Fig. 4: O-GlcNAcylation of c-Myc promotes its stability.
From: The OGT–c-Myc–PDK2 axis rewires the TCA cycle and promotes colorectal tumor growth
Immunoblotting analysis (A) and quantitative PCR analysis (B) of c-Myc expression in HT-29 and HCT116 cells expressing scramble or shOGT. n = 3; Data are presented as means ± SD. P values were determined by unpaired two-tailed Student’s t tests. C Analysis of c-Myc O-GlcNAcylation upon OSMI-4 or TMG treatment. D Mapping the site of O-GlcNAcylation on c-Myc using mass spectrometry. E Probing the major site of glycosylation on c-Myc using various site-directed mutants. Quantification was shown. n = 3; Data are presented as means ± SD. Statistical analyses were performed by unpaired two-tailed Student’s t tests. F Immunoblotting of c-Myc levels in WT or S415A c-Myc-reconstituted HT-29 cells by CHX treatment in the presence of inhibitors for OGT (OSMI-4) or proteasome (MG132). Quantification was shown. n = 3; Data are presented as means ± SD. P values were determined by unpaired two-tailed Student’s t tests. G Immunoblotting of c-Myc ubiquitination levels in the presence or absence of TMG treatment in WT or S415A c-Myc-reconstituted HT-29 cells. EV, expression of vehicles as a negative control. H Analysis of c-Myc-MAGI3 interaction in 293 T cells overexpressing WT or S415A Flag-tagged c-Myc and HA tag MAGI3 in the presence or absence of OGT overexpression. Immunoblot analyses were performed with the indicated antibodies. I Analysis of c-Myc-MAGI3 interaction in WT or S415A c-Myc-reconstituted HT-29 cells. Immunoblot analyses were performed with the indicated antibodies.
