Fig. 4: Constraints imposed on mutational resistance space by fitness space.
From: Decoding efficacy and resistance space at a drug binding site
a The schematic illustrates the MOT-library fitness profiling approach. The set of twenty ssODNs with degenerate codons were individually transfected into T. brucei MSH2 RNAi cells, either with two (diploid, 2n, green) or one (haploid, 1n, purple) proteasome β5 allele, 24 h after inducing knockdown with tetracycline. Cultures were assessed using deep sequencing of β5 amplicons and codon variant scoring prior to transfection and in duplicate, 6 h after transfection and 4 d after transfection, for each genotype. Created in BioRender. Altmann, S. (2025) https://BioRender.com/wi23cs9. b The plots show fitness, as revealed by the relative percentage of reads for stop codon edits remaining 4d after ssODN transfection, and at the sites indicated in the proteasome β5 subunit; in the diploid strain (upper panel) or in the haploid strain (lower panel). c The boxplot shows fitness, as revealed by the relative percentage of reads for mis-sense (non-synonymous) edited codons remaining 4d after ssODN transfection, at the sites indicated in the proteasome β5 subunit; averages from two replicate screens and >27 M reads mapped per site. Boxes indicate the interquartile range (IQR), the whiskers show the range of values within 1.5 × IQR, and a horizontal line indicates the median. The notches represent the 95% confidence interval for each median.
