Fig. 8. TRAF7 negatively regulates the expression of HOXA5 through ubiquitin-mediated degradation.

A TRAF7 expression in PCa tissues and normal tissues were obtained from the Gene Set Cancer analysis database. B TRAF7 mRNA level in PCa tissues and paired adjacent normal tissues. C Western blot analysis of TRAF7 and HOXA5 expression in PCa cells with TRAF7 overexpression. D Endogenous co-immunoprecipitation was performed to determine the interaction between TRAF7 and HOXA5 in PCa cells. TRAF7 was immunoprecipitated with a HOXA5 antibody and vice versa. E HOXA5 protein levels were determined after incubation with cycloheximide (CHX) for the indicated time periods in PC-3 and VCaP cells following TRAF7 overexpression. F PC-3 and VCaP cells were incubated with MG132 for 4 h. Cell lysates were immunoprecipitated with HOXA5 antibody and then immunoblotted with ubiquitin antibody. The increased cell proliferation (G), cell invasion (H), phospho-ERK expression (I), and percentage of CD133+ stem cell subpopulation (J) caused by TRAF7 overexpression were significantly inhibited by the ectopic HOXA5 expression. *P < 0.05, **P < 0.01. ^#P < 0.05, ^##P < 0.01. Results were presented as mean ± SD.