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Herpes simplex virus 1 Us3 deletion mutant is infective despite impaired capsid translocation to the cytoplasm.

Wild P, Leisinger S, de Oliveira AP, Schraner EM, Kaech A, Ackermann M, Tobler K - Viruses (2015)

Bottom Line: Determination of phenotype distribution by quantitative electron microscopy, and calculation per mean nuclear or cell volume revealed the following: (i) The number of R7041(∆US3) capsids budding at the inner nuclear membrane was significantly higher than that of wild type HSV-1; (ii) The mean number of R7041(∆US3) virions per mean cell volume was 2726, that of HSV-1 virions 1460 by 24 h post inoculation; (iii) 98% of R7041(∆US3) virions were in the perinuclear space; (iv) The number of R7041(∆US3) capsids in the cytoplasm, including those budding at Golgi membranes, was significantly reduced.Cell associated R7041(∆US3) yields were 2.37×10(8) and HSV-1 yields 1.57×10(8) PFU/mL by 24 h post inoculation.We thus conclude that R7041(∆US3) virions, which acquire envelope and tegument by budding at the inner nuclear membrane into the perinuclear space, are infective.

View Article: PubMed Central - PubMed

Affiliation: Institute of Veterinar Anatomy, Winterthurerstrasse 260, CH-8057 Zürich, Switzerland. pewild@access.uzh.ch.

ABSTRACT
Herpes simplex virus 1 (HSV-1) capsids are assembled in the nucleus bud at the inner nuclear membrane into the perinuclear space, acquiring envelope and tegument. In theory, these virions are de-enveloped by fusion of the envelope with the outer nuclear membrane and re-enveloped by Golgi membranes to become infective. Us3 enables the nucleus to cytoplasm capsid translocation. Nevertheless, Us3 is not essential for the production of infective progeny viruses. Determination of phenotype distribution by quantitative electron microscopy, and calculation per mean nuclear or cell volume revealed the following: (i) The number of R7041(∆US3) capsids budding at the inner nuclear membrane was significantly higher than that of wild type HSV-1; (ii) The mean number of R7041(∆US3) virions per mean cell volume was 2726, that of HSV-1 virions 1460 by 24 h post inoculation; (iii) 98% of R7041(∆US3) virions were in the perinuclear space; (iv) The number of R7041(∆US3) capsids in the cytoplasm, including those budding at Golgi membranes, was significantly reduced. Cell associated R7041(∆US3) yields were 2.37×10(8) and HSV-1 yields 1.57×10(8) PFU/mL by 24 h post inoculation. We thus conclude that R7041(∆US3) virions, which acquire envelope and tegument by budding at the inner nuclear membrane into the perinuclear space, are infective.

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One-Step growth curves of R7041(∆Us3) and wt HSV-1. Cells were harvested at times post inoculation as indicated. Infectious virus yields in pellets and supernatants were determined by plaque titration. Mean and standard deviation were calculated from 4 experiments.
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viruses-07-00052-f012: One-Step growth curves of R7041(∆Us3) and wt HSV-1. Cells were harvested at times post inoculation as indicated. Infectious virus yields in pellets and supernatants were determined by plaque titration. Mean and standard deviation were calculated from 4 experiments.

Mentions: HSV-1∆Us3 virions are infective though with delayed onset and reduced peaks of virus titers in HEp-2 cells [16] but with similar titers to wt HSV-1 in Vero cells [20]. Virus replication assays revealed also delay of R7041(∆Us3) growth (Figure 12). Infective virions were to a large extent cell associated. The mean total yield of R7041(∆Us3) at 24 hpi was 2.368 × 108 PFU/mL, that of wt HSV-1 4.54 × 108 PFU/mL. The intracellular yield of R7041(∆Us3) was 2.366 × 108 PFU/mL, that of wt HSV-1 1.571 × 108 PFU/mL, i.e., 1.5 times higher. The total number of R7041(∆Us3) virus particles (capsids budding at any membrane as well as capsids in the cytoplasm and all virions) was equal to the total number of wt HSV-1 virus particles at 24 hpi (Figure 13). However, the total number of R7041(∆Us3) virions was almost twice as high as that of wt HSV-1 virions by 24 hpi. 97.7% of R7041(∆Us3) virions were localized in the PNS at 24 hpi. Therefore, we conclude that (i) R7041(∆Us3) virions in the PNS are infective, as proposed earlier [44]; and (ii) almost all infectious virions acquire envelope and tegument including all essential proteins by budding at the INM.


