A pentameric protein ring with novel architecture is required for herpesviral packaging

Didychuk, Allison L.; Gates, S.N.; Gardner, Matthew R; Strong, Lisa M; Martin, Andreas; Glaunsinger, Britt A.

doi:10.7554/elife.62261

Cited by 11 publications

(14 citation statements)

References 68 publications

(135 reference statements)

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“…2B). These banding patterns of cleaved TR regions were in agreement with previous data which characterized an ORF68-KO reported by Glaunsinger et al (27, 28). Our data indicated that ORF7 plays a critical role in TR cleavage during lytic infection-induced viral DNA replication.…”

Section: Resultssupporting

confidence: 93%

“…Next, we asked whether the KSHV ORF7 cleaves the KSHV genome, thus contributing to KSHV terminase activity. To address this question, the cleavage of replicated KSHV DNA containing tandemly repeated genomes was detected by Southern blotting according to the method established by Glaunsinger et al (27, 28) with slight modifications (Fig. 2A).…”

Section: Resultsmentioning

confidence: 99%

“…The DNA samples (10 ug each) were digested overnight with EcoRI (Takara Bio) and SalI (TOYOBO) followed by electrophoresis on a 0.7% agarose 1xTBE gel. As for Southern blotting (27, 28), the gel was denatured in alkaline denaturing buffer (0.5 N NaOH, 1.5 M NaCl) for 45 min and transferred to Hybond-N+ (Cytiva, Tokyo, Japan) by capillary action overnight in the presence of alkaline transfer buffer (0.4 N NaOH, 1 M NaCl). The transferred DNA was cross-linked to the membrane with a CL-1000 Ultraviolet Crosslinker (UVP, Cambridge, UK).…”

Section: Discussionmentioning

confidence: 99%

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The contribution of Kaposi’s sarcoma-associated herpesvirus ORF7 and its zinc-finger motif to viral genome cleavage and capsid formation

Iwaisako

Watanabe

Sekine

et al. 2022

Preprint

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Kaposi’s sarcoma-associated herpesvirus (KSHV) is the causative agent of endothelial and B cell malignancies. During KSHV lytic infection, lytic-related proteins are synthesized, viral genomes are replicated as a tandemly repeated form, and subsequently, capsids are assembled. The herpesvirus terminase complex is proposed to package an appropriate genome unit into an immature capsid, by cleavage of terminal repeats (TRs) flanking tandemly linked viral genomes. Although the mechanism of capsid formation in α- and β-herpesviruses are well-studied, in KSHV, it remains largely unknown. It has been proposed that KSHV ORF7 is a terminase subunit, and ORF7 harbors a zinc-finger motif, which is conserved among other herpesviral terminases. However, the biological significance of ORF7 is unknown. We previously reported that KSHV ORF17 is essential for the cleavage of inner scaffold proteins in capsid maturation, and ORF17 knockout (KO) induced capsid formation arrest between the procapsid and B-capsid stages. However, it remains unknown if ORF7-mediated viral DNA cleavage occurs before or after ORF17-mediated scaffold collapse. We analyzed the role of ORF7 during capsid formation using ORF7-KO-, ORF7&17-double-KO (DKO)-, and ORF7-zinc-finger motif mutant-KSHVs. We found that ORF7 acted after ORF17 in the capsid formation process, and ORF7-KO-KSHV produced incomplete capsids harboring non-spherical internal structures, which resembled soccer balls. This soccer ball-like capsid was formed after ORF17-mediated B-capsid formation. Moreover, ORF7-KO- and zinc-finger motif KO-KSHV failed to appropriately cleave the TR on replicated genome and had a defect in virion production. Thus, our data revealed that ORF7 contributes to terminase-mediated viral genome cleavage and capsid formation.

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Section: Resultssupporting

confidence: 93%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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The contribution of Kaposi’s sarcoma-associated herpesvirus ORF7 and its zinc-finger motif to viral genome cleavage and capsid formation

Iwaisako

Watanabe

Sekine

et al. 2022

Preprint

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“…Orf50 encodes the replication and transcription activator protein, which initiates viral lytic gene expression 35 . Orf68 encodes a packaging protein that assists in moving the newly replicated viral genomes to the packaging motor, where they are loaded into capsids 36 . Orf73 encodes for the latency-associated nuclear antigen that is required for the establishment and maintenance of latent infection 37,38 .…”

Section: Resultsmentioning

confidence: 99%

Age-associated B cells are long-lasting effectors that restrain reactivation of latent γHV68

Mouat

Shanina

Horwitz

2021

Preprint

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Age-associated B cells (ABCs; CD19+CD11c+T-bet+) are increased during an array of viral infections, though their role during viral latency is unexplored. Here, we use murine gammaherpesvirus 68 (γHV68), a homolog of Epstein-Barr virus that latently infects B cells, to demonstrate that ABCs are necessary for the effective control of gamma-herpesvirus latency. We observe that ABCs expand in the spleen during acute infection and persist at least 150 days post-infection. During acute and latent infection ABCs secrete IFNγ and TNF. Using a strain of γHV68 that is cleared following acute infection, we show that ABCs persist in the absence of latent virus, though they secrete less IFNγ and TNF. With a fluorescent virus we demonstrate that ABCs are infected with γHV68 at similar rates to other mature B cells. We find that mice without ABCs display defects in anti-viral IgG2a/c antibodies and are less able to maintain γHV68 latency when challenged with heterologous infection. Together, these results indicate that ABCs are a persistent subset during latent viral infection that controls γHV68 reactivation from latency.

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“…Orf50 encodes the replication and transcription activator protein, which initiates viral lytic gene expression 36 . Orf68 encodes a packaging protein that assists in moving the newly replicated viral genomes to the packaging motor, where they are loaded into capsids 37 . Orf73 encodes for the latency-associated nuclear antigen that is required for the establishment and maintenance of latent infection 38,39 .…”

Section: Abcs Are Dispensable For the Control Of Acute Infection And ...mentioning

confidence: 99%

Age-associated B cells are long-lasting effectors that impede latent gHV68 reactivation

Mouat

Shanina

Horwitz

2022

Preprint

View full text Add to dashboard Cite

Age-associated B cells (ABCs; CD19+CD11c+T-bet+) are a unique population that are increased in an array of viral infections, though their role during latent infection is largely unexplored. Here, we use murine gammaherpesvirus 68 (γHV68) to demonstrate that ABCs remain elevated long-term during latent infection and express IFNγ and TNF. Using a recombinant γHV68 that is cleared following acute infection, we show that ABCs persist in the absence of latent virus, though their expression of IFNγ and TNF is decreased. With a fluorescent reporter gene-expressing γHV68 we demonstrate that ABCs are infected with γHV68 at similar rates to other previously activated B cells. We find that mice without ABCs display defects in anti-viral IgG2a/c antibodies and are more susceptible to reactivation of γHV68 following virus challenges that typically do not break latency. Together, these results indicate that ABCs are a persistent effector subset during latent viral infection that impedes γHV68 reactivation.

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A pentameric protein ring with novel architecture is required for herpesviral packaging

Cited by 11 publications

References 68 publications

The contribution of Kaposi’s sarcoma-associated herpesvirus ORF7 and its zinc-finger motif to viral genome cleavage and capsid formation

The contribution of Kaposi’s sarcoma-associated herpesvirus ORF7 and its zinc-finger motif to viral genome cleavage and capsid formation

Age-associated B cells are long-lasting effectors that restrain reactivation of latent γHV68

Age-associated B cells are long-lasting effectors that impede latent gHV68 reactivation

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