RNA-seq Analysis of Host and Viral Gene Expression Highlights Interaction between Varicella Zoster Virus and Keratinocyte Differentiation

Jones, Meleri; Dry, Inga; Frampton, Dan; Singh, Manuraj; Kanda, Ravinder K.; Yee, Michael B.; Kellam, Paul; Hollinshead, Michael; Kinchington, Paul R.; O’Toole, Edel A.; Breuer, Judith

doi:10.1371/journal.ppat.1003896

Cited by 63 publications

(77 citation statements)

References 61 publications

(77 reference statements)

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“…The nTERT cell line has previously been used for studies on human papillomavirus and varicella-zoster virus (18,19). However, at the beginning of this study the efficiency of HSV-1 infection and replication in these cells was unknown.…”

Section: Hsv-1 Replicates With Faster Kinetics In Ntert Keratinocytesmentioning

confidence: 99%

“…Crucially, nTERT cells are still dependent on epidermal growth factor, are able to differentiate, and remain noninvasive upon transplantation, indicating that they retain many characteristics of primary cells. Moreover, they have been used as a successful model system for other epitheliotropic viruses, such as varicella-zoster virus and human papillomavirus (18,19), suggesting they are an appropriate system to study HSV-1 infection of relevant human cells.…”

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confidence: 99%

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Herpes Simplex Virus 1 Enters Human Keratinocytes by a Nectin-1-Dependent, Rapid Plasma Membrane Fusion Pathway That Functions at Low Temperature

Sayers

Elliott

2016

J Virol

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Herpes simplex virus 1 (HSV-1) infects humans through stratified epithelia that are composed primarily of keratinocytes. The route of HSV-1 entry into keratinocytes has been the subject of limited investigation, but it is proposed to involve pH-dependent endocytosis, requiring the gD-binding receptor nectin-1. Here, we have utilized the nTERT human keratinocyte cell line as a new model for dissecting the mechanism of HSV-1 entry into the host. Although immortalized, these cells nonetheless retain normal growth and differentiation properties of primary cells. Using short interfering RNA (siRNA) depletion studies, we confirm that, despite nTERT cells expressing high levels of the alternative gD receptor HVEM, HSV-1 requires nectin-1, not HVEM, to enter these cells. Strikingly, virus entry into nTERT cells occurred with unusual rapidity, such that maximum penetration was achieved within 5 min. Moreover, HSV-1 was able to enter keratinocytes but not other cell types at temperatures as low as 7°C, conditions where endocytosis was shown to be completely inhibited. Transmission electron microscopy of early entry events at both 37°C and 7°C identified numerous examples of naked virus capsids located immediately beneath the plasma membrane, with no evidence of virions in cytoplasmic vesicles. Taken together, these results imply that HSV-1 uses the nectin-1 receptor to enter human keratinocyte cells via a previously uncharacterized rapid plasma membrane fusion pathway that functions at low temperature. These studies have important implications for current understanding of the relationship between HSV-1 and its relevant in vivo target cell. IMPORTANCEThe gold standard of antiviral treatment for any human virus infection is the prevention of virus entry into the host cell. In the case of HSV-1, primary infection in the human begins in the epidermis of the skin or the oral mucosa, where the virus infects keratinocytes, and it is therefore important to understand the molecular events involved in HSV-1 entry into this cell type. Nonetheless, few studies have looked specifically at entry into these relevant human cells. Our results reveal a new route for virus entry that is specific to keratinocytes, involves rapid entry, and functions at low temperatures. This may reflect the environmental conditions encountered by HSV-1 when entering its host through the skin and emphasizes the importance of studying virushost interactions in physiologically relevant cells. H erpes simplex virus type 1 (HSV-1) gains access to its human host through the epidermis at the oral mucosa, skin, or the cornea, where the majority of the cells are keratinocytes. Hence, although HSV-1 replication has been studied in many cell types in vitro, the role of the human keratinocyte in the HSV-1 life cycle makes it the most physiologically relevant cell type in which to unravel HSV-1 replication strategies important in its natural host. To date, there have been only a few studies on the mechanism of HSV-1 entry into human keratinocytes. While initial s...

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Section: Hsv-1 Replicates With Faster Kinetics In Ntert Keratinocytesmentioning

confidence: 99%

mentioning

confidence: 99%

Herpes Simplex Virus 1 Enters Human Keratinocytes by a Nectin-1-Dependent, Rapid Plasma Membrane Fusion Pathway That Functions at Low Temperature

Sayers

Elliott

2016

J Virol

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“…This small intergenic distance combined with almost equal distribution of DNA strand usage (37 genes map to the prototype strand and 34 genes map to the complementary strand) suggests that typical next-generation sequencing information without source strand identification (Jones et al, 2014) provides less information than next-generation sequencing in which mRNA source strand identification is maintained (Baird et al, 2014a). Taken together, our understanding of the HSV-1 and VZV transcriptomes, although significant, is not yet complete.…”

