Pfh1 Is an Accessory Replicative Helicase that Interacts with the Replisome to Facilitate Fork Progression and Preserve Genome Integrity

• DOI: 10.1371/journal.pgen.1006238
• Publication Date: 2016-09-09T18:44:19Z
• Authors: Karin R. McDonald; Amanda J. Guise; Parham Pourbozorg...; Ileana M. Cristea; Virginia A. Zakian; John A. Capra; Nasim Sabouri
• Keywords: Replisome; Origin recognition complex; Pre-replication complex

Replicative DNA helicases expose the two strands of double helix to replication apparatus, but accessory are often needed help forks move past naturally occurring hard-to-replicate sites, such as tightly bound proteins, RNA/DNA hybrids, and secondary structures. Although Schizosaccharomyces pombe 5'-to-3' helicase Pfh1 is known promote fork progression, its genomic targets, dynamics, mechanisms action largely unknown. Here we address these questions by integrating genome-wide identification binding comprehensive analysis effects depletion on damage, proteomic interaction partners immunoaffinity purification mass spectrometry. Of 621 high confidence Pfh1-binding sites in wild type cells, about 40% were slowing (as marked polymerase occupancy) and/or damage levels phosphorylated H2A). The integrity tRNA 5S rRNA genes, highly transcribed RNA II nucleosome depleted regions particularly Pfh1-dependent. association with at genes was S phase dependent, thus unlikely be an artifact transcription rates. affected suppressed discrete throughout genome, replicative similar extents both Pfh1-dependent independent suggesting that proximal machinery during phase. Consistent this interpretation, co-purified many key replisome components, including hexameric MCM helicase, polymerases, RPA, processivity clamp PCNA dependent manner. Thus, conclude interacts promotes suppresses sites. These data provide insight into which evolutionarily conserved helps preserve genome integrity.