Major current lines of research involve:
Host factors involved in cell-to-cell movement of plant viruses
C Carvalho, JE Wellink (MOB), JWM v Lent RW Goldbach
Cell-to-cell movement of plant viruses through plasmodesmata involves specific viral factors (movement proteins, MP) and is inevitably assisted by cellular mechanisms in MP targeting, plasma membrane (PM) anchoring and transport of the MP through the plasmodesmal pore. An important group of plant pathogenic viruses, e.g. Cowpea mosaic virus (CPMV) and Tomato spotted wilt virus (TSWV), employs a so-called tubule-guided movement mechanism, by which whole virions are guided through tubules into the neighbouring cells through modified plasmodesmata. In the past decade we have identified the virally encoded factors involved in this process, the next intriguing issue, i.e. the evident role of host factors, is to be resolved.
This project aims at the identification of host factors that participate in the virus movement process. Using purified HIS-tagged MPs as probes in a variety of assays (Far Western assays, column filtration of plant protein extracts, screening of an Arabidopsis thaliana expression library and as probes for in situ localisation at LM and EM level), host proteins with affinity for viral MPs will be identified and characterised. The outcome of this research will not only contribute to the understanding of virus movement, but will also give further insight in the host's cellular processes that are recruited by plant viruses.
Mechanism of (Phloem-mediated) long distance transport of cowpea mosaic virus
M Santos Silva, JE Wellink (MOB), JWM v Lent RW Goldbach
Successful infection of the host plant completely depends on the ability of (progeny) virus to move systemically from the initially infected cell into neighbouring cells and subsequently to other parts of the plant. Two clearly distinct events determine spread of virus through the plant: 1) movement of the virus from cell-to-cell (through plasmodesmata) to the vascular tissue and 2) transport of the virus over long distances through the vascular system to other plant parts. For local cell-to-cell transport plant viruses encode a so-called "movement protein" and the principles for this process, involving passage of virus-modified plasmodesmata, have been elucidated for our model virus, Cowpea mosaic virus (CPMV). Understanding the local cell-to-cell movement of virus at the cellular level was a logical and essential prerequisite to embark for analysis of systemic spread at tissue and whole plant level. Systemic movement is a key process in the viral life cycle, as plants in which viral infection is limited to the initially infected cells or even leaves are functionally resistant under natural conditions. Relatively little is known about the transport of viruses over long distances. It is generally accepted that most, if not all, plant viruses are transported from the initial site of infection to other plant parts by the phloem, as the pattern of virus spread coincides with the translocation of photoassimilates. Detailed research of this aspect of virus movement has been hampered by lack of technology needed to study this problem. Recent introduction of the reporter gene green fluorescent protein (GFP) from the jellyfish Aequorea victoria, opens unique opportunities to study virus trafficking in whole plants, e.g. using laser scanning microscopy.
The major goal of the project is to determine and investigate the site and mechanism by which CPMV enters the vascular system for long-distance transport, which viral factors such as the movement protein and coat proteins are involved and how unloading and re-infection of plant cells occurs. This project is financially support by NWO-ALW, and is performed in collaboration with the WU Laboratory of Molecular Biology.
For general information on these research subjects send an email to Jan van Lent