Herpes simplex virus 1 Us3 deletion mutant is infective despite impaired capsid translocation to the cytoplasm.

Wild P, Leisinger S, de Oliveira AP, Schraner EM, Kaech A, Ackermann M, Tobler K - Viruses (2015)

One-Step growth curves of R7041(∆Us3) and wt HSV-1. Cells were harvested at times post inoculation as indicated. Infectious virus yields in pellets and supernatants were determined by plaque titration. Mean and standard deviation were calculated from 4 experiments.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4306828&req=5

viruses-07-00052-f012: One-Step growth curves of R7041(∆Us3) and wt HSV-1. Cells were harvested at times post inoculation as indicated. Infectious virus yields in pellets and supernatants were determined by plaque titration. Mean and standard deviation were calculated from 4 experiments.
Mentions: HSV-1∆Us3 virions are infective though with delayed onset and reduced peaks of virus titers in HEp-2 cells [16] but with similar titers to wt HSV-1 in Vero cells [20]. Virus replication assays revealed also delay of R7041(∆Us3) growth (Figure 12). Infective virions were to a large extent cell associated. The mean total yield of R7041(∆Us3) at 24 hpi was 2.368 × 108 PFU/mL, that of wt HSV-1 4.54 × 108 PFU/mL. The intracellular yield of R7041(∆Us3) was 2.366 × 108 PFU/mL, that of wt HSV-1 1.571 × 108 PFU/mL, i.e., 1.5 times higher. The total number of R7041(∆Us3) virus particles (capsids budding at any membrane as well as capsids in the cytoplasm and all virions) was equal to the total number of wt HSV-1 virus particles at 24 hpi (Figure 13). However, the total number of R7041(∆Us3) virions was almost twice as high as that of wt HSV-1 virions by 24 hpi. 97.7% of R7041(∆Us3) virions were localized in the PNS at 24 hpi. Therefore, we conclude that (i) R7041(∆Us3) virions in the PNS are infective, as proposed earlier [44]; and (ii) almost all infectious virions acquire envelope and tegument including all essential proteins by budding at the INM.

Bottom Line: Determination of phenotype distribution by quantitative electron microscopy, and calculation per mean nuclear or cell volume revealed the following: (i) The number of R7041(∆US3) capsids budding at the inner nuclear membrane was significantly higher than that of wild type HSV-1; (ii) The mean number of R7041(∆US3) virions per mean cell volume was 2726, that of HSV-1 virions 1460 by 24 h post inoculation; (iii) 98% of R7041(∆US3) virions were in the perinuclear space; (iv) The number of R7041(∆US3) capsids in the cytoplasm, including those budding at Golgi membranes, was significantly reduced.Cell associated R7041(∆US3) yields were 2.37×10(8) and HSV-1 yields 1.57×10(8) PFU/mL by 24 h post inoculation.We thus conclude that R7041(∆US3) virions, which acquire envelope and tegument by budding at the inner nuclear membrane into the perinuclear space, are infective.

View Article: PubMed Central - PubMed

Affiliation: Institute of Veterinar Anatomy, Winterthurerstrasse 260, CH-8057 Zürich, Switzerland. pewild@access.uzh.ch.

ABSTRACT
Herpes simplex virus 1 (HSV-1) capsids are assembled in the nucleus bud at the inner nuclear membrane into the perinuclear space, acquiring envelope and tegument. In theory, these virions are de-enveloped by fusion of the envelope with the outer nuclear membrane and re-enveloped by Golgi membranes to become infective. Us3 enables the nucleus to cytoplasm capsid translocation. Nevertheless, Us3 is not essential for the production of infective progeny viruses. Determination of phenotype distribution by quantitative electron microscopy, and calculation per mean nuclear or cell volume revealed the following: (i) The number of R7041(∆US3) capsids budding at the inner nuclear membrane was significantly higher than that of wild type HSV-1; (ii) The mean number of R7041(∆US3) virions per mean cell volume was 2726, that of HSV-1 virions 1460 by 24 h post inoculation; (iii) 98% of R7041(∆US3) virions were in the perinuclear space; (iv) The number of R7041(∆US3) capsids in the cytoplasm, including those budding at Golgi membranes, was significantly reduced. Cell associated R7041(∆US3) yields were 2.37×10(8) and HSV-1 yields 1.57×10(8) PFU/mL by 24 h post inoculation. We thus conclude that R7041(∆US3) virions, which acquire envelope and tegument by budding at the inner nuclear membrane into the perinuclear space, are infective.

Show MeSH
Related in: MedlinePlus