Fidelity of DNA replication – cancer associated substitutions in DNA polymerase epsilon

Mutations in the proofreading domain of DNA Polymerase epsilon (builds half the human genome) have recently been defined as drivers in tumorigenesis. Here we ask whether mutations located elsewhere in Pol epsilon should be defined as drivers.

Two substitutions, located in the finger and thumb domain, were previously found associated with a high tumor mutation burden in a comprehensive analysis of 80000 tumors.

We propose that high-resolution structures, biochemical studies of processivity, nucleotide selectivity and proofreading activity will clarify whether the fidelity of Pol epsilon is affected in these two cases.

In addition, residues essential for the interaction between Pol epsilon and the processivity clamp, PCNA, will be identified. It is known that a reduced processivity of Pol epsilon results in a higher mutation rate. Will a loss of interaction between Pol epsilon and PCNA also lead to an increased mutation rate?

A novel in vitro replication assay will be developed, in which the fidelity of cancer associated variants in Pol epsilon can be studied in the context of the replisome.

Furthermore, this assay will be used to study the contribution of extrinsic proofreading to the overall fidelity when DNA is duplicated.

This hypothesis driven basic research project may have clinical implications in the diagnosis, prognosis and treatment of various cancers since driver mutations in Pol epsilon already have been suggested to be an indication for immune checkpoint therapy.