Fig. 5: USP10 stabilizes RUNX1 through deubiquitination.
From: USP10 deubiquitinates RUNX1 and promotes proneural-to-mesenchymal transition in glioblastoma
A Expression of USP10 and RUNX1 in LN229 and GBM2 cells after different treatments. B Western blot showing the expression of indicated proteins in LN229 and GBM2 cells after indicated treatments. C Expression of indicated proteins in U251 and GBM1 cells after different treatments. D LN229 and GBM2 cells were co-transfected with HA-Ub, ShCtrl or shUSP10 and cell lysates subjected to IP with RUNX1 antibody, followed by IB with antibodies against HA and RUNX1. Cells were treated with 20 μM MG132 for 8 h. E U251 and GBM1 cells were co-transfected with His-RUNX1, HA-Ub, and Flag-USP10 WT or USP10 C424A and cell lysates were subjected to IP with His antibody, followed by IB with indicated antibodies. Cells were treated with 20 μM MG132 for 8 h. F Unubiquitinated or ubiquitinated His-RUNX1 was incubated with GST-USP10 WT or GST-USP10 C424A coupled to glutathione-Sepharose beads. Proteins retained on Sepharose were then subjected to IB with indicated antibodies. Recombinant GST-USP10 WT or GST-USP10 C424A was analyzed by SDS-PAGE and Coomassie blue staining. U251 (G) and GBM1 (H) cells were co-transfected with His-RUNX1, Flag-USP10, and the indicated HA-Ub Lys0, Lys48-only, or Lys63-only plasmids, RUNX1 ubiquitylation linkage was analyzed. I LN229 and GBM2 cells co-transfected with Ub-WT or Ub-Lys48R and shCtrl or shUSP10, RUNX1 levels were analyzed.
