Research On The Molecular Mechanism Of Inhibition Of Enterovirus D68 By APOBEC3G

Enterovirus 68(EVD68),a member of the group D of enteroviruses genus,which belongs to the Picornaviridae family.From the beginning of the discovery in the 1960 s to 2005,there were only sporadic infections in the world.In 2014,EVD68 broke out in the United States,and there were a large number of infected people in other countries around the world,EVD68 threats to people's lives and health.EVD68 mainly infects children,causes respiratory failure and pneumonia,and the outbreak of EVD68 in the United States even causes neurological diseases such as acute flaccid myelitis.The harm of EVD68 cannot be underestimated,and there is no effective vaccine or drug to prevent EVD68 infection.In order to prevent large-scale outbreaks and biosafety threats,EVD68 research is of great significance.A3 G is a member of the APOBEC3 subfamily of the APOBEC family.It has cytosine deaminase activity and RNA and ss DNA binding activity.It can limit many types of viruses such as HIV,HPV and EV71,and which has broad-spectrum antiviral activity.Enterovirus nucleic acid usually has no cap structure at 5' end and cannot bind to ribosome to initiate translation,but the ribosome entry site of its 5' non-coding region will recruit intracellular proteins such as PCBP1 and PCBP2 to initiate nucleic acid translation.Previous studies have shown that EV71 5'UTR recruits PCBP1 to initiate nucleic acid translation,and A3 G inhibits EV71 replication by competing with PCBP1 to bind to the ribosome entry site of the 5'non-coding region of EV71.Whether the 5 'non-coding region of enterovirus EVD68 will recruit PCBP1 to translate,and whether A3 G competes with PCBP1 to combine the 5' non-coding region of D68 has not been studied.In the research,we will study whether EVD68 recruits PCBP1 for translation and replication and the interaction mechanism between A3 G and them.The research will provide a theoretical basis for the development of broad-spectrum antiviral drugs.In this study,we found that EVD68 5'UTR need to interact with PCBP1 to replicate effectively,and silencing PCBP1 has no effect on CA6 replication.Then we found that A3 G can inhibit EVD68 replication,but not CA6 replication.Further study on the mechanism of A3 G inhibiting EVD68,we found that A3 G suppresses EVD68 replication by competing with PCBP1 to interact with EVD68 5'UTR.By predicting the secondary structure of EVD68 5'UTR and truncating the secondary structure,we explore the position of the bond of A3 G and PCBP1 in the secondary structure,and find that A3 G can compete with PCBP1 to bond EVD68 5'UTR's Loop II and loop IV.At the same time,we also studied the nucleic acid binding sites of A3 G,and found that when Y124 A and W127 A mutations,A3 G lost the function of binding EVD68 5'UTR,so A3 G lost the effect of inhibiting EVD68.These results also show that the nucleic acid binding activity of A3 G is also very important for its antiviral effect.In previous studies,viruses have also evolved a set of strategies to antagonize A3 G.For example,Vif encoded by HIV can cause the degradation of A3 G,and 2C encoded by EV71 can degrade A3 G through autophagy.In view of the conservation of 2C protein in enteroviruses,we found that EVD68 2C can also cause A3 G degradation,and A3 G degradation was restored after adding 2C inhibitor.According to the 2C autophagy domain of EV71,we designed the 2C truncation of EVD68,and finally determined that the domain of 25-41 is the key region for the degradation of A3 G.Through the related research,we can better understand the interaction mechanism between enterovirus and host,and provide a theoretical basis for broad-spectrum antiviral drugs.