CHARACTERIZATION OF SYNTHETIC FOOT-AND-MOUTH-DISEASE VIRUS PROVIRIONSSEPARATES ACID-MEDIATED DISASSEMBLY FROM INFECTIVITY

One of the final steps in the maturation of foot-and-mouth disease vir us (FMDV) is cleavage of the VP0 protein to produce VP4 and VP2. The m echanism of this cleavage is unknown, but it is thought to function in stabilizing the virus particle and priming it for infecting cells. To investigate the cleavage process and to understand its role in virion maturation, we engineered synthetic FMDV RNAs with mutations at Ala-8 5 (A85) and Asp-86 (D86) of VP0, which border the cleavage site. BHK c ells transfected with synthetic RNAs containing substitutions at posit ion 85 (A85N or A85H) or at position 86 (D86N) yielded particles indis tinguishable from wild-type (WT) virus in sedimentation and electropho retic profiles. Viruses derived from these transfected cells were infe ctious and maintained their mutant sequences upon passage. However, BH K cells transfected with synthetic RNAs encoding Phe and Lys at these positions (A85F/D86K) or a Cys at position 86 (D86C) produced noninfec tious provirions with uncleaved VPO molecules. Despite their lack of i nfectivity, the A85F/D86K provirions displayed cell binding and acid s ensitivity similar to those of WT virus. However, acid breakdown produ cts of the A85F/D86K provirions differed in hydrophobicity from the co mparable WT virion products, which lack VP4. Taken together, these stu dies are consistent with a role for soluble VP4 molecules in release o f the viral genome from the endosomal compartment of susceptible cells .