title: Molecular mechanism of G-quadruplex unwinding helicase: sequential and repetitive unfolding of G-quadruplex by Pif1 helicase.
abstract: Recent advances in G-quadruplex (G4) studies have confirmed that G4 structures exist in living cells and may have detrimental effects on various DNA transactions. How helicases resolve G4, however, has just begun to be studied and remains largely unknown. In the present paper, we use single-molecule fluorescence assays to probe Pif1-catalysed unfolding of G4 in a DNA construct resembling an ongoing synthesis of lagging strand stalled by G4. Strikingly, Pif1 unfolds and then halts at the ss/dsDNA junction, followed by rapid reformation of G4 and 'acrobatic' re-initiation of unfolding by the same monomer. Thus, Pif1 unfolds single G4 structures repetitively. Furthermore, it is found that Pif1 unfolds G4 sequentially in two large steps. Our study has revealed that, as a stable intermediate, G-triplex (G3) plays an essential role in this process. The repetitive unfolding activity may facilitate Pif1 disrupting the continuously reforming obstructive G4 structures to rescue a stalled replication fork. The proposed mechanism for step-wise unfolding of G4 is probably applicable to other helicases that resolve G4 structures for maintaining genome stability.
authors.affiliation: *College of Life Sciences, Northwest A & F University, Xi'an, Shaanxi 712100, China.
authors.forename: Xi-Miao
authors.initials: XM
authors.lastname: Hou
authors.affiliation: *College of Life Sciences, Northwest A & F University, Xi'an, Shaanxi 712100, China.
authors.forename: Wen-Qiang
authors.initials: WQ
authors.lastname: Wu
authors.affiliation: *College of Life Sciences, Northwest A & F University, Xi'an, Shaanxi 712100, China.
authors.forename: Xiao-Lei
authors.initials: XL
authors.lastname: Duan
authors.affiliation: *College of Life Sciences, Northwest A & F University, Xi'an, Shaanxi 712100, China.
authors.forename: Na-Nv
authors.initials: NN
authors.lastname: Liu
authors.affiliation: *College of Life Sciences, Northwest A & F University, Xi'an, Shaanxi 712100, China.
authors.forename: Hai-Hong
authors.initials: HH
authors.lastname: Li
authors.affiliation: *College of Life Sciences, Northwest A & F University, Xi'an, Shaanxi 712100, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Fu
authors.affiliation: †Beijing National Laboratory for Condensed Matter Physics and CAS Key Laboratory of Soft Matter Physics, Institute of Physics, Chinese Academy of Sciences, Beijing 100190, China.
authors.forename: Shuo-Xing
authors.initials: SX
authors.lastname: Dou
authors.affiliation: †Beijing National Laboratory for Condensed Matter Physics and CAS Key Laboratory of Soft Matter Physics, Institute of Physics, Chinese Academy of Sciences, Beijing 100190, China.
authors.forename: Ming
authors.initials: M
authors.lastname: Li
authors.affiliation: *College of Life Sciences, Northwest A & F University, Xi'an, Shaanxi 712100, China.
authors.forename: Xu-Guang
authors.initials: XG
authors.lastname: Xi
mesh_terms: D055162:Biocatalysis; D004265:DNA Helicases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004261:DNA Replication; D004277:DNA, Single-Stranded / Q000737:chemistry* / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes; D054856:G-Quadruplexes; D015870:Gene Expression; D016681:Genome, Fungal; D042822:Genomic Instability; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D013194:Staining and Labeling
chemical_list: D004277:DNA, Single-Stranded; D005456:Fluorescent Dyes; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; C085463:PIF1 protein, S cerevisiae; D004265:DNA Helicases