Imaging the early events of virus infection in plants

This project will investigate the location and redistribution of fluorescent viral RNA immediately following its microinjection into living plant cells. The project will utilise real-time imaging approaches and newly developed FlAsH and ReAsH tags to track viral gene products immediately following their translation. These tagged proteins will be imaged against a transgenic background in which a range of cellular components (e.g. ER, microtubules, plasmodesmata) have been stably highlighted with green fluorescent protein (GFP). The aim is to provide unequivocal evidence for the pathway of viral RNA trafficking in plants during the first stages of infection, and to identify the viral and host factors that interact with the viral genome during transit. The project will be carried out in collaboration with Dr Ilan Davis, Wellcome Centre for Cell Biology, and will utilise real-time deconvolution imaging approaches developed in the Central Optical Imaging Laboratory (COIL), Edinburgh University.

The aim of this project is to image the earliest events of virus infection using real-time imaging approaches, and to identify the host factors that interact with viral RNA during its intra-and intercellular transport. Tobacco mosaic virus (TMV) viral RNA will be labelled with Alexa-UTP and microinjected into trichome cells to image the earliest events in viral RNA anchoring and trafficking. To image newly translated pools of viral gene products, viral vectors will be constructed that express the movement protein, coat protein and replicase proteins tagged with tetracysteine motifs that will be subsequently localized with FlAsH and ReAsH reagents. The project will also examine the phenomenon by which 'rescue' of viral RNA occurs in the presence of heterologous movement proteins using a combination of microinjection and deletion mutagenesis. The project will utilise real-time deconvolution imaging approaches for RNA tracking developed in the Central Optical Imaging Laboratory (COIL), Edinburgh University.