A genetically buffered helicase network promotes tolerance of G-quadruplex stabilization in Saccharomyces cerevisiae

Abstract

G-quadruplexes (G4s) are non-canonical DNA secondary structures that can impede DNA replication and transcription and provoke genome instability, and DNA helicases of the PIF1 and RecQ families have long been regarded as the principal enzymes that resolve them. To directly test the relative contributions of these families, we measured the growth of Saccharomyces cerevisiae helicase mutants in the presence of the G4-stabilizing ligand pyridostatin (PDS). Unexpectedly, no single PIF1- or RecQ-family mutant was sensitized to PDS relative to wild type. Sensitivity emerged only in double mutants, and it did so for combinations both within a single family and across the two families. This pattern indicates that G4 tolerance is buffered by the combined, partially interchangeable, activity of multiple helicases rather than by any one family. To ask whether this redundancy extends beyond the canonical players, we tested two additional helicases whose human orthologs are implicated in G4 metabolism: Chl1 (DDX11/ChlR1) and Srs2 (RTEL1). Loss of Chl1 alone did not sensitize cells, and chl1{Delta} combined with PIF1- or RecQ-family mutations recapitulated the redundancy pattern - with one informative exception: chl1{Delta} hrq1{Delta} remained PDS-tolerant, placing Chl1 and Hrq1 in a shared genetic route. In contrast, srs2{Delta} was the sole single mutant sensitized to PDS, defining a non-redundant requirement that no other helicase compensates. We integrate these results into a two-layer model in which a redundant helicase pool resolves G4-associated genomic stress, while a non-redundant Srs2 function manages its recombinogenic consequences. Our findings reframe G4 maintenance from a family-specific activity into a distributed, buffered network. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=79 SRC="FIGDIR/small/737069v1_ufig1.gif" ALT="Figure 1"> View larger version (21K): org.highwire.dtl.DTLVardef@d3db2borg.highwire.dtl.DTLVardef@d9eebborg.highwire.dtl.DTLVardef@1c868aaorg.highwire.dtl.DTLVardef@4f72d6_HPS_FORMAT_FIGEXP M_FIG C_FIG ARTICLE SUMMARYDNA helicases, enzymes that unwind DNA, are thought to dismantle G-quadruplexes (G4s), four-stranded DNA structures that can block DNA metabolism and destabilize genomes. In Saccharomyces cerevisiae, we used the chemical pyridostatin to stabilize G4s and measured the growth of helicase mutants. Losing any single helicase had no effect, but losing two together - even from different helicase families - impaired growth. The protein Chl1 works with the helicase Hrq1 in one shared pathway, while Srs2 is uniquely required on its own. G4 tolerance therefore depends on a redundant network of helicases. These findings interest researchers studying genome stability and related human cancer-predisposition disorders.