Discussion On The Mechanism Of Guanylate Binding Protein 1 Affecting The Replication Of Foamy Virus

Foamy viruses(FVs)were named as causing rapid foam-like multinucleated cytopathic effects in many cell types.With some unique characteristics different from traditional retroviruses,FVs were classified as the only genus of the Spumaretrovirinae subfamily of retroviruses.At present,the most of FV isolates were from non-human primates.First case about human foamy virus(HFV)isolated from a Kenyan man bitten by a monkey was reported in 1971.At the end of 20th century,sequences aligment showed that there was a high similarity between HFV and SFVcpz,so some researchers indicated that HFV was the result of SFV cross-species transmission,and named HFV as prototype foamy virus(PFV).FVs established a lifelong persistent infection in natural hosts or experimental infection animals with no associated pathologies,however the cytopathic effects in vitro would provoke the host innate immune.As the main defense factor of the innate immne,studies showed that IFN-y could effectively reduce the FVs cytopathic effect.The similar result was found in previous experiments performed in our laboratory.When stimulating PFV infection HT1080 with IFN-?,the expression of intracellular PFV Gag protein was greatly down-regulated compared with the control.While at the same time,the Guanylate-binding protein 1(GBP 1)expression was upregulated for more than four times.Until now,the anti FVs cytokines such as IFP35,PML,Nmi and APOBEC3 had been reported,but there was nothing about GBP1.It is worth to study out whether GBP1 mediates the anti PFV faction of IFN-? as one of the cytokines downstream of IFN-y.GBP1 was named as its ability to combine guanylic acid.GBP1 has GTPase activity which is able to hydrolysis GTP to GDP and GMP.Together with MX and dynamin,GBP 1 belongs to large GTP-binding proteins.As an interferon induced protein,GBP1 has certain antiviral function.It has been proved that GBP1 showed the inhibition effect on virues replication of VSV,EMCV,IAV and HCV which related to its GTPase activity.Whether GBP1 might inhibit the PFV replication has not been studied.In this study,exogenous overexpression of GBP1 and endogenous knockdown GBP1 were carried out to test the effects of GBP 1 to PFV replication.The final results showed that no matter instaneous or stable overexpression of GBP 1,in HT1080 cells transfected by pHSRV 13 carrying PFV genome or in HT1080-eGFP-GBP1 cells infected by PFV,PFV gag mRNA and Gag protein expressions were respectively upregulated nearly 2 times and 0.77 times compared with negative control group.Meanwhile,the cleaved P68 Gag protein relative to precursor P71 was obviously increased,and presented the GBP1 concentration dependence.Consistant with the above results,along with knockdown GBP1 by 56.7%in protein level,the expression of PFV Gag protein reduced 66.9%.So at the point of 48 hpi,GBP1 promoted the expression and cleavage of Gag protein.However,at the other 2 time ponits post PFV infection,72 hr and 96 hr,GBP1 seemed not always maintained this feature.But when the virus titer improved,the phenomenon was delayed.That is,under the condition of overexpression of GBP1,there was no significant difference of Gag expression between the experiment and the control only at the point of 96 hr.Later,the detection of PFV other proteins and PFV particles showed that with overexpression of GBPl,the expression of Pol and Bet proteins were upregulated obviously compared with the negative control groups.As well,the extracellular PFV titers at 3 time points,48 hr,72 hr and 96 hr,were increased nearly six times,more than seven times and about 4.3 times,respectively,relative to the negative control groups.For the 48 hr and 72 hr,the increase of PFV particles extracellular was far greater than intracellular.So according to all experiment results,the main function of GBP1 influence on PFV was speeding up the PFV replication cycle,and promoting PFV virus particle release or budding.This result showed opposite characteristic to the reported GBP1 antiviral function.That may be causing by different specificity of unique viruses or different microenvironment and immune regulation mechanism in host cells.Later,the function of GBP1 acting on PFV,perhaps correlating with GTPase or protein Bet would be worth doing to find out the real mechanism of GBP1 influence on PFV replication.