ATPR-induced differentiation and G0/G1 phase arrest in acute promyelocytic leukemia by repressing EBP50/NCF1 complex to promote the production of ROS.

Our previous study has demonstrated that 4-amino-2-trifluoromethyl-phenyl Retinate (ATPR) can induce human leukemia NB4 cells differentiation and G0/G1 phase arrest, but the underlying mechanism is still unclear. In this study, we used proteomics to screen differentially expressed protein profiles in NB4 cells before and after ATPR treatment in vitro. We analyzed the peptides digested from total cellular proteins by reverse phase LC-MS/MS and then performed label-free quantitative analysis. We found 27 significantly up-regulated proteins in the ATPR group compared to the control group. NCF1 was the most significantly changed protein. Immunoprecipitation and double immunofluorescent staining showed that EBP50 bind to NCF1. We further explored the potential molecular mechanism of EBP50/NCF1 complex in ATPR-induced differentiation and G0/G1 phase arrest. The results showed that ATPR remarkably reduced the expression of EBP50 in vivo and in vitro. Interestingly, the reduction of EBP50 contributed to ROS release by modulating the subcellular localization of NCF1. The reduction of EBP50 also contributed to G0/G1 phase arrest by inhibiting CyclinD1, CyclinA2 and CDK4, as well as promoting the differentiation of NB4 cells by increasing the expression of CD11b. Furthermore, we found that the overexpression of EBP50 restrained the effects of ATPR on differentiation and G0/G1 phase arrest in NB4 cells. These results suggest that ATPR-induced differentiation and G0/G1 phase arrest in acute promyelocytic leukemia (APL) by repressing EBP50/NCF1 complex to promote the production of ROS, and the results from in vivo experiments were consistent with those from in vitro studies. Therefore, our finding results suggest that EBP50 may be a new target for ATPR in the treatment of APL.