cells were cotransfected with either wild-type or mutant ATG7-3’UTR-psiCHECK2 plasmids along with bta-miR-221 mimics or inhibitors, followed by luciferase assay measurements 24 h posttransfection to assess regulatory impacts. C, Details of the experiment in MDBK cells, where transfection with bta-miR-221 mimics or inhibitors (along with their respective negative controls) preceded qPCR analysis at 12 and 24 h to quantify ATG7 mRNA levels. D and E, The influence of bta-miR-221 on ATG7 protein levels was tracked, and the intracellular concentration of mCherry-ATG7 protein in HEK-293T cells 24 h after cotransfection with bta-miR-221 mimics or inhibitors was determined