DNA helicases in homologous recombination repair

Helicases are in the spotlight of DNA metabolism and are critical for DNA repair in all domains of life. At their biochemical core, they bind and hydrolyze ATP, converting this energy to translocate unidirectionally, with different strand polarities and substrate binding specificities, along one strand of a nucleic acid. In doing so, DNA and RNA helicases separate duplex strands or remove nucleoprotein complexes, affecting DNA repair and the architecture of replication forks. In this review, we focus on recent advances on the roles and regulations of DNA helicases in homologous recombination repair, a critical pathway for mending damaged chromosomes and for ensuring genome integrity.

Introduction

Homologous recombination (HR) is critical for normal development and for stable propagation of the newly replicated genome. Defects in this process give rise to debilitating disorders, including predisposition to cancers, premature aging, reduced fertility and other congenital and developmental defects [1,2]. Functionally, HR is required to repair double strand breaks (DSBs), which arise due to replication errors or as a result of exogenous and endogenous DNA damage. In addition, HR factors stabilize stalled replication forks and guide filling of gaps arising during replication to the newly synthesized chromatid to facilitate replication completion [3]. The presence of daughter-strand gaps, rather than stalled or collapsed forks or DSBs, has recently emerged as the underlying feature of HR-defective cancers and of their chemotherapeutic sensitivity [4]. HR is most of the time error-free in outcome as the newly synthesized chromatid or the homologous chromosome is used as template for DNA repair [3]. However, in certain cases, HR can cause genome rearrangement and instability, for instance when induced at DNA repeat elements [5]. Importantly, formation, maturation and processing of HR intermediates, induced by both breaks and stalled replication forks, is intricately mediated or reversed by DNA helicases, which thus play critical roles in genome stability [6,7].

HR repair can be divided in several pathways, and the underlying mechanisms include gene conversion, when both sides of the break are homologous to the donor and participate in repair, and break-induced replication, when only one end of the break can find a homologous template [5]. The latter pathway of break-induced replication, recently discussed in a dedicated review [8], relates to situations arising at eroded telomeres or when replication forks collapse at nicked DNA. Another critical context for HR repair is the one of gap-filling by template switching, in which replication-associated DNA gaps ensured by repriming downstream the lesion [9,10] are being engaged by postreplicative repair coordinated by PCNA polyubiquitylation and HR factors. In HR-mediated gap-filling, the information from the newly synthesized sister chromatid is used to bypass the DNA damage postreplicatively [3,11^••]. In addition, HR factors play roles in stabilizing and restarting stalled replication forks through fork remodeling [12].

Several steps are common to HR repair pathways and include: (1) resection of DSB ends or extension of the gaps before gap-filling; (2) assembly of the Rad51 helical filament known as the presynaptic complex; (3) homology search and strand invasion with the formation of a D-loop (Figure 1). In gene conversion, the D-loop is extended via DNA synthesis after which it can be disassembled by DNA helicases in a process known as synthesis-dependent strand annealing (SDSA) leading to noncrossover recombinants (Figure 2). Alternative to SDSA, D-loop extension can be combined with second end capture, leading to the formation of a double Holliday junction (dHJ) (Figure 2) or a pseudo-dHJ intermediate if the context is gap-filling. Subsequently, dHJs are either dissolved by a helicase-topoisomerase complex (Sgs1-Top3-Rmi1, known as STR in budding yeast, and BLM-TOPIIIα-RMI1-RMI2 in vertebrates) to noncrossovers or resolved by structure-specific endonucleases, with the potential of forming crossovers [13] (Figure 2). Importantly, most steps involved in HR repair rely on the action of DNA helicases (Figure 1, Figure 2).

Basic functions of DNA helicase and how they affect various or specific HR steps have been recently described in a review [14]. Here, we focus on recent insights on how DNA helicases modulate HR, how their activity is influenced by posttranslational modifications and interaction with other proteins, and how in certain cases the helicase activity modulation is context or cell cycle specific. Although we occasionally refer to recently gained insights from other model systems, we largely focus on recent work in budding yeast related to helicase-mediated repair of DSBs and filling of daughter-strand gaps arising during replication as a consequence of repriming.