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Herpes simplex virus-1 bottleneck during genome replication

Final Report Summary - HSV-1 BOTTLENECK (Herpes simplex virus-1 bottleneck during genome replication)

Herepsviruses are a large group of DNA viruses that cause significant morbidity and mortality in human and animals. As all herpesviruses have a similar replication process in the nucleus of the infected host cell, Herpes simplex virus-1 (HSV-1), a common human pathogen, is considered a prototype for studying the replication of the herpesviridae family. The replication of the viral genome is regulated by both viral and host proteins. Interestingly, some host proteins contribute to the viral replication whereas others have inhibitory effect on the replication process. We set up to study a recently described bottleneck phenomenon during the HSV replication, the finding that only a limited number of incoming viral genomes can initiate expression and replication. We developed two systems that allowed us to explore this phenomenon. First, we used a three-colors infection system that allows to estimate the average number of viral genomes initiating expression per cell. This system allowed perturbation the infection process and testing the effect of the perturbation on the limited number of genomes observed previously. Second, we developed a system that allow direct quantitation of the number of viral genomes replicated in the single cell level.
One of the goals set in the research proposal was the establishment of a functional herpesviruses research group. Indeed, I was able to establish a research group ( in Tel Aviv University that includes graduate and undergraduate students. Work in my lab yielded four scientific publications in scientific journals, including high impact journal as PNAS. The findings of the funded project were recently published in Frontiers in microbiology (Shapira, L. et al. 2016) and in PLoS Pathogens (Cohen E. and Kobiler O. 2016).

To study the mechanisms regulating the number of herpes viral genomes initiating replication in an infected cell, we first tested the effect of histone deacetylase inhibitors (HDACi’s) on infection. We found that HDACi treatment decreases the number of viral genomes establishing infection per cell independent of the cell type. We further identified that HDACi treatment increases the levels of antiviral ND10 proteins. In support of this findings we identified that In the absence of ICP0 (a viral protein known to reduce ND10 proteins levels), a significant decrease in the number of viral genomes that initiate expression compared to wild type infection was detected in all MOIs tested.
Using a novel single cell assay we found that viral gene expression correlates with the number of viral genomes replicating per cell. On the other hand, We found that in cells with high gene expression from host house-keeping promoters the gene expression from the virus is usually low and vice versa.
The results obtained in our lab provide a novel level of regulation on the process of herpesvirus infection. Our finding provide evidence that the bottleneck limiting the number of genomes initiating a successful infection, is regulated by host genes. We speculate that specific intrinsic antiviral proteins are key in regulating this bottleneck.

Potential socio-economic impact:
Herpesviruses are still a major source for morbidity and mortality in developed countries, especially for newborns and immunocompromised populations. Unfortunately, in these populations, we detect resistance to current antiviral treatments (up to 20%), thus there is a need for new therapeutic options. Understanding the fundamental mechanisms involved in the replication of these viruses, can provides new mechanisms that can be targeted by drugs. Our findings suggest that even small modifications in the levels of host protein can have great effect on viral infection. Targeting host factors instead of viral proteins may reduce the development of drug resistance problems.