Understanding the reprogramming of host mRNA translation during calicivirus infection

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Caliciviruses, a family of small RNA viruses, are important pathogens of man and animals yet they remain poorly characterised. They use a novel mechanism of viral protein synthesis that involves the recruitment of cellular initiation factors to a virus-encoded protein attached to the 5' end of the viral genome, namely VPg. We have recently discovered that calicivirus infection regulates the activity of eIF4E via phosphorylation. We also found that host mRNA translation undergoes a specific reprogramming as a result of p-eIF4E associating with polysomes. Our preliminary data would indicate that this biased recruitment is focused on mRNAs involved in the cellular response to viral infection and may facilitate virus replication.

Our overall aim is to dissect how norovirus infection alters the gene expression profile of infected cells contributing to viral pathogenesis, and how the regulation of eIF4E activity participates in this response to infection.
First, given our observations on the translation of specific genes, we will characterize the global regulation of the host mRNA translation by using polysome profiling and RNA sequencing. We will confirm our analysis by using proteomics to further characterize the impact of this translational control on the antiviral response during calicivirus infection. Then, we will unravel the specific contribution of eIF4E phosphorylation in this process. We will define the function of eIF4E phosphorylation by identifying mRNA specifically translated by p-eIF4E using polysomes profiling and animal models. Finally, we will establish the contribution of the kinase responsible for eIF4E phosphorylation, MNK, and characterize the interactions between MNK-eIF4G-eIF4E and VPg.
This will provide an unprecedented understanding of the regulation of host gene expression by caliciviruses, from mRNA production to protein synthesis, and shed light on how these viruses may modulate the antiviral response.

Impact Summary-

The preliminary data presented in this application, and the experiments planned to build on our findings, will lead to a step-change in our understanding of the control of host translational and the host antiviral response in calicivirus infection. Our findings are also likely to shed light on fundamental processes underpinning the control of gene expression and the host response to infection. This research will have a direct scientific impact in the fields of virology, translational control and host/virus interactions. It also has strong potential for economic/societal impact. As caliciviruses are important human and animal pathogens, our work may identify new targets for treatment of these economically important infections and therefore has the potential to impact on UK health, society and economy.

Industrial and Economic Impact
The regulation of translation initiation and the signalling pathways associated with translation are well established targets for therapeutic intervention against diseases that arise as a result of translational deregulation. The MAPK pathway has already been the focus for the development of drugs against cancer (e.g: Selumetinib for small cell lung cancer or Trametinib for metastatic melanoma). In addition, numerous cancers have been identified in which eIF4E is over-expressed. The dissection of the points in which translation is regulated during viral infection will therefore provide the pharmaceutical industry with new leads in i) the development of specific antiviral therapies and ii) the development of drugs for pathologies linked to deregulation of translation, such as cancer. Importantly for the pharmaceutical industry (Pfizer, GSK, AstraZeneca), some of the drugs that are currently being developed to control cell proliferation (e.g: Selumetinib, Trametinib, PD0325901, LY2228820) could also act as potent antivirals through the control they exert on eIF4E phosphorylation. This could expand the portfolio of application for existing drugs, greatly reducing the development time, benefiting the UK and worldwide population.

Public sector and Societal Impact
Noroviruses, often referred to as 'winter vomiting disease', are a significant public health problem. Outbreaks in hospitals alone often compromise patient care at a time of year when NHS Trusts are already under pressure, namely the winter months. In addition, norovirus outbreaks in schools, cruise ships, care homes and restaurants have a significant socioeconomic impact. The 2012-2013 winter season saw in excess of 1 million cases in the UK, which is typical of the annual norovirus season. The findings from our work will be publicised widely via the respective university press offices but also via our outreach activities raising awareness of noroviruses in the general public.

Training of skilled researchers
Two research assistants, one at post-doctoral level and one graduate RA, will be recruited as part of this project. Both will receive extensive training in state of the art molecular methods for the study of gene expression (RNA sequencing, polysome profiling for PDRA1 and proteomics for RA2). They will also be trained extensively in molecular biology and virology techniques. They will gain experience in the systems biology-type approaches that integrate large data sets. We aim to provide a holistic set of skills that will equip the research assistants for challenges relevant to a wide range of careers both in academic research and industrial, increasing their long-term career prospects. In addition, our laboratories regularly host both undergraduate and post-graduate students, who will also benefit from exposure to the BBSRC funded research.