A novel innate signalling pathway and its viral antagonism

Human cytomegalovirus (HCMV) infects most people worldwide, and establishes a persistent, lifelong infection. Although most people do not suffer from serious illness, immunocompromised individuals such as transplant recipients can suffer from devastating disease including organ rejection, multi-organ failure and even death. Furthermore, unborn babies are at a particular risk from HCMV, both in mothers who become infected during pregnancy and in mothers who already have the virus. Overall ~1/100 pregnancies are affected, and HCMV is the leading infectious cause of deafness and intellectual disability in children, and one of the most significant viral causes of birth defects.

One of the first lines of defence against HCMV and other viruses are specific 'sensing proteins', which can detect infection and activate 'signalling proteins' to trigger critical immune responses. To be able to replicate, some viruses such as HCMV have developed mechanisms to subvert these proteins.

Our aim is to understand how our cells detect HCMV and other pathogens by characterising previously undiscovered signalling proteins. I have developed new technologies to identify signalling proteins based on how they move within cells to warn of infection, and whether they are destroyed by viruses.

In initial experiments, we have discovered two novel viral signalling proteins. Crucially, these proteins are activated not only by infection with HCMV but also infection with an evolutionally distinct virus - Sendai. This is suggestive of a general response to viral infection, and potentially a general response to infection by diverse types of pathogen. Intriguingly, whereas one protein triggers the production of antiviral molecules that can inhibit HCMV, and is rapidly destroyed by the virus upon infection, the other acts to promote infection - and the virus even uses one of its own proteins to assist this process. We now wish to understand how these proteins work in much more detail, specifically how they are activated by sensing proteins, exactly which antiviral or proviral molecules they cause to be produced, and how they regulate and are subverted by HCMV and other viruses. To achieve these aims, we will study skin cells and immune sensing cells, which both play vital yet distinct roles in HCMV infection. Both can be infected with HCMV and Sendai virus in our laboratory.

By understanding how HCMV and other viruses subvert signalling proteins, we will ultimately be able to develop vital new treatments to prevent diseases caused by viral infection. Crucially, this knowledge will also dramatically improve our understanding of immunity to viruses in general.

Appropriate cell-intrinsic recognition of intracellular pathogens is essential for the generation of effective innate and adaptive antiviral immunity. Viral sensors and their signalling components thus provide a crucial first line of host defence. Using new proteomic approaches, we have identified two novel signalling components (proteins 1 and 2) pivotal in understanding innate immune recognition of diverse DNA and RNA viruses, in addition to how HCMV persists in vivo. Importantly, activation of protein 1 represents an entirely novel trigger for interferon-independent induction of antiviral gene products, whereas protein 2 promotes expression of proviral factors. We will now determine:

1. How do proteins 1 and 2 respond to viral infection?
Using biochemical approaches, RNAseq and proteomics in human fibroblasts and primary dendritic cells infected with HCMV or Sendai virus, we will:
a. identify upstream sensors and kinases;
b. characterise cellular transcriptional effects and transcription complex components;
c. characterise cell-type specific similarities and differences.

Using cellular gene knockout and viral mutagenesis, we will then:
d. determine which protein 1-dependent cellular proteins restrict HCMV, and characterise the proviral effect of protein 2 on viral and cellular transcription over time.

2. How do HCMV and other viruses subvert proteins 1 and 2?
Using interaction studies, viral mutagenesis and biochemical/structural approaches, we will:
a. characterise the viral antagonist targeting protein 1
b. determine the mechanism of HCMV-mediated subversion of protein 2
c. collaborate to define protein 1/2 subversion by EBV, HSV and SARS-CoV-2.

Overall, these studies will provide detailed characterisation of crucial host antiviral defence pathways, revealing fundamental insights into innate immunity. Furthermore, the small-molecule therapeutic opportunities offered by identification of these key viral-host interactions may be unparalleled.