miR-927 has pro-viral effects during acute and persistent infection with dengue virus type 2 in C6/36 mosquito cells

Dengue virus (DENV) is an important flavivirus that is transmitted to humans by Aedes mosquitoes, where it can establish a persistent infection underlying vertical and horizontal transmission. However, the exact mechanism of persistent DENV infection is not well understood. Recently miR-927 was found to be upregulated in C6/36-HT cells at 57 weeks of persistent infection (C6-L57), suggesting its participation during this type of infection. The aim of this study was to determine the role of miR-927 during infection with DENV type 2. The results indicate an overexpression of miR-927 in C6-L57 cells and acutely infected cells according to the time of infection and the m.o.i. used. The downregulation of miR-927 in C6-L57 cells results in a reduction of both viral titre and viral genome copy number. The overexpression of miR-927 in C6-L40 and C6/36 cells infected at an m.o.i. of 0.1 causes an increase in both viral titre and viral genome copy number, suggesting a pro-viral activity of miR-927. In silico prediction analysis reveals target mRNAs for miR-927 are implicated in post-translational modifications (SUMO), translation factors (eIF-2B), the innate immune system (NKIRAS), exocytosis (EXOC-2), endocytosis (APM1) and the cytoskeleton (FLN). The expression levels of FLN were the most affected by both miR-927 overexpression and inhibition, and FLN was determined to be a direct target of miR-927 by a dual-luciferase gene reporter assay. FLN has been associated with the regulation of the Toll pathway and either overexpression or downregulation of miR-927 resulted in expression changes of antimicrobial peptides (Cecropins A and G, and Defensin D) involved in the Toll pathway response.

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The Latent Herpes Simplex Virus Type 1 Genome Copy Number in Individual Neurons Is Virus Strain Specific and Correlates with Reactivation

Journal of Virology ◽

10.1128/jvi.72.7.5343-5350.1998 ◽

1998 ◽

Vol 72 (7) ◽

pp. 5343-5350 ◽

Cited By ~ 118

Author(s):

N. M. Sawtell ◽

D. K. Poon ◽

C. S. Tansky ◽

R. L. Thompson

Keyword(s):

Herpes Simplex ◽

Viral Genome ◽

Copy Number ◽

Genetic Factors ◽

Genome Copy ◽

Latently Infected ◽

Genome Copy Number ◽

Simplex Virus

ABSTRACT The viral genetic elements that determine the in vivo reactivation efficiencies of fully replication competent wild-type herpes simplex virus (HSV) strains have not been identified. Among the common laboratory strains, KOS reactivates in vivo at a lower efficiency than either strain 17syn+ or strain McKrae. An important first step in understanding the molecular basis for this observation is to distinguish between viral genetic factors that regulate the establishment of latency from those that directly regulate reactivation. Reported here are experiments performed to determine whether the reduced reactivation of KOS was associated with a reduced ability to establish or maintain latent infections. For comparative purposes, latent infections were quantified by (i) quantitative PCR on DNA extracted from whole ganglia, (ii) the number of latency-associated transcript (LAT) promoter-positive neurons, using KOS and 17syn+ LAT promoter–β-galactosidase reporter mutants, and (iii) contextual analysis of DNA. Mice latently infected with 17syn+-based strains contained more HSV type 1 (HSV-1) DNA in their ganglia than those infected with KOS strains, but this difference was not statistically significant. The number of latently infected neurons also did not differ significantly between ganglia latently infected with either the low- or high-reactivator strains. In addition to the number of latent sites, the number of viral genome copies within the individual latently infected neurons has recently been demonstrated to be variable. Interestingly, neurons latently infected with KOS contained significantly fewer viral genome copies than those infected with either 17syn+ or McKrae. Thus, the HSV-1 genome copy number profile is viral strain specific and positively correlates with the ability to reactivate in vivo. This is the first demonstration that the number of HSV genome copies within individual latently infected neurons is regulated by viral genetic factors. These findings suggest that the latent genome copy number may be an important parameter for subsequent induced reactivation in vivo.

