| Bovine parainfluenza virus type 3（BPIV3）,also known as Bovine respirovirus 3,belongs to the Paramyxoviridae Respirovirus genus.According to reports,BPIV3 has become a bovine pathogen that causes severe economic losses,and it is extremely harmful to both adult cattle and calves.In addition,together with other viruses and pathogens,BPIV3 can cause complications and formation of Bovine Respiratory Disease Complex（BRDC）with symptoms of cough,fever,etc,which has become a major problem affecting the health of cattle worldwide.At present,inactivated or attenuated vaccines are mainly used for immune prevention in dairy developed countries,but there is still not suitable vaccine in our country,previous studies have demonstrated that infection of BPIV3 causes innate immune response within the host cell.Therefore,it is necessary to further study the molecular mechanism of virus replication,especially the research hotspots such as the relationship between virus and innate immunity,so that to find drug targets and provide ideas for the development of new antiviral drugs.β-catenin protein is a multifunctional protein with cell adhesion and signal transduction,and it is widely found in fibroblasts,endothelial cells,osteoblasts and other cells.β-catenin is a key component of the Wnt/β-catenin signal pathway.Wnt/β-catenin signaling pathway is not only essential for regulating the early development of embryos,maintenance of tissue homeostasis in adults,and regeneration of the central nervous system,but also closely related to cell proliferation,differentiation,migration and apoptosis.As report goes,the abnormal regulation of Wnt/β-catenin signaling is tightly associated with many diseases.However,the viruses can affect the Wnt/β-catenin signaling pathway,several studies have showed the relationship of the Wnt/β-catenin signaling pathway with virus replication according to the existing research,such as the human immunodeficiency virus（HIV）,the porcine circovirus-like virus P1,the Human Cytomegalovirus（HCMV）,the hepatitis C virus（HCV）and so on.However,the interaction betweenβ-catenin and BPIV3 has not been reported.In this study,the effect ofβ-catenin gene on BPIV3 replication and its mechanism were studied,and the main research progress was as follows:1.After BPIV3 infection,β-catenin mRNA level was up-regulated,while protein level was down regulated.Especially in the late stage of infection,β-catenin protein was down regulated significantly.The samples of Madin-Darby Bovine Kidney Cells（MDBK）at different time points after BPIV3 infection were detected by qRT-PCR and Western blot respectively,the results showed that the mRNA level ofβ-catenin was up-regulated,but the protein level ofβ-catenin was down regulated,which indicated that BPIV3 infection could regulate the expression ofβ-catenin at the post translation level.2.Using gene overexpression and silencing,it was found thatβ-catenin could inhibit BPIV3.The stable cell lines ofβ-catenin overexpression and silencing were constructed to infect the virus.TCID50 was used to detect the change of virus titer.The results showed that the overexpression ofβ-catenin could inhibit the proliferation and replication of BPIV3,while the silencing ofβ-catenin promoted the proliferation of BPIV3,indicating thatβ-catenin has the function of anti BPIV3.3.As a transcription factor,β-catenin can enter the nucleus and combine with TCF/LEF,and jointly promote the expression of IFN-βand its downstream ISGs（such as OAS1）,so as to played an antiviral role.As the specific inhibitor of GSK3β,LiCl was used to inhibit the activity of GSK3β.Western blot and immunofluorescence detection showed that LiCl could enhance the expression ofβ-catenin,promote its accumulation in the cytoplasm and then enter the nucleus,thus further promoting the expression of IFN-βand its downstream interference gene OAS1,so as to play the role of resistance to BPIV3 and inhibit the replication of the virus.4.BPIV3 infection can significantly up-regulate the expression of GSK3β,thus accelerating the degradation ofβ-catenin in the cytoplasm.Through nuclear and cytoplasmic separation experiments,compared with the uninfected group,after BPIV3 infection,the expression ofβ-catenin decreased significantly,while the expression of GSK3βincreased significantly,indicating that BPIV3 infection can up-regulate the expression of GSK3β,thus down-regulate the expression ofβ-catenin.After LiCl treatment,the expression ofβ-catenin in cytoplasm and nucleus was promoted.It was suggested that the down-regulation ofβ-catenin protein expression caused by BPIV3 infection was due to GSK3βmediated degradation ofβ-catenin by proteasome.In summary,this study first discovered the role and mechanism of BPIV3infection in degradingβ-catenin protein andβ-catenin against BPIV3 replication by studying the relationship betweenβ-catenin and BPIV3.These findings can provide perspectives and ideas for further exploration of the pathogenesis of BPIV3,at the same time,it also provides a target molecule for the research and development of new drugs against BPIV3. |
