Fig. 4: RUNX2 disrupts FBXW7-mediated c-Myc ubiquitination and degradation.
From: Overexpression of RUNX2 promotes breast cancer multi-organ metastasis through stabilizing c-Myc
A RUNX2 and/or FBXW7 was knocked down with lentiviral infection in MDA-MB-231 and 4T1 cells. The mRNA and protein levels of c-Myc, FBXW7, and RUNX2 in the indicated cells were detected by RT–qPCR and immunoblot. B c-Myc ubiquitination levels in the indicated cells were detected by IP and immunoblot following MG-132 treatment. C mRNA levels of c-Myc target genes in the indicated cells were detected by RT–qPCR. D HA-tagged c-Myc and a gradient dose of Flag-tagged RUNX2 or vector control were co-transfected into 293T cells. The protein levels of RUNX2 and FBXW7 interacting with c-Myc were detected by Co-IP and immunoblot. E Density maps for ChIP–seq (GSE190248) and CUT&RUN–seq (GSE95303) data showing c-Myc target genes genomic locus enriched by c-Myc and RUNX2 in MDA-MB-231 cells. The genome browser map is displayed using the IGV software. F Enrichment of c-Myc and RUNX2 at the promoter regions of c-Myc target genes in T-47D cells with RUNX2 overexpression and MDA-MB-231 cells was detected by Re-ChIP–qPCR analysis. Data are shown as the mean ± SD. Statistical analyses were performed with the unpaired Student’s t-test. *P < 0.05 compared with control cells. #P < 0.05 shFBXW7 plus shRUNX2 compared with shRUNX2 cells, or shFBXW7 plus shRUNX2 compared with shFBXW7 cells. IP immunoprecipitation, ChIP–seq chromatin immunoprecipitation–sequencing.
