Fig. 4: ASB10 stabilizes HSP70 via competitively inhibiting STUB1-mediated ubiquitination of HSP70.

A Protein expression of HSP70 after overexpressing Asb10 in NRVMs as determined by western blot. The right panel showed the quantifications. N = 6. B Protein expression of HSP70 after overexpressing ASB10 in 293 T cells as determined by western blot. The right panel showed the quantifications. N = 6. C Relative expressions of Asb10 and Hspa1 in NRVMs overexpressing Asb10 as determined by qPCR. N = 5. D Expression changes of HSP70 in NRVMs overexpressing Asb10 and treated with different time courses of CHX. The right panel showed the quantifications. N = 3. E Expression changes of HSP70 in 293T cells overexpressing ASB10 and treated with different time courses of CHX. The right panel showed the quantifications. N = 3. F Expression changes of HSP70 in NRVMs overexpressing Asb10 and treated with CQ or MG132 for 6 h. The right panel showed the quantifications. N = 3. G Co-immunoprecipitation assay to detect the changes of the STUB1/HSP70 complex and HSP70 ubiquitination level in NRVMs overexpressing Asb10. N = 3. H Co-immunoprecipitation assay to detect the changes of the STUB1/HSP70 complex and HSP70 ubiquitination level in NRVMs transfected with STUB1 siRNA and Ad-Asb10. N = 3. I Co-immunoprecipitation assay to detect the changes of the STUB1/HSP70 complex and HSP70 ubiquitination level in 293T cells transfected with STUB1-EGFP, ASB10-FLAG, HSPA1-Myc and Ub-HA. N = 3. Unpaired Student’s t-test (A–E) was conducted. One-way ANOVA followed by Turkey’s multiple comparisons test (F) was conducted. Data were represented as Mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, ^nsP > 0.05.