Genome wide nucleosome mapping for HSV-1 shows nucleosomes are deposited at preferred positions during lytic infection.

Jaewook Oh,John W Tobias,Eric Z Chen,Hongzhe Li,Sindura Penubarthi,Nigel W Fraser,Hao Sun,Iryna F Sanders,Don A Baldwin,R Benjamin Isett,Michael Nevels

doi:10.1371/journal.pone.0117471

Abstract

HSV is a large double stranded DNA virus, capable of causing a variety of diseases from the common cold sore to devastating encephalitis. Although DNA within the HSV virion does not contain any histone protein, within 1 h of infecting a cell and entering its nucleus the viral genome acquires some histone protein (nucleosomes). During lytic infection, partial micrococcal nuclease (MNase) digestion does not give the classic ladder band pattern, seen on digestion of cell DNA or latent viral DNA. However, complete digestion does give a mono-nucleosome band, strongly suggesting that there are some nucleosomes present on the viral genome during the lytic infection, but that they are not evenly positioned, with a 200bp repeat pattern, like cell DNA. Where then are the nucleosomes positioned? Here we perform HSV-1 genome wide nucleosome mapping, at a time when viral replication is in full swing (6hr PI), using a microarray consisting of 50mer oligonucleotides, covering the whole viral genome (152kb). Arrays were probed with MNase-protected fragments of DNA from infected cells. Cells were not treated with crosslinking agents, thus we are only mapping tightly bound nucleosomes. The data show that nucleosome deposition is not random. The distribution of signal on the arrays suggest that nucleosomes are located at preferred positions on the genome, and that there are some positions that are not occupied (nucleosome free regions -NFR or Nucleosome depleted regions -NDR), or occupied at frequency below our limit of detection in the population of genomes. Occupancy of only a fraction of the possible sites may explain the lack of a typical MNase partial digestion band ladder pattern for HSV DNA during lytic infection. On average, DNA encoding Immediate Early (IE), Early (E) and Late (L) genes appear to have a similar density of nucleosomes.

Highlights

Herpes simplex virus (HSV-1) is a large double stranded DNA virus (152kb genome)
Our data show that the HSV-1 genome is partially covered by nucleosomes at 6h Post Infection (PI) in SY5Y tissue culture cells explaining the lack of micrococcal nuclease (MNase) partial digest band pattern
The distribution of nucleosomes in the population of viral genomes is not random because distinct regions of the genome exist where we detect nucleosomes and there are other regions that are nucleosome free (NFR). This is similar to cellular genes which are being expressed where nucleosome free regions (NFRs) regions are seen

Summary

Introduction

Herpes simplex virus (HSV-1) is a large double stranded DNA virus (152kb genome). On infecting a host it forms lytic infections in epithelial tissue, from which it can spread to the innervating sensory neurons of the peripheral nervous system where it can establish a life long latent infection, from which it can be reactivated to cause recurrent disease (for review see: [1]). DNA viruses protect their genomes in compact protein structures called capsids. Small polyoma and SV40 virions have been shown to contain nucleosomes in their capsids [2,3]. Larger Adenovirus does not have nucleosomes in its capsid, but acquires them during lytic infection [4]. Small DNA viruses contain nucleosomes in their capsids, HSV, like Adenovirus, does not [5,6]. HSV DNA in virions has its charge partly neutralized by polyamines [5]. Following infection of tissue culture cells and entry of the cell nucleus, nucleosomes are deposited on HSV DNA within 1 hour [7], at which time the linear viral genome is circularized and gene expression initiated [8]

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