The Construction And Study Of A Novel Recombinant Vesicular Stomatitis Virus With A Mutant Of Matrix Protein Amino Acid Sites

Recombinant vesicular stomatitis virus（rVSV）has many advantages such as efficient expression of foreign proteins,offer high level of protection against viral infections and its ability to induce strong cellular and humoral host immune responses.So,rVSV is widely used in vaccine and oncology research.However,wild-type VSV is both neurotropic and neurovirulent in mice,and can cause neurological disease when injected directly into the brain of cows and horses.Therefore,we urgently need to transform the VSV vector vaccine to decrease its pathogenesis.Viral matrix protein is a multifunctional protein that is involved in the shutoff of host transcription,nuclear cytoplasmic transport,and translation during virus infection.Wild-type VSV effectively suppress the host innate immune response through the inhibition of type I interferon（IFN-α/β）expression by the M protein.So,M protein is an important virulence factor of VSV and is closely related to the pathogenicity of the virus.Studies have shown that the pathogenicity of VSV can be attenuated by the mutation of amino acid 51（methionine）of M protein to arginine(VSV_M51R)or deletion(VSV_ΔM51).In addition,Hoffmann et al.demonstrated that another VSV-M mutant strain with two amino acid substitutions（V221F and S226R）could also attenuate.However,these two sites have definite significance in the pathogenesis of VSV,especially in animals no detailed study and research in vivo yet.In this paper,recombinant VSV aiming at amino acids of 221 and 226 in matrix protein were constructed with their pathogenicity characterized in vivo and in vitro.In vitro experiments include:（1）Multi-step growth curve of recombinant VSVs in PC3 or PEF cells;（2）Expression of type I interferons by different viruses;（3）Analysis of VSV-mediated shutoff of host mRNA expression.In vivo,we used BABL/c mice to evaluate virulence of different VSVs,including the vaccination of mice,animal body weight losses,survival and the content of virus in mouse lung and brain tissue.In this study,in contrast to wild-type VSV,VSV with double mutation occurring at amino acid sites of 221 and226 can significantly attenuate,which can be due to efficient induction of typeⅠinterferon in host cells due the viral infection,which indicates that VSV_M221,226 can be a promising vector for developing vaccine.