Mutational Landscape of Spontaneous Base Substitutions and Small Indels in Experimental Caenorhabditis elegans Populations of Differing Size

Abstract

Experimental investigations into the rates and fitness effects of spontaneous mutations are fundamental for our understanding of the evolutionary process. To gain insights into the molecular and fitness consequences of spontaneous mutations, we conducted a mutation accumulation (MA) experiment at varying population sizes in the nematode Caenorhabditis elegans, evolving 35 lines in parallel for 409 generations at three population sizes (N = 1, 10, 100 individuals). Here, we focus on nuclear SNPs and small indels under minimal influence of selection, as well as their accrual rates in larger populations under greater selection efficacy. The spontaneous rates of base substitutions and small indels are 1.84 × 10−9 substitutions and 6.84 × 10−10 changes /site/generation, respectively. Small indels exhibit a deletion-bias with deletions exceeding insertions by three-fold. Notably, there was no correlation between the frequency of base substitutions, nonsynonymous substitutions or small indels with population size. These results contrast with our previous analysis of mtDNA mutations and nuclear copy-number changes in these MA lines, and suggest that nuclear base substitutions and small indels are under less stringent purifying selection compared to the former mutational classes. A transition bias was observed in exons as was a near universal base substitution bias towards A/T. Strongly context-dependent base substitutions, where 5’–T and 3’–As increase the frequency of A/T → T/A transversions, especially at the boundaries of A or T homopolymeric runs, manifest as higher mutation rates in (i) introns and intergenic regions relative to exons, (ii) chromosomal cores versus arms and tips, and (iii) germline-expressed genes.

Mutability of mononucleotide repeats, not oxidative stress, explains the discrepancy between laboratory-accumulated mutations and the natural allele-frequency spectrum inC. elegans

“…In that study, motifs with C or G in the mutant (3’) site were more mutable on average than motifs with A or T in the mutant site, with one exception: the triplet 5’-ttA-3’ was the most mutable of any of the 64 motifs. Subsequent MA studies with the N2 strain of C. elegans have found a similarly high mutability of the 5’-ttA-3’ motif (Konrad et al 2019; Volkova et al 2020).…”

“…The ratio of transitions to transversions (Ts/Tv) in MA lines is consistently around 0.6-0.8, whereas the Ts/Tv ratio among wild isolates is around 1.1-1.2. Two early studies reported that G:C→T:A transversions are overrepresented in MA data relative to the standing nucleotide diversity (Denver et al 2009; Denver et al 2012); two more recent studies with more data reported that A:T→T:A transversions are also overrepresented in MA data relative to the standing nucleotide diversity (Konrad et al 2019; Saxena et al 2019).…”

Parallel genetics of regulatory sequences using scalable genome editing in vivo

“…On average, samples contained 57.9% deletions, 22.9% insertions, and 19.3% complex events (combinations of insertions, deletions, and substitutions) (Figure 2B). These proportions are similar for naturally occurring germline indels in C. elegans (75% deletions, 25% insertions) (Konrad et al, 2019) and human (50% deletions, 35% insertions) (Collins et al, 2020).…”

Mutation rate and spectrum in Caenorhabditis elegans mutation accumulation lines subjected to RNAi-induced knockdown of the mismatch repair gene msh-2

“…For each line, a trio of one female and two male worms were used to establish a population that was expanded for two to three additional generations to generate sufficient worm tissue for genomic DNA (gDNA) extraction. Sequencing followed the methodologies previously described (Konrad et al 2017(Konrad et al , 2018(Konrad et al , 2019. The PureGene Genomic DNA Tissue Kit (QIAGEN no.…”

“…The identification of small mutations (SNPs and indels up to 100 bp long) followed the same methodology previously used to identify mutations in the wildtype MA lines relative to their ancestral control (Konrad et al 2019). Briefly, alignment files from Phaster and BWA were analyzed separately.…”