Polymerase-mediated ultramutagenesis in mice produces diverse cancers with high mutational load

Abstract

Mutations underlie all cancers, and their identification and study are the foundation of cancer biology. We describe what we believe to be a novel approach to mutagenesis and cancer studies based on the DNA polymerase ε (POLE) ultramutator phenotype recently described in human cancers, in which a single amino acid substitution (most commonly P286R) in the proofreading domain results in error-prone DNA replication. We engineered a conditional PoleP286R allele in mice. PoleP286R/+ embryonic fibroblasts exhibited a striking mutator phenotype and immortalized more efficiently. PoleP286R/+ mice were born at Mendelian ratios but rapidly developed lethal cancers of diverse lineages, yielding the most cancer-prone monoallelic model described to date, to our knowledge. Comprehensive whole-genome sequencing analyses showed that the cancers were driven by high base substitution rates in the range of human cancers, overcoming a major limitation of previous murine cancer models. These data establish polymerase-mediated ultramutagenesis as an efficient in vivo approach for the generation of diverse animal cancer models that recapitulate the high mutational loads inherent to human cancers.

DNA polymerase δ proofreads errors made by DNA polymerase ε

“…Mutations in the POLE gene, which encodes the catalytic subunit of Pole in humans, are found in 5-8% of sporadic colorectal and endometrial cancers and define a unique subset of these cancers with a so-called ultramutated phenotype (62). The POLE mutations predominantly affect the exonuclease domain of Pole and cause strong mutator and cancer susceptibility phenotypes in model systems (39,63,64). Although MMR defects are also common in colorectal and endometrial tumors, strong POLE mutators are never seen in combination with MMR deficiency, suggesting that MMR is critical to keep the mutation rate at a level compatible with cell survival.…”

Mismatch repair and DNA polymerase δ proofreading prevent catastrophic accumulation of leading strand errors in cells expressing a cancer-associated DNA polymerase ϵ variant

“…This is 50-fold higher than the mutator effect of Polϵ proofreading deficiency and also overwhelmingly exceeds the effect of any previously studied Polϵ mutation. Furthermore, Pole P286R mice are dramatically more cancer-prone than mice deficient in Polϵ proofreading and, in fact, more cancer-prone than any existing monoallelic animal model ( 24 , 25 ).…”

Excessive excision of correct nucleotides during DNA synthesis explained by replication hurdles

“…Exo-site mutants of replicative Pol e and Pol d DNAPs are found in tumors, and many of them show an ultramutational phenotype (Albertson et al, 2009). Interestingly, the ultramutational phenotype is not explained entirely by the loss of proofreading activity (Kane & Shcherbakova, 2014;Li et al, 2018), suggesting that there are additional roles of the exonuclease activity that prevent mutagenicity.…”