Break-induced replication requires all essential DNA replication factors except those specific for pre-RC assembly

Abstract

Break-induced replication (BIR) is an efficient homologous recombination (HR) pathway employed to repair a DNA double-strand break (DSB) when homology is restricted to one end. All three major replicative DNA polymerases are required for BIR, including the otherwise nonessential Pol32 subunit. Here we show that BIR requires the replicative DNA helicase (Cdc45, the GINS, and Mcm2-7 proteins) as well as Cdt1. In contrast, both subunits of origin recognition complex (ORC) and Cdc6, which are required to create a prereplication complex (pre-RC), are dispensable. The Cdc7 kinase, required for both initiation of DNA replication and post-replication repair (PRR), is also required for BIR. Ubiquitination and sumoylation of the DNA processivity clamp PCNA play modest roles; in contrast, PCNA alleles that suppress pol32Δ's cold sensitivity fail to suppress its role in BIR, and are by themselves dominant inhibitors of BIR. These results suggest that origin-independent BIR involves cross-talk between normal DNA replication factors and PRR. © 2010 by Cold Spring Harbor Laboratory Press.