VPgGeneAmplification Correlates withInfective Particle Formation inFoot-and-Mouth Disease Virus

Inordertoanalyze thefunction ofVPgamplification inaphthoviruses, we haveundertaken thefirst mutational analysis oftherepetitive VPg-coding region using an improved foot-and-mouth disease virus (FMDV)cDNAclone fromwhichinfective viral RNA was synthesized. A setofVPgmutants was constructed bysite-directed mutagenesis whichincludes different VPgdeletion mutations, aVPginsertion mutation, and aminoacidresidue replacement mutations thatinterfere withbinding oftheVPgprotein totheviral RNA and withitsproteolytic processing. Ourresults revealed thatan amazing flexibility inthenumberofVPgsis tolerated inFMDV.Optimal viability isgiven whenthree VPgsareencoded. Deletion aswell asinsertion ofone VPggenestill resulted ininfective particle production. Infective particle formation was observed aslong asone VPgremained intact. Noobvious differences intheindividual VPgmolecules withregard totheir promoting viral RNA synthesis wereobserved, indicating thatallthree VPgscanactequally inFMDV replication. Mutant polyprotein processing was comparable tothatofthewild-type virus. However, VPgmutantsshowedreduced viral RNA synthesis levels after infection. Thelevels ofviral RNA synthesis andinfective particle formation were foundtocorrelate withthenumberoffunctional VPgsleft inthemutantvirus. Thesefindings suggest a direct VPggenedosage effect on viral RNA synthesis, witha secondary effect on infective particle formation.