Negative selection allows human primary fibroblasts to tolerate high somatic mutation loads induced by...

With advanced sequencing methods, it has now been shown that thousands of mutations accumulate with age in most human tissues. While there is ample evidence that some mutations can clonally amplify and lead to disease, the total burden of mutations a cell can tolerate without functional decline remains unknown. Here we directly addressed this question by exposing serially cultured human primary fibroblasts to multiple, low doses of N-ethyl-N-nitrosourea and analyzing somatic mutation burden using single-cell whole genome sequencing. The results indicate that mitotically active cells can sustain at least ∼56,000 single-nucleotide variants with only a slight adverse effect on growth rate. We provide evidence that such high levels of mutations are tolerated through negative selection against variants in gene coding and non-coding regions, and in sequences associated with genetic pathways for maintaining relevant cellular functions such as growth and cell identity. Since most tissues in adults are non-dividing, these results suggest that somatic mutations in the absence of negative selection may have functionally adverse effects.