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 ess ential heterotrimeric DNA helicase-primase that is responsible for con comitant DNA unwinding and primer synthesis at the viral DNA replicati on fork. The viral single-strand DNA-binding protein (ICP8) can stimul ate DNA unwinding by the helicase-primase as a result of a physical in teraction that is mediated by the UL8 subunit, In this study, we inves tigated the ability of the helicase-primase to unwind a fork-like subs trate that contains an intrastrand d(GpG) DNA cross-link produced by t he antitumor drug cisplatin. We also examined the ability of ICP8 to m odulate the effect of the cisplatin lesion. The data show that the les ion inhibited the helicase-primase when located on the DNA strand alon g which it translocates, However, the lesion did not represent a perma nent obstacle to its progression. In contrast, the adduct did not affe ct 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 damage d DNA, Addition of competitor DNA to helicase reactions led to substan tial reduction of DNA unwinding by the helicase-primase, suggesting th at the enzyme is distributive. ICP8 did not abolish the competition, i ndicating that it did not stimulate the helicase by increasing its pro cessivity, Rather, ICPS may stimulate DNA unwinding and enable bypass of cisplatin damaged DNA by recruiting the helicase-primase to the DNA .