We study viruses and explore the creative strategies they use to maneuver their host cells. We are interested in deciphering the roles different viral elements play during infection, as well as how viruses interact with and commandeer cellular pathways to control gene expression. We study these complex interactions using mainly cytomegalovirus (CMV), a herpesvirus that infects most of the world's population, but also additional viruses, such as SARS-CoV2. Leveraging high-throughput technologies, we approach these complex questions with an unbiased perspective, and uncover new aspects of virus-host interactions, as well as reveal new cell biology principles.
Finkel Y., Nachshon A., Aharon E., Arazi T., Simonovsky E., Dobešová M., Saud Z., Gluck A., Fisher T., Stanton R. J., Schwartz M. & Stern-Ginossar N.
(2024)
Nature.
630,
8017,
p. 712-719
Genetic screens have transformed our ability to interrogate cellular factor requirements for viral infections1,2, but most current approaches are limited in their sensitivity, biased towards early stages of infection and provide only simplistic phenotypic information that is often based on survival of infected cells24. Here, by engineering human cytomegalovirus to express single guide RNA libraries directly from the viral genome, we developed virus-encoded CRISPR-based direct readout screening (VECOS), a sensitive, versatile, viral-centric approach that enables profiling of different stages of viral infection in a pooled format. Using this approach, we identified hundreds of host dependency and restriction factors and quantified their direct effects on viral genome replication, viral particle secretion and infectiousness of secreted particles, providing a multi-dimensional perspective on virushost interactions. These high-resolution measurements reveal that perturbations altering late stages in the life cycle of human cytomegalovirus (HCMV) mostly regulate viral particle quality rather than quantity, establishing correct virion assembly as a critical stage that is heavily reliant on virushost interactions. Overall, VECOS facilitates systematic high-resolution dissection of the role of human proteins during the infection cycle, providing a roadmap for in-depth study of hostherpesvirus interactions.
Schwartz M., Shnayder M., Nachshon A., Arazi T., Kitsberg Y., Levi Samia R., Lavi M., Kuint R., Tsabari R. & Stern-Ginossar N.
(2023)
Nature Microbiology.
8,
3,
p. 455-468
Human cytomegalovirus (HCMV) can result in either productive or non-productive infection, with the latter potentially leading to viral latency. The molecular factors dictating these outcomes are poorly understood. Here we used single-cell transcriptomics to analyse HCMV infection progression in monocytes, which are latently infected, and macrophages, considered to be permissive for productive infection. We show that early viral gene expression levels, specifically of those encoding immediate early proteins IE1 and IE2, are a major factor dictating productive infection. We also revealed that intrinsic, not induced, host cell interferon-stimulated gene expression level is a main determinant of infection outcome. Intrinsic interferon-stimulated gene expression is downregulated with monocyte to macrophage differentiation, partially explaining increased macrophage susceptibility to productive HCMV infection. Furthermore, non-productive macrophages could reactivate, making them potential latent virus reservoirs. Overall, we decipher molecular features underlying HCMV infection outcomes and propose macrophages as a potential HCMV reservoir.
Fisher T., Gluck A., Narayanan K., Kuroda M., Nachshon A., Hsu J. C., Halfmann P. J., Yahalom-Ronen Y., Tamir H., Finkel Y., Schwartz M., Weiss S., Tseng C. K., Israely T., Paran N., Kawaoka Y., Makino S. & Stern-Ginossar N.
(2022)
Cell reports (Cambridge).
39,
11,
110954.
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) leads to shutoff of protein synthesis, and nsp1, a central shutoff factor in coronaviruses, inhibits cellular mRNA translation. However, the diverse molecular mechanisms employed by nsp1 as well as its functional importance are unresolved. By overexpressing various nsp1 mutants and generating a SARS-CoV-2 mutant, we show that nsp1, through inhibition of translation and induction of mRNA degradation, targets translated cellular mRNA and is the main driver of host shutoff during infection. The propagation of nsp1 mutant virus is inhibited exclusively in cells with intact interferon (IFN) pathway as well as in vivo, in hamsters, and this attenuation is associated with stronger induction of type I IFN response. Therefore, although nsp1s shutoff activity is broad, it plays an essential role, specifically in counteracting the IFN response. Overall, our results reveal the multifaceted approach nsp1 uses to shut off cellular protein synthesis and uncover nsp1s explicit role in blocking the IFN response.
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Nsp1 inhibits translation, induces mRNA degradation, and blocks nuclear mRNA exportNsp1 effects on translation, degradation, and export represent distinct functionsNsp1 is SARS-CoV-2 main shutoff factor, and it broadly targets translated cellular mRNAsNsp1 functional importance lies explicitly in the attenuation of the interferon response
Here, Fisher et al. show the importance of nsp1, a central shutoff factor in coronaviruses, during SARS-CoV-2 infection. nsp1 has broad activity, and it targets all translated cellular mRNA, but its functional importance in SARS-CoV-2 infection lies explicitly in blocking the IFN response.
