Fig. 7

BIG promotes protein-protein interactions. A, E, J, Q Sublocalizations of Bcl-2 with Beclin-1, Bcl-2 with CH25H, Bcl-2 with c-MYC, and CH25H with RORɑ were analysed by immunofluorescence assay. Laser confocal microscopy images confirmed that Bcl-2 and Beclin-1, Bcl-2 and CH25H, Bcl-2 and c-MYC, and CH25H with RORɑ were colocalized with the nucleus and in the cell matrix. Chondrocytes pretreated with BIG showed a significantly enhanced fluorescence intensity. The scale bars represent 20 μm. B–D Chondrocyte lysates were prepared by IP lysis and coimmunoprecipitated with Bcl-2 antibody and IgG as a negative control. The results show the interaction between Bcl-2 with Beclin-1 and c-Myc. F–I The coimmunoprecipitation results similarly show the CH25H with RORɑ interaction, CH25H with RORɑ and Bcl-2. K–M, P Using c-MYC, Beclin-1, and CH25H as bait proteins for coimmunoprecipitation, the results confirm that Bcl-2 interacted with c-MYC, Beclin-1, and CH25H. O The chemical-molecular structure map of BIG (red: oxygen; green: carbon). N Ribbon diagram of the monomeric c-MYC catalytic domain complexed with BIG (pink: the c-MYC active domain). Docking of BIG into the c-MYC catalytic pocket (the map shows the details of molecular docking). Data are presented as the mean ± SEM (n = 3) and were analysed by the Kruskal‒Wallis test, *P < 0.05, **P < 0.01, ***P < 0.001