Understanding a viral polymerase complex: a multienzyme 'molecular motor' that drives virus replication

Viruses can only successfully infect animals by hijacking normal cellular functions and using these to make many more copies of the virus. However, cells do not contain all of the machinery necessary to copy the virus genes, and so the virus has to carry some of this with it when it infects the animal. As this part of the virus has no cellular equivalent, but is necessary for the virus to reproduce, it is a very good target for medicine that interferes with virus infection. Bluetongue virus causes a devastating disease in sheep where up to 70% if infected animals die. For this virus we know all of the genes in the virus, understand the overall structure of the pathogen, and have demonstrated the function of some of the parts of the machinery that copies the virus at a test tube level. What we do not know is precisely how these individual pieces of the machinery work together and that is the focus of this research study. The approach taken will involve a study of the structure of the machinery at a very small scale, as well as experiments that test how well each part of the overall machine is working. Outputs from this research will provide opportunities for the design of new medicine that interferes with the virus machinery and prevents it from copying itself.

The structure of the virus core of dsRNA viruses within the family Reoviridae has revealed a functional molecular machine dedicated to the efficient transcription of RNA upon cell entry, so initiating the infectious cycle. The present level of understanding has required a merging of structural and biophysical studies to map the protein-protein interactions within the core with biochemistry of individual reactions within the protein microenvironment and represents a novel and holistic approach to uncovering the action of the virion core. It is still the case however that the described activity of each individual core protein is incomplete and that a complete understanding of how the core proteins act in concert is lacking. This proposal will address these issues with a comprehensive analysis of each of the viral enzymes associated with transcription from the virus core. The proposed work will reveal how these proteins come together to provide the complete viral transcription function and, in an important and unique objective, we will reconstitute a functional transcriptase its component parts in vitro / a first in the field. The output from the work will contribute to the growing area of the structural basis of transcription and pave the way for the rational design of viral transcriptase inhibitors with the potential to block orbivirus replication and provide lead compounds of relevance to many related viruses of pathogenic significance to man and animals.