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Oral Inoculation of Chickens with a Candidate Fowl Adenovirus 9 Vector

Clinical and Vaccine Immunology ◽

10.1128/cvi.00187-13 ◽

2013 ◽

Vol 20 (8) ◽

pp. 1189-1196 ◽

Cited By ~ 16

Author(s):

Li Deng ◽

Shayan Sharif ◽

Éva Nagy

Keyword(s):

Gene Expression ◽

Antibody Response ◽

Virus Replication ◽

Viral Genome ◽

Copy Number ◽

Cytokine Gene Expression ◽

Genome Copy ◽

Virus Shedding ◽

Genome Copy Number

ABSTRACTFowl adenoviruses (FAdVs) are a potential alternative to human adenovirus-based vaccine vectors. Our previous studies demonstrated that a 2.4-kb region at the left end of the FAdV-9 genome is nonessential for virus replication and is suitable for the insertion or replacement of transgenes. Ourin vivostudy showed that the virus FAdV-9Δ4, lacking six open reading frames (ORFs) at the left end of its genome, replicates less efficiently than wild-type FAdV-9 (wtFAdV-9) in chickens that were infected intramuscularly. However, the fecal-oral route is the natural route of FAdV infection, and the oral administration of a vaccine confers some advantages compared to administration through other routes, especially when developing an adenovirus as a vaccine vector. Therefore, we sought to investigate the effects of FAdV-9 in orally inoculated chickens. In the present study, we orally inoculated specific-pathogen-free (SPF) chickens with FAdV-9 and FAdV-9Δ4 and assessed virus shedding, antibody response, and viral genome copy number and cytokine gene expression in tissues. Our data showed that FAdV-9Δ4 replicated less efficiently than did wtFAdV-9, as evidenced by reduced virus shedding in feces, lower viral genome copy number in tissues, and lower antibody response, which are consistent with the results of the intramuscular route of immunization. Furthermore, we found that both wtFAdV-9 and FAdV-9Δ4 upregulated the mRNA expression of alpha interferon (IFN-α), IFN-γ, and interleukin-12 (IL-12). In addition, there was a trend toward downregulation of IL-10 gene expression caused by both viruses. These findings indicate that one or more of the six deleted ORFs contribute to modulating the host response against virus infection as well as virus replicationin vivo.

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Replication of Herpes Simplex Virus Type 1 within Trigeminal Ganglia Is Required for High Frequency but Not High Viral Genome Copy Number Latency

Journal of Virology ◽

10.1128/jvi.74.2.965-974.2000 ◽

2000 ◽

Vol 74 (2) ◽

pp. 965-974 ◽

Cited By ~ 56

Author(s):

Richard L. Thompson ◽

N. M. Sawtell

Keyword(s):

