Negative selection allows human primary fibroblasts to tolerate high somatic mutation loads induced by N-ethyl-N-nitrosourea

High-throughput sequencing at the single-cell and single-molecule level has shown that thousands of mutations accumulate with age in most human tissues. While there is ample evidence that some of these mutations can clonally amplify and lead to disease, the total burden of mutations a cell tolerates without functional decline remains unknown. Here we addressed this question by exposing human primary fibroblasts multiple times to low doses of N-ethyl-N-nitrosourea (ENU) and analyzing somatic mutation burden using single-cell whole genome sequencing. The results indicate that individual cells can sustain ~60,000 single-nucleotide variants (SNVs) with only a slight adverse effect on growth rate. Evidently high levels of mutations are tolerated through negative selection against potentially deleterious variants in gene coding regions, and sequences associated with genetic pathways relevant for maintaining basic cellular function and growth. Since most tissues in adults are non-dividing, these results suggest that somatic mutations in the absence of negative selection can have functionally adverse effects.