The Study On The Role Of Wnt/?-Catenin Signaling Pathways In The Replication Of Avian Leukosis Virus Subgroup J

Wnt signaling pathway is a signaling pathway that remains relatively stable in biological evolution.It is mainly summarized in three forms:Wnt/?-catenin classic pathway,planar cell polarity pathway,and Wnt/Ca2+ pathway.In recent years,the classical signal pathway of Wnt/P-catenin has been extremely hot in the study of tumor diseases,which is mainly manifested in the close relationship between this signaling pathway and the occurrence of tumor,invasion and metastasis,and poor prognosis.Avian leukosis virus(ALV)causes immunosuppression in chicken,causing a variety of tumors in the clinical and causing huge economic losses to the world poultry industry.Until now,there is no effective protective vaccine.To establish the healthy breeding flocks through the eradication of the core group of chickens is the main measure to control the disease.Previous transcriptional studies results in our research group indicate that the related gene expression of Wnt/?-catenin pathway was unusual regulated in the ALV-J infected tissues,which speculated that Wnt/?-catenin pathway might be play some role in ALV-J virus replication and associate with the pathogenicity of the virus.Therefore in this study we investigate the role of Wnt/?-catenin signaling pathway and replication of subgroup J avian leukosis virus and its possible mechanisms.It will provide a theoretical basis to further study the relationship between Wnt/?-catenin pathway and pathogenic and tumorigenic mechanisms of ALV-J.1.Comparison of the basic activation state of Wnt/beta-catenin signal pathway in poultry cells.Using real-time PCR,indirect immunofluorescence assay and Western blot methods we detect protein localization and gene expression levels of beta-catenin in chicken embryo fibroblast(CEF),chicken embryo kidney cells(CEK),DF-1,and chicken liver cancer cell(LMH)to reflect the underlying activation of the Wnt/?-catenin signaling pathway in the resting state of cells.The results of various experiments demonstrated that the protein content of beta-catenin in CEF cells was mainly located in the cell membrane,and the protein content of cytoplasm and nucleus was the lowest,and the activation status of Wnt/p-catenin signaling pathway was very low also.Therefore,in this study CEF cells were selected as cell models to study the relationship between the ALV-J infection and Wnt/?-catenin signaling pathway.2.The Wnt/?-catenin signaling pathway in CEF cells was activated by J subgroup avian leukosis virus infection and the effect of interference with P-catenin expression on the replication of ALV-J in the CEF cells.After infection with different MOI drop of subgroup J avian leukosis virus(JSGY07)in CEF cells,the TOP flash luciferase assay,transcription factor LEF/TCF expression level and the downstream gene expression of signaling pathway were carried out at post infection 12h,24h,48h,and 72h to reflect the activation status of signaling pathways.At the same time,the replication of the virus was investigated after interference RNA was used to inhibit the expression of beta-catenin in the cell.The results showed that the high titer virus infection could activate the Wnt/beta-catenin signaling pathway in CEF cells.Moreover,interference with the expression of beta-catenin can inhibit the replication of ALV-J in CEF cells.3.The effect on the replication of J subgroup avian leukosis virus after activation of the Wnt/beta-catenin signal pathway in CEF cells.According to previous reports,LiCl can activate the Wnt/?-catenin signaling pathway,while GSK-3? also can activate the same pathway by inhibition the formation of beta-catenin-AXIN-APC and GSK-3? complexes,which cause the beta-catenin gathered into the nucleus.In this study,we used two signaling pathways activators and one inhibitor,JW74 to study the effect on the replication of J subgroup avian leukosis virus after activation of the Wnt/?-catenin signal pathway in CEF cells.The results showed that LiCl significantly inhibited the replication of ALV-J in CEF cells,while GSK-3? inhibitor significantly promoted the replication of ALV-J.We think that these two different observations may be related to the different mechanisms of activators.The specific mechanism need further study to elucidate.