FIG 5.

Mutations in the P-side of the 1/2 site affect the rescue of SFV^CP and SFV^CP2 and the processing of the corresponding ns-polyproteins. (A) Sequences of the P6-P1′ regions of the 1/2 sites of CHIKV, SFV, SFV^CP, and SFV^CP2. Mutations introduced to obtain SFV^CP+1G536V, SFV^{CP+1Y533D+1H534R}, SFV^CP2+1G536V, and SFV^{CP2+1Y533D+1H534R} are shown in boldface and italics. (B) Localization of swapped regions and introduced mutations in the ns-regions of the chimeric genomes. ICA titers are shown as PFU per 1 μg RNA and represent the averages of the values obtained in three independent experiments; <2 indicates values below the detectable limit. Plaque sizes are indicated as follows: large, >3 mm; very small, <1 mm in diameter. P₀ titers are shown as PFU per ml and represent the averages of the values obtained in three independent experiments. The data for WT SFV, SFV^CP, and SFV^CP2 are replotted from Fig. 1A. (C) Results of in vitro translation and processing. The assay was conducted as described in the legend to Fig. 4. Each number represents the mean ± SD from three independent experiments. * indicates complete processing of P123 (corresponding density at background level); the numbers in parentheses indicate a lack of increase in the amount of product with the same mobility as that of nsP2 during the chase.