description
- The rice blast fungus M. oryzae has emerged as a paradigm for the understanding of fungal aerial attack. M. oryzae also infect roots. Consequently, M. oryzae-rice interaction represents an excellent pathosystem for comparative studies of the distinct organ-specific mechanisms that occur during fungal colonisation of rice leaves and roots. We have characterised two pathogenicity-deficient mutants in M. oryzae lacking the karyopherin Msn5p/EXP5 [Plant Cell 22, 953 (2010)] and a novel RNA-binding protein (RBP35) present exclusively in filamentous fungi [PLoS Pathogens, under review]. This project aims to investigate both organ-specific and pos-transcriptional mechanisms that regulate the ability of M. oryzae to infect rice using the resources generated previously in my group. The work outlined in this proposal will extend this area of research through: 1. Deep sequencing experiments using infected rice leaves and roots with a wild-type M. oryzae strain to follow changes in gene expression in the fungus and in the host plant. 2. Tandem affinity purification experiments using EXP5-tagged with HA-FLAG to identify proteins and RNAs that directly interact with EXP5. 3. Tandem affinity purification experiments using highly conserved proteins of the M. oryzae polyadenylation machinery to identify additional protein components and use a combination of SELEX and bioinformatic approaches to discover alternative polyadenylation signals in M. oryzae genes. 4. In vivo visualisation of infection-related mRNAs including fungal effectors using the lambdaN/GFP system to analyse the involvement of RBP35 in mRNA transport and/or localisation. At present there is a lack of knowledge of the EXP5 cargoes in filamentous fungi and very little is known about the post-transcriptional mechanisms that regulate fungal plant infection. Further, this research will open exciting avenues about local control of translation of proteins significant for M. oryzae plant invasion (fungal effectors).