Fig 1: Identification of betulinic acid (BA) as a Skp2 inhibitor. A, Plant-derived compounds were docked to the binding pocket of Skp2-Skp1. Skp2 and compounds are shown as cartoon and line, respectively. B, Co-immunoprecipitation (co-IP) experiments were performed to examine the effects of four selected compounds and SZL-P1-41 on Skp2-Skp1 binding in 293T cells transfected with Skp2 and Flag-Skp1. C, Endogenous Skp2-Skp1 interactions with or without BA in H1299 cells were examined by co-IP. D, 293T cells were transfected with Flag-Skp1 to examine the Skp2-Skp1 interaction with or without BA treatment. E, Ubiquitination assay of exogenous p27 in 293T cells transfected with Myc-p27, HA-Ub, and Skp2 upon BA or SZL-P1-41 treatment. F, Ubiquitination assay of endogenous p27 in A549 cells upon BA or SZL-P1-41 treatment. The asterisk indicates the bands of IgG light chain. G, Ubiquitination assay of exogenous E-cadherin in 293T cells transfected with Flag-E-cadherin, HA-Ub, and Skp2 upon BA treatment. H, Ubiquitination assay of endogenous E-cadherin in A549 cells upon BA treatment. I, J, A cellular thermal shift assay (CETSA) of Skp2 was conducted by western blotting (I); quantification of the cellular thermal curve shift of Skp2 protein (J)