NO: A nitric oxide synthase generated chemical effector of fungal pathogenesis

The world relies heavily on a handful of food crops. Of these, rice forms the staple diet of half of the world's population. However, rice and 50 other known grasses, are vulnerable to attack by the rice blast fungus. Such attack can lead to very significant crop losses. This may have implications for UK cereal crops - if global climate change leads to a rise in ambient temperature, rice blast disease will infiltrate Europe. Currently, disease is moderately well controlled by fungicides or by the use of rice varieties bred to carry extra disease resistance genes. Both are problematic: fungicides are costly, antifungal activity can be lost and inbred disease resistance can be lost due to the emergence of new and aggressive races of the fungus. Whilst we have made great strides towards understanding fungal invasion, much remains to be discovered. Detailed knowledge of the processes underpinning fungal attack will allow us to identify weak links during the early stages of fungal attack. This will empower us with the knowledge to design more environmentally benign modes of crop protection. This project aims to look at signals the fungus uses to drive the formation of its infection structures. More specifically, it will look at the generation and consequence of chemical effectors of fungal disease.

The research aims to determine whether generation of the transient and simple molecule NO, by nitric oxide synthase, influences germination and appressorium formation in Magnaporthe grisea. The work builds on preliminary data which shows that the transcript level of one of 4 putative NOS genes in M. grisea is highly up-regulated coincident with penetration (NOS3) and one significantly up-regulated coincident with germination (NOS2). Gene knockout showed highly attenuated disease levels in the NOS3 mutant. Appressorium formation and disease levels were restored by exogenous cAMP and by the NO donors SNP and PAPANONOate. These results demonstrate a link between NOS and NO and with signalling cascades. The proposed work is to a) to generate single and double NOS knockout and recomplementation strains b) to determine whether NOS2 and NOS3 synthesise NO, and when and where NO burst is c) to determine the cellular localisation of the proteins d) to unmask links between NO/NOS and signal relay pathways e) to identify both down-stream targets and genes involved in NO/NOS activity.