The Role Of TRIM62 In Inhibiting Avian Reticuloendotheliosis Virus Infection

Reticuloendotheliosis virus(REV)belongs to avian retrovirus.In addition to the formation of tumors,it can also lead to immunosuppression,causing secondary infections and mixed infections.There has been a major loss in the poultry industry,and there are no vaccines or drugs for commercial use.TRIM62 is a member of the TRIM/RBCC family and includes the highly conserved RING,B-box,and Coiled coil domains of the TRIM family,with the C-terminus being the SPRY domain.Many proteins of the TRIM family have antiretroviral effects and their mechanisms of action are closely related to their domains.As a natural immune factor,the anti-retroviral effect of TRIM62 on other species has been studied,such as human TRIM62 protein against HIV-1 replication,and its RING domain deletion enhances its inhibition of HIV-1.Based on this,this study designed a study to study the inhibitory effect of chicken TRIM62 on retroviral REV replication,clarify the influence of its domain,and analyze the interaction protein of TRIM62 in inhibiting REV replication,in order to further study chicken TRIM62 inhibition of avian The mechanism for transcriptional viral replication lays the foundation.Fluorescence quantitative PCR and Western Blot were used to detect the expression of TRIM62 in cells at different time points and in chicken tissues,and the effect of REV replication on TRIM62 expression and the correlation between them were confirmed.TRIM62 lentiviral vector and shRNA expression vector were transfected respectively,and REV vaccination was performed to detect the expression of TRIM62 and REV,and the inhibitory effect of TRIM62 on REV was determined from an external perspective;The supernatant of the poisoned cells overexpressing TRIM62 was inoculated again,and the expression of REV was detected to determine the effect of TRIM62 on virion release;The chicken embryos of 60 h embryos were transfected with the recombinant vector of TRIM62 lentivirus in vivo.The chicks were exposed to the virus at 3 days old.The expression of REV in each tissue was detected,and the inhibitory effect of TRIM62 on REV was determined from the internal perspective;The TRIM62 domain deletion mutant lentivirus was constructedand transfected into CEF cells,and REV was inoculated to detect REV expression,and the effect of each domain on TRM62 inhibition of REV replication was determined;After over-expressing TRIM62 cells,the immunoprecipitation was carried out,and the small peptide was separated by SDS-PAGE.The protein-containing gel was cut by Coomassie blue staining,and the protein was qualitatively analyzed by liquid chromatography tandem mass spectrometry.A qualitative change in the protein with which TRIM62 interacts with REV replication is analyzed.The results showed that the expression of TRIM62 in REV-infected cells decreased first,and gradually increased with the prolonged exposure time,and decreased again after 72 h,indicating that TRIM62 expression levels were different at different stages of viral replication.In the organs of liver,kidney,spleen and bursa of the poisoned chicken,the expression of TRIM62 was significantly decreased in one week,and the expression was reversed at the 7th week with the prolongation of the intoxication time,suggesting that TRIM62 is involved in the anti-REV process of the body as an important protein.This indicates that TRIM62 is involved in the body's anti-REV process as an important immune protein.In vitro TRIM62 overexpression and inhibition of TRIM62 expression confirmed that TRIM62 can effectively inhibit viral replication and reduce virion release.Overexpression of TRIM62 in vivo can effectively inhibit REV replication in various tissues and organs.The four domains of TRIM62 have the effect of inhibiting REV,followed by SPRY domain>RING domain>B-box domain>Coiled coil domain.Differences in protein qualitatively analyzed by mass spectrometry.The final screened proteins interacting with TRIM62 during viral replication were 18 species,including Arp2/3.This study showed that TRIM62 can effectively inhibit REV replication and reduce virion release.The SPRY domain has the greatest effect on TRIM62 inhibition of REV replication.TRIM62 induces qualitative changes in 18 interacting proteins including Arp2/3during anti-REV,which provides a basis for subsequent studies on TRIM62 inhibition of avian retrovirus replication.