The Regulation Of P53Pathway By DNAJB1or UXT And Their Potential Effect On Breast Cancer

Objective:P53is an important tumour suppressor. MDM2/MDMX complex is the most essential negative regulator in P53pathway. In order to identify MDM2and MDMX different roles more accurately, Dr.Yuan’s lab first identified several proteins that can bind to MDM2or MDMX through Y2H(Yeast Two Hybird). Considering of the research background and clinical features, we further research DNAJB1which can bind to MDM2AD but not MDMX, as well as UXT which can bind to MDMX but not MDM2.Methods:1. The IP, IF, WB techniques were performed to investigate the interaction of target protein and MDM2/MDMX.2. After inserting target gene into proper plasmid through gene recombination, the target gene was expressed/overexpressed in certain cell lines by transefection. The western blot was employed to verify the regulation of P53pathway by DNAJB1or UXT.3. In order to investigate the effect on P53pathway under the condition of lower protein level, the target gene was konckdowned through RNAi(siRNA or shRNA) in breast cancer cell line MCF7.4. The cellular experiments and Xenograft tumor mouse model were performed to figure out the target gene’s contribution to P53pathway in vivo and in vitro.Results:1. DNAJB1and MDM2are localized in nuclear where DNAJB1can bind to MDM2through its c-terminal and stabilize MDM2at a post-translational level. DNAJB1can inhibit the MDM2-mediated ubiquitination and degradation of P53protein, as well as increase the P21protein level in a P53-dependent manner. The depletion of DNAJB1promotes breast cancer cell line MCF7’s growth in vitro and in vivo.2. UXT can bind to and stabilize MDMX, it can increase the P21protein level in a P53-dependent manner. Overexpressing UXT can suppress the cellular response to DNA damage, ribosomal stress or overexpressed oncogenes. Knocking down UXT can enhance the cell’s response to various stimulations. In vitro experiments, the depletion of UXT suppresses breast cancer cell’s growth, while overexpressing UXT can promote cancer cell’s growth.Conclusions: 1. DNAJB1is a P53-dependent tumor suppressor.2. UXT is a P53-dependent oncogene.