RT Journal Article
SR Electronic
T1 A simple bypass assay for DNA polymerases shows hypermutating variants associated with cancer show mechanistic differences in vitro
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2022.01.10.475213
DO 10.1101/2022.01.10.475213
A1 Gilles Crevel
A1 Stephen Kearsey
A1 Sue Cotterill
YR 2022
UL http://biorxiv.org/content/early/2022/01/10/2022.01.10.475213.abstract
AB Errors made by DNA polymerases contribute to both natural variation and, in extreme cases, to genome instability and its associated diseases. Recently the importance of polymerase misincorporation in disease has been highlighted by the identification of cancer-associated polymerase variants and the recognition that a subgroup of these variants have a hypermutation phenotype in tumours. We have developed a bypass assay to rapidly determine the tendency of a polymerase to misincorporate in vitro. We have used the assay to compare misincorporation by wild-type, exonuclease defective and two hypermutating DNA polymerase e variants, P286R and V411L. The assay clearly distinguished between the misincorporation rates of wild type, exonuclease dead and P286R polymerases. However, the V411L polymerase showed different misincorporation characteristics to P286R, suggesting that these variants cause hypermutation by different mechanisms. Using this assay misincorporation opposite a templated C nucleotide was consistently higher than for other nucleotides, and this caused predominantly C to T transitions. This is consistent with the observation that C to T transitions are commonly seen in POLE mutant tumours.Competing Interest StatementThe authors have declared no competing interest.