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Defence and defence suppression in plant pathogen interactions: the role of CaMV protein P6

Reference	BB/D017319/1
Principal Investigator / Supervisor	Dr Joel Milner
Co-Investigators / Co-Supervisors	Professor Ari Sadanandom
Institution	University of Glasgow
Department	School of Life Sciences
Funding type	Research
Value (£)	413,598
Status	Completed
Type	Research Grant
Start date	01/05/2006
End date	31/07/2009
Duration	39 months

Abstract

Plants possess an armoury of defence mechanisms that biotrophic pathogens must either evade or suppress in order to multiply successfully in their hosts. We have previously shown that infection by Cauliflower mosaic virus (CaMV), a compatible pathogen of Arabidopsis, activates expression of markers of Salicylic Acid (SA)-mediated defence signalling. We have also found that mutants that over-produce SA and are constitutively activated for SA-mediated defence show enhanced resistance to CaMV. However, the converse is not true for SA-deficient mutants, and SA-mediated defence responses, although activated, are not effective in limiting spread of CaMV in wild-type backgrounds. This can be explained by our recent discovery that a virus-encoded protein, P6, which has functions essential for replication and is also the major genetic determinant of pathogenicity, is a very potent suppressor of SA-dependent defence responses, blocking signalling downstream of SA, and also greatly increasing susceptibility to non-viral pathogens. We will express native, and mutant forms of P6 in transgenic Arabidopsis and then assay the plants for SA-responsiveness in order to delineate the domains of P6 that are required for defence suppression. We will also test the extent to which expression of P6 effects susceptibility to virulent and avirulent bacterial and fungal pathogens, the effect on suscptibility to a necrotrophic fungus and whether we can extend the host-range to organisms that are normally non-pathogens of Arabidopsis. To determine the mechanism of action of P6, we will use yeast-two hybrid and immune pulldowns to identify interacting partners. To minimize interactions with proteins involved in translational transactivation, we will use as bait mutant forms of P6 selected on the basis of our domain-characterization studies. We will characterize the interactions between P6 and its interactors both in vitro and in planta using a variety of approaches.

Summary

Plants possess a battery of defence mechanisms that protect them against the organisms that cause disease, these include viruses, fungi and bacteria. The plant must first recognize the presence of invading pathogens and then engage defence responses that interfere with the ability of the pathogen to multiply and spread. Pathogens in turn must either evade these defence responses or suppress them, and are constantly evolving mechanisms of their own to do so. Increasing evidence suggests that the ability to suppress host defences may be an essential feature of all biotrophic pathogens, i.e. those that can grow in living tissue. Many of the defence responses used by plants involve Salicylic Acid (SA) as a signalling intermediate (i.e. trigger). The presence of a pathogen, stimulates the synthesis and accumulation of SA, this in turn signals the activation of various defence responses throughout the plant. We have found that when a viral pathogen of Arabidopsis, Cauliflower mosaic virus (CaMV), infects the plants, it triggers the activation of SA-dependent responses. However, these do not appear to be effective in restricting the spread of the virus. This is because a protein, P6, which is encoded by the virus and synthesized in infected cells, acts very efficiently to block the defence responses that would ordinarily be activated by the increased amounts of SA. We have found that P6 can not only block defence against CaMV but also against a bacterial pathogen, allowing it to grow in an ecotype of Arabidopsis which is normally resistant. In the proposed programme we have set out to understand how P6 acts on defence-signalling. P6 is known to have several functions which are essential for the replication of CaMV, but which do not appear to be related to defence suppression. We will investigate whether the parts of the protein that are responsible for these replicative functions are separate from those that are responsible for defence suppression. This will be achieved by expressing the whole P6 protein or parts of it in transgenic plants. We will also investigate the effect of expressing P6 on the susceptibility to a range of fungal and bacterial pathogens. For example, can P6 extend the host range of pathogens that ordinarily cannot grow in Arabidopsis? P6 must work by interacting with other Arabidopsis proteins that are involved in defence signalling. We will identify the interacting partners using two parallel approaches, yeast two hybrid and immune-pulldown. We will use this information to try to determine how these interacting proteins might be functioning in defence signalling and how P6 blocks their action. This work falls directly within the BBSRC priority area of understanding the interactions between plants and their pathogens. Our long term aims are to characterize the mechanisms by which viruses and other biotrophic pathogens can evade the defence responses by which plants defend themselves.

Committee	Closed Committee - Plant & Microbial Sciences (PMS)
Research Topics	Crop Science, Microbiology, Plant Science
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

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