Fig. 3

K42 is crucial for the degradation of SOX2 induced by the MLL1/WDR5 complexes. a Sequence alignment of SOX2 peptides, histone H3 N-terminal peptide, and the MLL1 Win peptide. b K42R mutant was non-sensitive to the MLL1/WDR5 methyltransferase complexes. Candidate lysines in SOX2 were mutated to arginine. The wild-type SOX2 and mutants were tagged with an N-terminus Flag, and stably expressed in PA-1 cells through retroviral gene transfer and expression system. Cells were harvested after knockdown of MLL1 or WDR5 for 48 h, and proteins of interest were analyzed by western blotting. The relative protein levels of SOX2 were densitometry measured and plotted, which were normalized to tubulin. c Forced expression of WDR5 promotes the ubiquitination of SOX2. GFP-SOX2 was stably expressed in 293 cells. The cells were co-transfected with pRK5-HA-ubiquitin and pCMV10-3Flag or pCMV10-3Flag-WDR5 respectively. MG-132 was applied to accumulate the ubiquitinated proteins. Co-immunoprecipitation was performed using anti-SOX2 antibody, and the ubiquitination was analyzed by anti-HA antibody. The ubiquitinated SOX2 was densitometry quantified by Gel Image analysis software. *p < 0.05, **p < 0.01, ***p < 0.001. The data were represented as mean ± SD