Fig. 4
SENP6-mediated ANXA1 de-SUMOylation inhibits IKKα protein degradation. A Ni2+-NTA agarose affinity pull-down assay results representing ANXA1 SUMOylation in HEK293T cells transfected with plasmids expressing SENP6 or sh. SENP6. B Coimmunoprecipitation (co-IP) assay showing the interaction of ANXA1 with IKKα and NBR1 in primary cultured microglia infected with adenovirus encoding vector or SENP6. C Co-IP assay showing the interaction of ANXA1 with IKKα and NBR1 in primary cultured microglia infected with adenovirus encoding sh. NC or sh. SENP6. D Primary cultured microglia were infected with adenovirus expressing SENP6 or vector. The interaction between IKKα and NBR1 in microglia was evaluated by a co-IP assay. E A Co-IP assay was performed to explore the interaction of IKKα and NBR1 in microglia infected with sh. NC or sh. SENP6. F RT–qPCR assay results show the Chuk mRNA levels in microglia infected with adenovirus encoding vector or SENP6. G RT–qPCR shows the Chuk mRNA levels in microglia infected with adenovirus encoding sh. NC or sh. SENP6. H Sh. SENP6 cells reduce the half-life of ectopically expressed IKKα in primary microglia. I Quantitative analysis of the half-life of ectopically expressed IKKα in I. J Effects of increasing the dose of SENP6 on ANXA1-mediated degradation of endogenous IKKα. Data are presented as the mean ± S.E.M. from at least three dependent experiments and analysed by one-way ANOVA followed by Tukey’s post-hoc test (F and G) or repeated-measures (RM) ANOVA followed by Tukey’s post-hoc test (I). ns for P > 0.05, **P < 0.01