Replication and Recombination of Herpes Simplex Virus DNA

Department of Medical Biochemistry and Cell Biology, Institute of Biomedicine, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden.
Journal of Biological Chemistry (Impact Factor: 4.57). 03/2011; 286(18):15619-24. DOI: 10.1074/jbc.R111.233981
Source: PubMed


Replication of herpes simplex virus takes place in the cell nucleus and is carried out by a replisome composed of six viral proteins: the UL30-UL42 DNA polymerase, the UL5-UL8-UL52 helicase-primase, and the UL29 single-stranded DNA-binding protein ICP8. The replisome is loaded on origins of replication by the UL9 initiator origin-binding protein. Virus replication is intimately coupled to recombination and repair, often performed by cellular proteins. Here, we review new significant developments: the three-dimensional structures for the DNA polymerase, the polymerase accessory factor, and the single-stranded DNA-binding protein; the reconstitution of a functional replisome in vitro; the elucidation of the mechanism for activation of origins of DNA replication; the identification of cellular proteins actively involved in or responding to viral DNA replication; and the elucidation of requirements for formation of replication foci in the nucleus and effects on protein localization.

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    • "ICP8 also binds to RNA albeit with reduced affinity compared to its ssDNA binding activity [19]. Functionally, ICP8 performs a critical role at the viral DNA replication fork by maintaining ssDNA regions and by interacting with various other replisome components via specific protein–protein interactions [reviewed in [20] [21] [22] [23] [24]]. In addition, ICP8 has been demonstrated to function as a recombinase, capable of promoting strand annealing and strand invasion as well as mediating strand exchange reactions in conjunction with the HSV-1 helicase-primase or UL12 nuclease [25] [26] [27] [28]. "
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    ABSTRACT: We have adapted the thermal shift assay to measure the ligand binding properties of the herpes simplex virus-I single-strand DNA binding protein, ICP8. By measuring SYPRO Orange fluorescence in microtiter plates using a fluorescence-enabled thermal cycler, we have quantified the effects of oligonucleotide ligands on the melting temperature of ICP8. We found that single-stranded oligomers raise the melting temperature of ICP8 in a length- and concentration-dependent manner, ranging from I C for (dT)(5) to a maximum of 9 C with oligomers >= 10 nucleotides, with an apparent K-d of <1 mu M for (dT)(20). Specifically, the results indicate that ICP8 is capable of interacting with oligomers as short as 5 nucleotides. Moreover, the observed increases in melting temperature of up to 9 degrees C, indicates that single-strand DNA binding significantly stabilizes the structure of ICP8. This assay may be applied to investigate the ligand binding proteins of other single-strand DNA binding proteins and used as a high-throughput screen to identify compounds with therapeutic potential that inhibit single-strand DNA binding. As proof of concept, the single-strand DNA binding agent ciprofloxacin reduces the ligand induced stabilization of the melting temperature of ICP8 in a dose-dependent manner.
    Preview · Article · Nov 2014 · Biochemical and Biophysical Research Communications
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    • "Following nuclear entry, the linear genome circularizes before the advent of viral protein production (Garber et al., 1993; Strang & Stow, 2005). During lytic infection, early genome replication is achieved by a theta replication mechanism initiated at three redundant origins of replication (two copies of oriS and one copy of oriL) but rolling circle replication later predominates, forming ‘endless’ DNA lacking termini (reviewed by Boehmer & Nimonkar, 2003; Muylaert et al., 2011). This concatemeric DNA is later cleaved to monomeric units during packaging into newly formed capsids. "
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