Herpes Simplex Virus ◽

Herpes Simplex ◽

Herpes Simplex Virus Type ◽

Viral Genome ◽

Copy Number ◽

Trigeminal Ganglia ◽

Genome Copy ◽

Genome Copy Number ◽

Simplex Virus

ABSTRACT The replication properties of a thymidine kinase-negative (TK−) mutant of herpes simplex virus type 1 (HSV-1) were exploited to examine the relative contributions of replication at the body surface and within trigeminal ganglia (TG) on the establishment of latent infections. The replication of a TK− mutant, 17/tBTK−, was reduced by ∼12-fold on the mouse cornea compared to the rescued isolate 17/tBRTK+, and no replication of 17/tBTK− in the TG of these mice was detected. About 1.8% of the TG neurons of mice infected with 17/tBTK− harbored the latent viral genome compared to 23% of those infected with 17/tBRTK+. In addition, the latent sites established by the TK− mutant contained fewer copies of the HSV-1 genome (average, 2.3/neuron versus 28/neuron). On the snout, sustained robust replication of 17tBTK− in the absence of significant replication within the TG resulted in a modest increase in the number of latent sites. Importantly, these latently infected neurons displayed a wild-type latent-genome copy number profile, with some neurons containing hundreds of copies of the TK− mutant genome. As expected, the replication of the TK− mutant appeared to be blocked prior to DNA replication in most ganglionic neurons in that (i) virus replication was severely restricted in ganglia, (ii) the number of neurons expressing HSV proteins was reduced 30-fold compared to the rescued isolate, (iii) cell-to-cell spread of virus was not detected within ganglia, and (iv) the proportion of infected neurons expressing late proteins was reduced by 89% compared to the rescued strain. These results demonstrate that the viral TK gene is required for the efficient establishment of latency. This requirement appears to be primarily for efficient replication within the ganglion, which leads to a sixfold increase in the number of latent sites established. Further, latent sites with high genome copy number can be established in the absence of significant virus genome replication in neurons. This suggests that neurons can be infected by many HSV virions and still enter the latent state.

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Replication of Herpes Simplex Virus Type 1 within Trigeminal Ganglia Is Required for High Frequency but Not High Viral Genome Copy Number Latency

Journal of Virology ◽

10.1128/jvi.74.24.12003a-12003a.2000 ◽

2000 ◽

Vol 74 (24) ◽

pp. 12003-12003

Author(s):

Richard L. Thompson ◽

N. M. Sawtell

Keyword(s):

Herpes Simplex Virus ◽

Herpes Simplex ◽

High Frequency ◽

Viral Genome ◽

Copy Number ◽

Trigeminal Ganglia ◽

Genome Copy ◽

Genome Copy Number ◽

Simplex Virus

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Nuclear-cytoplasmic balance: whole genome duplications induce elevated organellar genome copy number

10.1101/2021.06.08.447629 ◽

2021 ◽

Author(s):

Matheus Fernandes Gyorfy ◽

Emma R Miller ◽

Justin L Conover ◽

Corrinne E Grover ◽

Jonathan F Wendel ◽

...

Keyword(s):

Copy Number ◽

Nuclear Genome ◽

Plant Genome ◽

Whole Genome ◽

Genome Copy ◽

Relative Copy Number ◽

Organellar Genome ◽

Plastid Genomes ◽

Copy Numbers ◽

Genome Copy Number

The plant genome is partitioned across three distinct subcellular compartments: the nucleus, mitochondria, and plastids. Successful coordination of gene expression among these organellar genomes and the nuclear genome is critical for plant function and fitness. Whole genome duplication events (WGDs) in the nucleus have played a major role in the diversification of land plants and are expected to perturb the relative copy number (stoichiometry) of nuclear, mitochondrial, and plastid genomes. Thus, elucidating the mechanisms whereby plant cells respond to the cytonuclear stoichiometric imbalance that follow WGDs represents an important yet underexplored question in understanding the evolutionary consequences of genome doubling. We used droplet digital PCR (ddPCR) to investigate the relationship between nuclear and organellar genome copy numbers in allopolyploids and their diploid progenitors in both wheat and Arabidopsis. Polyploids exhibit elevated organellar genome copy numbers per cell, largely preserving the cytonuclear stoichiometry observed in diploids despite the change in nuclear genome copy number. To investigate the timescale over which cytonuclear stoichiometry may respond to WGD, we also estimated organellar genome copy number in Arabidopsis synthetic autopolyploids and in a haploid-induced diploid line. We observed corresponding changes in organellar genome copy number in these laboratory-generated lines, indicating that at least some of the cellular response to cytonuclear stoichiometric imbalance is immediate following WGD. We conclude that increases in organellar genome copy numbers represent a common response to polyploidization, suggesting that maintenance of cytonuclear stoichiometry is an important component in establishing polyploid lineages.

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