FIG. 3.
Expression of KUNV genes/gene cassettes by the SFV replicon vector results in correct processing of the individual proteins and reveals a requirement of the viral protease for cleavage of NS4A and NS4B. BHK cells were electroporated and radiolabeled at 11 h posttransfection with [35S]methionine-cysteine. Individual KUNV proteins were then isolated from the collected lysates by RIP using monospecific rabbit antisera (A and B). Protein migration was assessed against a [35S]methionine-cysteine-radiolabeled lysate generated from KUN-infected BHK cells and by the predicted molecular mass. Note that the expression of the NS2B-3pro construct yields an NS3 species of ∼20 kDa corresponding to the first 187 amino acids of the NS3 protein that is isolated after RIP with anti-NS3 antibodies. Uncleaved NS4A-NS4B polyproteins are indicated by asterisks in panel A. Mock lysates are shown in panel C.