A speed-fidelity trade-off determines the mutation rate and virulence of an RNA virus

William Fitzsimmons; Robert J. Woods; John T. McCrone; Andrew Woodman; Jamie J. Arnold; Madhumita Yennawar; Richard Evans; Craig E. Cameron; Adam S. Lauring

. CC-BY 4.0 International license is made available under a The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It . https://doi.org/10.1101/309880 doi: bioRxiv preprint Figure S1 . CC-BY 4.0 International license is made available under a The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It . https://doi.org/10.1101/309880 doi: bioRxiv preprint Figure S1 . In vitro assay of polymerase mediated single nucleotide incorporation. (A) Schematic of GTP misincorporation assay (G opposite the U). Primer-template (sym-subU) and polymerase are assembled in the absence of nucleotide. GTP and a 25-fold excess of unlabeled trap RNA are then added after an incubation period (Δt). Excess of RNA trap ensures that if the polymerase dissociates from primer-template it is taken up by the trap and cannot re-assemble. (B) Kpol and Kd for GMP misincorporation. Concentrations of GTP were used over time-points to calculate Kpol and Kd. Each GTP time-course was plotted to single exponential, then combined plot to hyperbola. Note that all polymerase variants have the I92T mutation. The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It . https://doi.org/10.1101/309880 doi: bioRxiv preprint Figure S2 . Log fitness versus passage in adaptability experiment. Each point is the fitness mean ± standard deviation of three replicate competition assays. (A) WT and 3DG64S on HeLa (B) WT and 3DG64S on PVR-3T3. For the graphs in (B), the relationship of log fitness vs. passage was not linear past passage 16, and only passages 1-12 are shown. The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It . https://doi.org/10.1101/309880 doi: bioRxiv preprint Figure S3 . Mutagen sensitivity of 2C-V127L variants. HeLa were infected at an MOI of 0.1 with the indicated viruses in presence of various concentrations of ribavirin. After 24 hours, titers of mock and ribavirin-treated populations were determined by TCID50 and those of ribavirin-treated populations were normalized to mock-treated controls (mean of 5 measurements per virus). Shown are the changes in titer (y-axis, mean, 5 replicates) for each virus at each drug concentration (x-axis). A greater reduction in titer (more negative number) indicates higher mutagen sensitivity and suggests a higher baseline mutation rate.