Herpes simplex virus type-1 single-strand DNA-binding protein (ICP8) enhances the ability of the viral DNA helicase-primase to unwind cisplatin-modified DNA.
The herpes simplex virus type-1 UL5, UL8, and UL52 genes encode an essential heterotrimeric DNA helicase-primase that is responsible for concomitant DNA unwinding and primer synthesis at the viral DNA replication fork. The viral single-strand DNA-binding protein (ICP8) can stimulate DNA unwinding by the helicase-primase as a result of a physical interaction that is mediated by the UL8 subunit. In this study, we investigated the ability of the helicase-primase to unwind a fork-like substrate that contains an intrastrand d(GpG) DNA cross-link produced by the antitumor drug cisplatin. We also examined the ability of ICP8 to modulate the effect of the cisplatin lesion. The data show that the lesion inhibited the helicase-primase when located on the DNA strand along which it translocates. However, the lesion did not represent a permanent obstacle to its progression. In contrast, the adduct did not affect the helicase-primase when located on the opposite DNA strand. ICP8 specifically stimulated DNA unwinding by the helicase-primase. Coating concentrations of ICP8 were necessary for optimal unwinding of damaged DNA. Addition of competitor DNA to helicase reactions led to substantial reduction of DNA unwinding by the helicase-primase, suggesting that the enzyme is distributive. ICP8 did not abolish the competition, indicating that it did not stimulate the helicase by increasing its processivity. Rather, ICP8 may stimulate DNA unwinding and enable bypass of cisplatin damaged DNA by recruiting the helicase-primase to the DNA.
Tanguy Le Gac
N
N
Institut de Pharmacologie et de Biologie Structurale, CNRS, 205 route de Narbonne, 31077 Toulouse cedex, France.
Villani
G
G
Boehmer
P E
PE
D001483:Base Sequence; D002945:Cisplatin / Q000378:metabolism* / Q000494:pharmacology*; D018736:DNA Adducts / Q000378:metabolism*; D004249:DNA Damage; D004265:DNA Helicases / Q000378:metabolism*; D019915:DNA Primase; D004264:DNA Topoisomerases, Type I / Q000378:metabolism*; D004279:DNA, Viral / Q000187:drug effects*; D004268:DNA-Binding Proteins; D018259:Herpesvirus 1, Human / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D013379:Substrate Specificity; D014764:Viral Proteins / Q000378:metabolism*
D018736:DNA Adducts; D004279:DNA, Viral; D004268:DNA-Binding Proteins; C045462:ICP8 protein, Simplexvirus; D014764:Viral Proteins; C057270:cisplatin-DNA adduct; D019915:DNA Primase; C062021:helicase-primase, Human herpesvirus 1; D004265:DNA Helicases; D004264:DNA Topoisomerases, Type I; D002945:Cisplatin