The Effect Of Cellular Unfolded Protein Response On Avain Reovirus Replication And Inducing Cell Apoptosis

Avian reovirus(ARV)is one of the important infectious disease in poultry,causing viral arthritis,tenosynovitis and chronic respiratory disease,inducing immunosuppression and growth dysplasia in chickens.In recent years,ARV infection has been frequently found in commercial broiler farms and breeder farms in Jiangsu,Shandong,Fujian and other places,decreasing huge profitability of poultry industry.The unfolded protein response(UPR),a series of compensatory regulatory mechanisms initiated by cells,is aimed in maintaining endoplasmic reticulum homeostasis.UPR is regulated by a variety of signal pathways,of which three classic pathways are ATF6,PERK and IRE1 pathway.The activation of UPR can promote the capability of cells such as folding unfolded proteins and degradating misfolded proteins,as well as enhancing the secretion ability under endoplasmic reticulum stress to maintain endoplasmic reticulum homeostasis and promoting cell survival or regulating cell apoptosis.The molecular mechanism of avian reovirus activating cell UPR ATF6 pathway is still unclear at present.In this study,vero cells were used to investigate the molecular mechanism of avian reovirus inducing unfolded protein response in cells and its relationship with virus replication and apoptosis.The experimental results are as follows:1.ARV infection of vero cells activates the three pathways of cell UPR.QRT-PCR and Western blot were used to detect the transcription and translation expression levels of key factors in the three UPR pathways in cells at different time points of ARV infection.The results showed that the expressions of GRP78,ATF6,and CHOP were significantly up-regulated and XBP1 had a splicing effect,indicating that ARV infection activates all pathways of cell UPR.2.UPR can promote ARV proliferation.?C protein is an important part of ARV synthetic virus particles,and can be used as a detection gene for virus proliferation.UPR inducer tunicamycin was used to activate cell UPR,endoplasmic reticulum stress inhibitor 4-PBA and shRNA-mediated knockdown of ATF6 expression,after inoculation with ARV virus,the ARV replication level was detected by qRT-PCR method.The results indicate that inhibition of intracellular UPR can inhibit the level of ARV replication.3.ARV can induce host cell apoptosis by inducing UPR.The samples of vero cells infected with ARV at different time points were collected,and the apoptosis rate and the protein level changes of Caspase-12 and cleaved-Caspase-3 were detected by flow cytometry method and Western blot method.The results showed that while using endoplasmic reticulum stress inhibitor 4-PBA and shRNA mediated knockdown of ATF6 expression,the level of virus replication in ARV-infected vero cells decreased,while the rate of apoptosis decreased significantly,indicating that UPR and its ATF6 pathway play an important role in cell apoptosis induced by ARV.Therefore,ARV infection can activate the cell UPR signaling pathway in cells.Inhibition of UPR and ATF6 pathway plays an important role in inhibiting ARV proliferation and cell apoptosis induced by ARV.