Lagging-strand replication shapes the mutational landscape of the genome

Abstract

The origin of mutations is central to understanding evolution and of key relevance to health. Variation occurs non-randomly across the genome, and mechanisms for this remain to be defined. Here, we report that the 5′-ends of Okazaki fragments have significantly elevated levels of nucleotide substitution, indicating a replicative origin for such mutations. With a novel method, emRiboSeq, we map the genome-wide contribution of polymerases, and show that despite Okazaki fragment processing, DNA synthesised by error-prone Pol-α is retained in vivo, comprising ~1.5% of the mature genome. We propose that DNA-binding proteins that rapidly re-associate post-replication act as partial barriers to Pol-δ mediated displacement of Pol-α synthesised DNA, resulting in incorporation of such Pol-α tracts and elevated mutation rates at specific sites. We observe a mutational cost to chromatin and regulatory protein binding, resulting in mutation hotspots at regulatory elements, with signatures of this process detectable in both yeast and humans.

Sister chromatid telomere fusions, but not NHEJ-mediated inter-chromosomal telomere fusions, occur independently of DNA ligases 3 and 4

“…5C,D) might reflect the distinct DNA polymerases that replicate leading-versus lagging-strand DNA. Leading-strand DNA synthesis is coordinately mediated by DNA polymerases delta and epsilon, resulting in minimal error incorporation (Nick McElhinny et al 2008;Lujan et al 2012;Johnson et al 2015), whereas lagging-strand replication occurs by ligation of Okazaki fragments synthesized by DNA polymerases alpha and delta with reduced proof-reading capacity Reijns et al 2015). We hypothesized that the shorter chromatid partner of each 17p intra-chromosomal fusion might result predominantly from lagging-strand DNA synthesis and thus display a higher frequency of nonconstitutive nucleotide changes (Supplemental Fig.…”

“…We were able to uncover a skew in error-incorporation segregating with short versus long paired chromatids that may reflect the ligation of leading with lagging-strand DNA. The reduction in proofreading capacity of lagging-strand polymerases (Reijns et al 2015), coupled with increased fork stalling at telomeric locations (Lormand et al 2013) could result in an incompletely replicated template (Chow et al 2012) that is ligated with the leading strand fulllength chromatid to create a nonpalindromic fusion (Stohr et al 2010). The higher incidence of templated insertions we detected at intra-chromosomal junctions also suggests an active contribution of DNA synthesis to repair (Lowden et al 2011), conceivably involving the A-NHEJ-associated DNA polymerase theta (Yousefzadeh et al 2014;Ceccaldi et al 2015;Mateos-Gomez et al 2015) or alpha (Reijns et al 2015) by way of incomplete primer removal.…”

Acetylation regulates DNA repair mechanisms in human cells

“…Recent localization of errors in the yeast and human genomes corresponded to synthesis by DNA polymerase a demonstrating that the errorprone patches are not always removed. 55 This suggests that protein acetylation is a likely regulator of fidelity during eukaryotic DNA replication.…”

Crystal Structure of the Human Pol α B Subunit in Complex with the C-terminal Domain of the Catalytic Subunit

“…DNA fragments synthesized by Pol ␣ contain errors because it does not have proofreading exonuclease activity, and, if uncorrected, these errors contribute to the overall fidelity of DNA replication (14). It was estimated that patches synthesized by Pol ␣ comprise ϳ1.5% of newly replicated human genome, making Pol ␣ an important factor modulating genome stability (15).…”