Sabilization Of The UHRF2 Protein By HBx Promotes Hepatocarcinogenesis

ObjectiveThe purpose of this study was to verify the expression of UHRF2 in Hepatitis B virus(HBV)positive Hepatocellular carcinoma(HCC),and analyze its relationship with the prognosis of HBV positive HCC patients,and to further clarify UHRF2 through HBV-HBx-Ets1-CDK2-UHRF2 pathway promotes the molecular mechanism of hepatocellular carcinoma.Based on this study,the feasibility of UHRF2 as a biomarker and potential therapeutic target for HCC was explored,which provided a theoretical basis for the early diagnosis and prognosis of HCC patients.MethodsWestern blot(WB)was used to detect the expression of UHRF2 in 20 cases of HBV-positive HCC tissues and adjacent tissues;Immunohistochemistry(Immunohistonchemistry,IHC)was used to verify the expression of UHRF2 in 87 cases of HBV-positive HCC tissues and adjacent tissues.Circumstances;WB test method was used to verify the effect of HBV on UHRF2 in four HCC cells,HepG2,HepG2.2.25,Hep AD38 and HepG2-NTCP.UHRF2 was knocked down and overexpressed in Hep AD38 and HepG2.2.25 cells,respectively.Real-time quantitativePCR(qRT-PCR)and WB technology were used to detect the knockdown and overexpression efficiency of UHRF2 gene.The scratch test,Transwell test and EdU(5-Ethynyl-2’-deoxyuridine)test were used to detect the migration,invasion and proliferation ability of HBV-positive HCC cells,respectively.Mechanistically,the WB experiment method,Co-Immunoprecipitation(Co-IP)experiment technology and Immunofluorescence(IF)experiment were used to verify the HBV x protein(virus transactivator x,HBx),CDK2(Cyclin-dependent kinases),the relationship between UHRF2 and Ets1(c-Ets-1).ResultsWB and immunohistochemistry showed that UHRF2 was significantly overexpressed in HBV-positive HCC,and it was determined that its high expression was negatively correlated with the prognosis of 87HBV-positive HCC patients.The results of scratch test,Transwell test and EdU test showed that overexpression of UHRF2 in HBV-positive HCC cells can promote their migration,invasion and proliferation;knockdown of UHRF2 inhibits the migration,invasion and proliferation of HBV-positive HCC cells.WB experiments,co-immunoprecipitation experiments and immunofluorescence results show that HBx up-regulates CDK2 via Ets1 protein;CDK2 interacts with UHRF2 and activates the phosphorylation of the 643 serine residue of UHRF2.Prevent the degradation of UHRF2 to maintain the stability of UHRF2,and then increase UHRF2.Conclusion(1)UHRF2 protein is up-regulated in HBV-positive HCC tissues,and its expression level is negatively correlated with the prognosis of HCCpatients,revealing that UHRF2 may be involved in the occurrence and development of HCC and other processes.(2)Using an in vitro HBV-positive HCC cell carcinoma model,it was found that overexpression of UHRF2 can promote cell migration,invasion and value-added,suggesting that overexpression of UHRF2 can play a cancer-promoting role in HCC.(3)UHRF2 may promote the development of HCC through the HBV-HBx-Ets1-CDK2-UHRF2 pathway,revealing the specific mechanism by which UHRF2 may promote the development of HCC.