Extensive loss of cell-cycle and DNA repair genes in an ancient lineage of bipolar budding yeasts
Fig 6
Analyses of base substitutions and indels reveal a higher mutational load in the FEL compared to the SEL.
(A) Analyses of substitution patterns among codon-based alignments of 1,034 OGs revealed a higher number of base substitutions in the FEL compared to the SEL (F(1) = 196.88, p < 0.001; multifactor ANOVA) and an asymmetric distribution of base substitutions at codon sites (F(2) = 1,691.60, p < 0.001; multifactor ANOVA). A Tukey honest significance differences post hoc test revealed a higher proportion of substitutions in the FEL compared to the SEL at the first (n = 240,565; p < 0.001), second (n = 318,987; p < 0.001), and third (n = 58,151; p = 0.02) codon positions. (B) Analyses of the direction of base substitutions (i.e., G|C → A|T or A|T → G|C) revealed significant differences between the FEL and SEL (F(1) = 447.1, p < 0.001; multifactor ANOVA) as well as differences in the directionality of base substitutions (F(1) = 914.5, p < 0.001; multifactor ANOVA). A Tukey honest significance differences post hoc test revealed a significantly higher proportion of substitutions were G|C → A|T compared to A|T → G|C among sites that are G|C (n = 232,546) and A|T (n = 385,157) (p < 0.001), suggesting a general AT bias of base substitutions. Additionally, there was a significantly higher proportion of sites with base substitutions in the FEL compared to the SEL (p < 0.001). Specifically, a higher number of base substitutions was observed in the FEL compared to the SEL for both G|C → A|T (p < 0.001) and A|T → G|C mutations (p < 0.001), but the bias toward AT was greater in the FEL. (C) Examinations of transition/transversion ratios revealed a lower transition/transversion ratio in the FEL compared to the SEL (p < 0.001; Wilcoxon rank–sum test). (D) Comparisons of insertions and deletions revealed a significantly greater number of insertions (p < 0.001; Wilcoxon rank–sum test) and deletions (p < 0.001; Wilcoxon rank–sum test) in the FEL (; ) compared to the SEL (; ). (E and F) When adding the factor of size per insertion or deletion, significant differences were still observed between the lineages (F(1) = 2,102.87, p < 0.001; multifactor ANOVA). A Tukey honest significance differences post hoc test revealed that most differences were caused by significantly more small insertions and deletions in the FEL compared to the SEL. More specifically, there were significantly more insertions in the FEL compared to the SEL for sizes 3–18 (p < 0.001 for all comparisons between each lineage for each insertion size), and there were significantly more deletions in the FEL compared to the SEL for sizes 3–21 (p < 0.001 for all comparisons between each lineage for each deletion size). Black lines at the top of each bar show the 95% confidence interval for the number of insertions or deletions for a given size. (G) Evolutionarily conserved homopolymers of sequence length 2 (n = 17,391), 3 (n = 1,062), 4 (n = 104), and 5 (n = 5) were examined for substitutions and indels. Statistically significant differences of the proportion mutated bases (i.e., [base substitutions + deleted bases + inserted bases]/total homopolymer bases) were observed between the FEL and SEL (F(1) = 27.68, p < 0.001; multifactor ANOVA). Although the FEL had more mutations than the SEL for all homopolymers, a Tukey honest significance differences post hoc test revealed differences were statistically significant for homopolymers of two (p = 0.02) and three (p = 0.003). Analyses of homopolymers using additional factors of mutation type (i.e., base substitution, insertion, deletion) and homopolymer sequence type (i.e., A|T and C|G homopolymers) can be seen in S10 Fig. (H) G → T or C → A mutations are associated with the common and abundant oxidatively damaged base, 8-oxo-dG. When examining all substituted G positions for each species and their substitution direction, we found significant differences between different substitution directions (F(2) = 5,682, p < 0.001; multifactor ANOVA). More importantly, a Tukey honest significance differences post hoc test revealed an over-representation of G → T or C → A in the FEL compared to the SEL (p < 0.001). (I) Signatures of UV-damage–associated single and double substitutions (i.e., C → T at CC sites and CC → TT) double substitutions are greater in the FEL compared to the SEL (p < 0.001 for both tests; Wilcoxon rank–sum test). figshare: https://doi.org/10.6084/m9.figshare.7670756.v2. FEL, faster-evolving lineage; OG, orthologous gene; Pro., Proportion; SEL, slower-evolving lineage.