Fig. 6: HOIL-1 inhibits the degradation of HBx via inhibiting K48 ubiquitination. | Experimental & Molecular Medicine

Fig. 6: HOIL-1 inhibits the degradation of HBx via inhibiting K48 ubiquitination.

From: The LUBAC subunit HOIL-1 promotes the progression of HBV-associated hepatocellular carcinoma independently of linear ubiquitination

Fig. 6

a The Co-IP and blotting of anti-HOIL-1 or anti-Flag in HepG2 cells transduced with HBx-Flag, LHBs-Flag, MHBs-Flag, SHBs-Flag and HBc-Flag plasmids, respectively. The red star symbol indicates specific bands. b The Co-IP and blotting of anti-Flag in HEK293T cells transduced with HOIL-1-HA plasmids combined with HBx-Flag, LHBs-Flag, MHBs-Flag, SHBs-Flag and HBc-Flag plasmids, respectively. The red star symbol indicates specific bands. c The Co-IP and blotting of anti-HOIL-1 or anti-HBx in HepG2.2.15 cells. d The western blot analysis of indicated proteins in the HOIL-1-knockdown and control cells. e The GSEA for HOIL-1 expression in HBx-related gene signatures (WU_HBx_Target_2_UP, WU_HBx_Target_3_UP) in the GSE14520 cohort. f The western blot of HBx in HepG2.2.15 cells incubated with cycloheximide at different time points. g The western blot of the indicated tag protein detected in HepG2 cells with exogenous HBx-plasmid transfection and incubated with cycloheximide at different time points. h The Co-IP for the K48-linked ubiquitination of HBx in the HepG2.2.15 cells after 8 h of MG132 treatment (10 μM). i, j The effect of the HBx knockdown on the cell proliferation of HepG2.2.15 cells transfected with LV-HOIL-1 or LV-Ctrl detected by CCK-8 (i) and EdU incorporation (j) assays. k The CCK-8 assays detected the effect of HBx overexpression on proliferation of HepG2.2.15 cells. l The representative images of IHC staining of HOIL-1 and HBx in HCC tissues and the correlation between HOIL-1 and HBx levels, n = 10. m The correlation between HOIL-1 and HBx mRNA expression in the TCGA LIHC cohort. n The Kaplan–Meier analysis for the OS and DFS of two groups of patients with HCC defined by low and high coexpression of HOIL-1 and HBx in the TCGA LIHC cohort. The Mann–Whitney U test was used in f, g and i–k, Spearman’s rank correlation test in l and the log-rank Mantel–Cox test in n. Data are shown as mean ± s.d., *P < 0.05, **P < 0.01. CHX cycloheximide, Co-IP Co-immunoprecipitation assays, IB immunoblot, IF immunofluorescence, IP immunoprecipitated, MG132 N- (benzyloxycarbonyl) leucinylleucinylleucina.

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