Virus-mediated nucleolar polyadenylation: a novel mechanism of RNA processing compartmentalisation to escape global mRNA degradation

RNA has to undergo a series of processing events prior to its nuclear export and translation into protein. In mammalian cells, the half-life of RNA can vary between a few minutes to many hours. This mRNA turnover can be regulated by its 3' processing which can either lead to the stabilisation of the RNA or enhance its degradation. This process is particularly important as the regulation of RNA turnover provides an effective way to alter the amount of RNA and thus the amount of protein produced. Moreover, many human diseases are due to aberrant RNA processing and as such understanding the fundemental mechanisms by which viruses can overcome these RNA degradation pathways may provide clues for therapeutic interventions in the future.

It is not surprising that viruses have evolved ways to control RNA 3' processing events and thus RNA turnover. For example Kaposi's sarcoma-associated herpesvirus (an oncogenic herpesvirus) lytic infection leads to a dramatic and rapid shutoff of host cell gene expression; where the majority of cellular mRNAs are degraded due to the expression of the virally-encoded SOX protein. SOX affects the 3' processing of cellular RNAs by causing the RNAs to have extra long poly(A) tails, which in turn leads to their instability and resulting degradation. However, an intriguing question is how do these viral RNAs evade this process. The viral mRNAs are transcribed and processed similar to cellular mRNAs and in essence should also be degraded at the same time as the cellular mRNAs.
We have exciting preliminary data to suggest that a virally-encoded protein, ORF57, redistributes cellular 3' processing factors into the nucleolus, a distinct sub-structure in the nucleus, which provides an alternative environment for the correct processing of viral RNAs, therefore bypassing the KSHV SOX-mediated degradation mechanism for cellular mRNAs. As such, this highlights a novel mechanism employed by a virus to evade a global RNA degradation process.

We now aim to further investigate these observations and identify the components of the cellular RNA processing complex that are redistributed into the nucleolus by the ORF57 protein. Moreover, we will investigate how the multi-protein complex is moved to the nucleolus and also assess the role of the cellular proteins in viral RNA processing and whether their function is essential for virus replication. If so, this may provide new strategies for the therapeutic intervention of herpesvirus infections.

Post-transcriptional events which regulate mRNA biogenesis are central to the regulation of gene expression. Not surprisingly viruses have evolved ways to manipulate these pathways. KSHV lytic infection leads to an extensive destruction of the host cell transcriptome, due to viral-mediated aberrant polyadenylation of cellular transcripts. These transcripts have extended poly(A) tails which leads to their enhanced hyperpolyadenylation-linked mRNA turnover.

One intriguing question let to be addressed is how the KSHV mRNAs evade this shutoff mechanism. This is an important issue, as delineation of this mechanism may provide clues as to how cellular quality control checkpoints can be bypassed during viral infection and other human disease.

We have exciting preliminary observations which lead to an intriguing hypothesis of how KSHV mRNAs bypass global cellular transcriptome degradation. Specifically, we present data suggesting that the KSHV ORF57 protein redistributes cellular 3' processing factors into the nucleolus, which provides an alternative environment for the correct processing of viral RNAs, therefore bypassing the KSHV SOX-mediated host cell shutoff mechanism. As such, this highlights a novel mechanism employed by a virus to evade a global transcriptome degradation process.

We now aim to further investigate this process and identify the components of the nucleolar-localised viral mRNA 3' processing complex. Moreover, we will determine how the 3' processing complex is localised to the nucleolus and also assess the role of the cellular proteins in viral 3' processing and virus replication.

This project will delineate how a virus can subvert cellular RNA biogenesis pathways. In addition, it will provide evidence for a previously undescribed role of the nucleolus in the 3' processing. As such this will provide fundemental new insights into cell biology processes.

Who will benefit from this research and how ?

The proposal builds upon previous novel work which has focussed on applying quantitative proteomic-based strategies to understanding the interactions between viruses and the host cell. The aim of this current proposal is to test the hypothesis that a herpesvirus protein provides an alternative environment for the 3' processing of viral mRNAs to avoid a global cellular mRNA degradation process.

Whilst this study is fundamental in nature, the impact of the research will be wide reaching. Aberrant RNA processing and nucleolar function are implicated in a number of human diseases, therefore any clues as to how cellular RNA processing quality control checkpoints are bypassed generated from this project will be of interest to the pharmaceutical industry. Moreover, a key element of this project is the identification of essential virus-host cell interactions which will provide avenues for novel antiviral strategies. For example, the nucleophosmin inhibitor, NSC348884 which is currently being developed as a potential cancer therapy, may also have potential as a novel KSHV antiviral agent. Moreover, many other viruses target the nucleolus during their replication, for example HIV mRNAs are also trafficked through the nucleulus, therefore this project may provide clues to why other viruses interact with the nucleolus. Therefore, these discoveries may foster new collaborations with the pharmaceutical and other commercial industries to exploit these findings for new therapeutic strategies.

In the longer term, exploitation of these findings by the commercial sector may lead to new treatments for a wide range of diseases and virus infections, and this will provide benefits to the quality of life of the general public. Moreover, exploitation of the research findings by the commercial sector is also likely to have a direct impact on the prosperity of the general public of the UK, through increased investment and employment opportunities that will arise from new therapeutic drugs.