Section: Introductionmentioning

confidence: 99%

“…1). This annotation represents the most current understanding of the VZV proteome, and has formed the basis for analyses of VZV transcripts in mRNA for infected tissue culture cells by PCR-based arrays (Cohrs et al, 2003b), long-oligonucleotide-based arrays (Kennedy et al, 2005), multiplex PCR ) and next-generation-based deep sequencing (Baird et al, 2014a;Jones et al, 2014). Transcripts mapping to all annotated VZV ORFs have been detected in virus-infected cells, but evidence from PCRbased VZV transcription arrays, Northern blot analysis, cDNA PCR (Cohrs et al, 2003b) and strand-specific cDNA deep sequencing (Baird et al, 2014a) suggests that unannotated VZV genes remain.…”

Section: Introductionmentioning

confidence: 99%

A comparison of herpes simplex virus type 1 and varicella-zoster virus latency and reactivation

Kennedy

Rovnak

Badani

et al. 2015

Journal of General Virology

133

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Herpes simplex virus type 1 (HSV-1; human herpesvirus 1) and varicella-zoster virus (VZV; human herpesvirus 3) are human neurotropic alphaherpesviruses that cause lifelong infections in ganglia. Following primary infection and establishment of latency, HSV-1 reactivation typically results in herpes labialis (cold sores), but can occur frequently elsewhere on the body at the site of primary infection (e.g. whitlow), particularly at the genitals. Rarely, HSV-1 reactivation can cause encephalitis; however, a third of the cases of HSV-1 encephalitis are associated with HSV-1 primary infection. Primary VZV infection causes varicella (chickenpox) following which latent virus may reactivate decades later to produce herpes zoster (shingles), as well as an increasingly recognized number of subacute, acute and chronic neurological conditions. Following primary infection, both viruses establish a latent infection in neuronal cells in human peripheral ganglia. However, the detailed mechanisms of viral latency and reactivation have yet to be unravelled. In both cases latent viral DNA exists in an 'end-less' state where the ends of the virus genome are joined to form structures consistent with unit length episomes and concatemers, from which viral gene transcription is restricted. In latently infected ganglia, the most abundantly detected HSV-1 RNAs are the spliced products originating from the primary latency associated transcript (LAT). This primary LAT is an 8.3 kb unstable transcript from which two stable (1.5 and 2.0 kb) introns are spliced. Transcripts mapping to 12 VZV genes have been detected in human ganglia removed at autopsy; however, it is difficult to ascribe these as transcripts present during latent infection as early-stage virus reactivation may have transpired in the post-mortem time period in the ganglia. Nonetheless, low-level transcription of VZV ORF63 has been repeatedly detected in multiple ganglia removed as close to death as possible. There is increasing evidence that HSV-1 and VZV latency is epigenetically regulated. In vitro models that permit pathway analysis and identification of both epigenetic modulations and global transcriptional mechanisms of HSV-1 and VZV latency hold much promise for our future understanding in this complex area. This review summarizes the molecular biology of HSV-1 and VZV latency and reactivation, and also presents future directions for study. Received 5 January 2015Accepted 18 March 2015 IntroductionHerpes simplex virus type 1 (HSV-1; human herpesvirus 1) and varicella-zoster virus (VZV; human herpesvirus 3) are human neurotropic alphaherpesviruses usually acquired early in life. Primary HSV-1 infection is usually localized and may be asymptomatic, although it can produce a more widespread systemic infection in neonates and immunocompromised adults, whilst primary VZV infection is systemic and results in childhood varicella (chickenpox). During primary infection, both viruses gain access to neurons most likely through retrograde transport from the site of cutaneous lesion...

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“…Recently, high throughput RNA sequencing (RNA-seq) technology has been used to profile host transcriptomes during viral infections and diseases [15][16][17][18][19][20]. The application of RNAseq technology is potentially very useful for elucidation of the dynamics of the pathogen genome and the systemic changes in host gene expression in response to infection.…”

Section: Introductionmentioning

confidence: 99%

Insights into the role of immunosenescence during varicella zoster virus infection (shingles) in the aging cell model

Kim

Park

Kumar

et al. 2017

Journal of Dermatological Science

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RNA-seq Analysis of Host and Viral Gene Expression Highlights Interaction between Varicella Zoster Virus and Keratinocyte Differentiation

Cited by 63 publications

References 61 publications

Herpes Simplex Virus 1 Enters Human Keratinocytes by a Nectin-1-Dependent, Rapid Plasma Membrane Fusion Pathway That Functions at Low Temperature

Herpes Simplex Virus 1 Enters Human Keratinocytes by a Nectin-1-Dependent, Rapid Plasma Membrane Fusion Pathway That Functions at Low Temperature

A comparison of herpes simplex virus type 1 and varicella-zoster virus latency and reactivation

Insights into the role of immunosenescence during varicella zoster virus infection (shingles) in the aging cell model

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