# The genetic and biochemical basis of human leading strand synthesis

## TL;DR

This study reveals how DNA Polymerase Epsilon functions during DNA replication by identifying key factors and mechanisms that support its activity on the leading strand.

## Contribution

The paper introduces a two-tier mechanism involving PCNA loading and dsDNA binding to explain Polε processivity on the leading strand.

## Key findings

- Genes involved in iron metabolism are essential for sustaining ISC-dependent Polε activity.
- A synthetic lethal interaction exists between POLE3-POLE4 and the CHTF18-RFC2/5 complex.
- Two tiers of regulation are required for Polε processivity: PCNA loading and dsDNA gripping.

## Abstract

The maintenance of genome stability requires efficient leading strand synthesis by DNA Polymerase Epsilon (Polε). By performing CRISPR genetic screens in cells lacking the POLE4 subunit of Polε we define a genetic map of the factors required to support Polε function in the absence of its accessory subunits. A set of genes involved in iron metabolism emerge as required to sustain Iron Sulphur Cluster (ISC)-dependent Polε activity. We then dissect a synthetic lethal interaction between POLE3-POLE4 and the CHTF18-RFC2/5 complex. By combining cell biology, structural modelling and biochemistry, we define the existence of two tiers of regulation of Polε processivity: leading strand-specific loading of PCNA by CHTF18-RFC2/5 and “gripping” of newly synthesised dsDNA by POLE3-POLE4. The combined loss of these functions is incompatible with leading strand synthesis and viability. In summary, we describe the biochemical basis of human leading strand synthesis and the consequence of its dysfunction in genome stability.

How DNA Polymerase Epsilon accomplishes continuous leading strand synthesis during DNA replication is not understood. Here, the authors describe a two tiers mechanism required to sustain Pol Epsilon processivity: CHTF18-dependent loading of PCNA at leading strand and dsDNA binding by its POLE3-POLE4 subunits.