Fig. 6

Expression of USP28 K99R induces an increase in HIF-1α protein and HIF-1α transcriptional activation. a HCT116 cells were transfected with Flag-tagged vector control, Flag-tagged USP28 WT or Flag-tagged USP28 K99R for 36 h, cells were then lysed and subjected to western blot with indicated antibody. b 293T cells were co-transfected with HA-HIF-1α, pGL3-HRE-Luc, pSV40-renilla and Flag-tagged vector control, Flag-tagged USP28 WT or Flag-tagged USP28 K99R for 36 h, after which the luciferase activities in each group were measured and compared, respectively. c Proposed model for the SENP1–USP28–HIF-1α positive feedback loop. HIF-1α is able to induce the expression of SENP1, which in turn deSUMOylates and stabilizes HIF1a. We show SENP1 also deSUMOylates USP28 to enhance its deubiquitinating activity towards HIF-1α, and thereby preventing FBXW7-dependent degradation of HIF-1α during hypoxia