(A) Prior to the competition experiment, ubiquitin alleles were specifically associated with unique barcodes through a paired end sequencing. To monitor the frequency of different alleles during the …
The fitness of all ubiquitin mutants was measured in a single culture by shutting off the galactose-driven wild type copy. This allows a constitutively expressed mutant to be the sole source of …
(A) Heatmap showing the fitness of observed ubiquitin alleles. Scores presented are the average of three biological replicates. Wild type amino acids are shown in green and mutations without fitness …
We calculated the standard deviation of the distributions of barcode fitness scores that contribute to each amino acid mutant fitness score. Large errors of the stop codon substitutions are due to …
A linear regression (R2 = 0.785) and Pearson’s correlation coefficient (CC = 0.886) were calculated between the fitness scores determined in DMSO and the previously published unperturbed dataset (Ros…
The difference in fitness between DMSO and a perturbation for each Ub allele: (A) Caffeine, (B) DTT (C) Hydroxyurea (D) MG132. Wild type amino acids are shown in red and mutations without fitness …
Heatmaps showing the fitness of observed Ub alleles under: (A) Caffeine, (B) DTT (C) Hydroxyurea (D) MG132. Wild type amino acids are shown in green and mutations without fitness values (due to lack …
Heatmaps showing the error estimates for each observed Ub alleles under; (A) Caffeine, (B) DTT (C) Hydroxyurea (D) MG132. Wild type amino acids are shown in green and mutations without fitness …
We determined the concentration to inhibit SUB328 growth by 25% by monitoring optical density. Error bars represent standard deviation of multiple measurements.
The residuals between datasets shows are shown with the Lorentzian representing the biological replicates of DMSO in red. When compared to DMSO, three perturbations (Caffeine, DTT and HU) shift the …
Fitness scores for each of the 5 sets of experimental conditions are shown along the y-axis as boxplots. Scores are grouped first by their respective experimental condition, and then by the change …
Cartoon model of Ub (PDB 1UBQ) with important residues colored as follows: Lys48 - orange, Lys63 - light blue, Lys11 - green, other Lys residues - yellow, hydrophobic patch (Leu8, Val40, Ile 44) - …
(A) Based on the average fitness score, positions were binned into tolerant (>=-0.075 - Blue), intermediate (<-0.075 to > -0.35 - Pink) and sensitive (<= -0.35 - Red). (i) DMSO (ii) Caffeine (iii) …
(A) DMSO fitness - Caffeine fitness vs. DMSO fitness - DTT fitness. The markers are colored based on DMSO fitness - Hydroxyurea fitness. (B) DMSO fitness - Caffeine fitness vs. DMSO fitness - …
Shared response mutants representing mutations that are equally perturbed by all three sensitizing perturbations.
Mutants in the shared response were determined by fitting a line to the fitness scores. The distance from each point to that line was calculated. If the distance was less than 0.1 and the average Δ (DMSO - Perturbation) fitness was less than -0.2 the mutant was considered part of the shared response. E1 activity relative to WT Ub (Roscoe and Bolon, 2014) is listed and may explain the sensitization of some of the shared response mutants.
Perturbation specific mutations represent alleles that are differentially affected by Caffeine, DTT and Hydroxyurea.
Perturbation specific mutations were determined by fitting a line to the delta (DMSO - perturbation) fitness scores. The distance from each point to that line was calculated. If the distance was greater than 0.35 the mutant was classified as perturbation specific. Mutants with high experimental errors were deemed outliers and removed from this list.
Specific information regarding highlighted mutants.