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Anterograde transport of herpes simplex virus capsids in neurons by both separate and married mechanisms.

ABSTRACT: Anterograde transport of herpes simplex virus (HSV) from neuronal cell bodies into, and down, axons is a fundamentally important process for spread to other hosts. Different techniques for imaging HSV in axons have produced two models for how virus particles are transported in axons. In the Separate model, viral nucleocapsids devoid of the viral envelope and membrane glycoproteins are transported in axons. In the Married model, enveloped HSV particles (with the viral glycoproteins) encased within membrane vesicles are transported in the anterograde direction. Earlier studies of HSV-infected human neurons involving electron microscopy (EM) and immunofluorescence staining of glycoproteins and capsids supported the Separate model. However, more-recent live-cell imaging of rat, chicken, and mouse neurons produced evidence supporting the Married model. In a recent EM study, a mixture of Married (75%) and Separate (25%) HSV particles was observed. Here, we studied an HSV recombinant expressing a fluorescent form of the viral glycoprotein gB and a fluorescent capsid protein (VP26), observing that human SK-N-SH neurons contained both Separate (the majority) and Married particles. Live-cell imaging of rat superior cervical ganglion (SCG) neuronal axons in a chamber system (which oriented the axons) also produced evidence of Separate and Married particles. Together, our results suggest that one can observe anterograde transport of both HSV capsids and enveloped virus particles depending on which neurons are cultured and how the neurons are imaged.

SUBMITTER: Wisner TW

PROVIDER: S-EPMC3126305 | biostudies-literature | 2011 Jun

REPOSITORIES: biostudies-literature

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Anterograde transport of herpes simplex virus capsids in neurons by both separate and married mechanisms.

Wisner Todd W TW Sugimoto Ken K Howard Paul W PW Kawaguchi Yasushi Y Johnson David C DC

Journal of virology 20110330 12

Anterograde transport of herpes simplex virus (HSV) from neuronal cell bodies into, and down, axons is a fundamentally important process for spread to other hosts. Different techniques for imaging HSV in axons have produced two models for how virus particles are transported in axons. In the Separate model, viral nucleocapsids devoid of the viral envelope and membrane glycoproteins are transported in axons. In the Married model, enveloped HSV particles (with the viral glycoproteins) encased withi ...[more]

PMID: 21450818

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Project description:Herpes simplex virus replicates in the nucleus, where new capsids are assembled. It produces procapsids devoid of nucleic acid but containing the preVP22a scaffold protein. These thermo-unstable particles then mature into A-, B- or C-nuclear icosahedral capsids, depending on their ability to shed the proteolytically processed scaffold and incorporation of the viral genome. To study how these viral capsids differ, we performed proteomics studies of highly enriched HSV-1 A-, B- and C-nuclear capsids, relying in part on a novel and powerful flow virometry approach to purify C-capsids. We found that the viral particles contained the expected capsid components and identified several tegument proteins in the C-capsid fraction (pUL21, pUL36, pUL46, pUL48, pUL49, pUL50, pUL51 and pUS10). Moreover, numerous ribosomal, hnRNPs and other host proteins, absent from the uninfected controls, were detected on the capsids with some of them seemingly specific to C-capsids (glycogen synthase, four different keratin-related proteins, fibronectin 1 and PCBP1). A subsequent proteomics analysis was performed to rule out the presence of protein complexes that may share similar density as the viral capsids but do not otherwise interact with them. Using pUL25 or VP5 mutant viruses incapable of assembling C-nuclear or all nuclear capsids, respectively, we confirmed the bulk of our initial findings. Naturally, it will next be important to address the functional relevance of these proteins.IMPORTANCE Much is known about the biology of herpesviruses. This includes their unique ability to traverse the two nuclear envelopes by sequential budding and fusion steps. For HSV-1, this implies the pUL31/pUL34 and pUL17/pUL25 complexes that may favor C-capsid egress. However, this selection process is not clear, nor are all the differences that distinguish A-, B- and C-capsids. The present study probes what proteins compose these capsids, including host proteins. This should open up new research avenues to clarify the biology of this most interesting family of viruses. It also reiterates the use of flow virometry as an innovative tool to purify viral particles.

Project description:BackgroundHerpes simplex virus type 1 strain 129 (H129) has represented a promising anterograde neuronal circuit tracing tool, which complements the existing retrograde tracers. However, the current H129 derived tracers are multisynaptic, neither bright enough to label the details of neurons nor capable of determining direct projection targets as monosynaptic tracer.MethodsBased on the bacterial artificial chromosome of H129, we have generated a serial of recombinant viruses for neuronal circuit tracing. Among them, H129-G4 was obtained by inserting binary tandemly connected GFP cassettes into the H129 genome, and H129-ΔTK-tdT was obtained by deleting the thymidine kinase (TK) gene and adding tdTomato coding gene to the H129 genome. Then the obtained viral tracers were tested in vitro and in vivo for the tracing capacity.ResultsH129-G4 is capable of transmitting through multiple synapses, labeling the neurons by green florescent protein, and visualizing the morphological details of the labeled neurons. H129-ΔTK-tdT neither replicates nor spreads in neurons alone, but transmits to and labels the postsynaptic neurons with tdTomato in the presence of complementary expressed TK from a helper virus. H129-ΔTK-tdT is also capable to map the direct projectome of the specific neuron type in the given brain regions in Cre transgenic mice. In the tested brain regions where circuits are well known, the H129-ΔTK-tdT tracing patterns are consistent with the previous results.ConclusionsWith the assistance of the helper virus complimentarily expressing TK, H129-ΔTK-tdT replicates in the initially infected neuron, transmits anterogradely through one synapse, and labeled the postsynaptic neurons with tdTomato. The H129-ΔTK-tdT anterograde monosynaptic tracing system offers a useful tool for mapping the direct output in neuronal circuitry. H129-G4 is an anterograde multisynaptic tracer with a labeling signal strong enough to display the details of neuron morphology.

Dataset Information

Anterograde transport of herpes simplex virus capsids in neurons by both separate and married mechanisms.

Publications

Anterograde transport of herpes simplex virus capsids in neurons by both separate and married mechanisms.

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