The transcriptional landscape of plant infection by the rice blast fungus Magnaporthe oryzae reveals distinct families of temporally co-regulated and structurally conserved effectors

Abstract

The rice blast fungus Magnaporthe oryzae causes a devastating disease which threatens global rice production. In spite of intense study, the biology of plant tissue invasion during blast disease remains poorly understood. Here we report a high resolution, transcriptional profiling study of the entire plant-associated development of the blast fungus. Our analysis revealed major temporal changes in fungal gene expression during plant infection. Pathogen gene expression could be classified into 10 modules of temporally co-expressed genes, providing evidence of induction of pronounced shifts in primary and secondary metabolism, cell signalling and transcriptional regulation. A set of 863 genes encoding secreted proteins are differentially expressed at specific stages of infection, and 546 were predicted to be effectors and named MEP (Magnaportheeffector protein) genes. Computational prediction of structurally-related MEPs, including the MAX effector family, revealed their temporal co-regulation in the same co- expression modules. We functionally characterised 32 MEP genes and demonstrate that Mep effectors are predominantly targeted to the cytoplasm of rice cells via the biotrophic interfacial complex (BIC), and use a common unconventional secretory pathway. Taken together, our study reveals major changes in gene expression associated with blast disease and identifies a diverse repertoire of effectors critical to successful infection.

Rgs1 is a regulator of effector gene expression during plant infection by the rice blast fungus Magnaporthe oryzae

“…MEP2 was identified in a recent study that characterised the effector repertoire of M. oryzae based on their differential expression during pathogenesis (25). MEP2 shows peak expression 48h after infection based on RNA-seq analysis, corresponding to a time of rapid plant tissue biotrophic colonisation by the fungus (25). We generated a MEP2:GFP gene fusion, transformed it into a M. oryzae wild type rice pathogenic strain Guy11 and selected a transformant with a single integration of the reporter gene construct.…”

“…S7 A ). This cluster also contains many effector-encoding genes (25). Consistent with this, we found 98 predicted or known effector-encoding genes upregulated in conidia of Δ rgs1 mutants, of which 60 were also up-regulated in conidia of the cer7 mutant (Fig.…”

“…This suggests that in addition to MEP2 , Rgs1 may regulate a much larger group of effectors, that are de-repressed in conidia when the function of Rgs1 is compromised. Among this group, there were 8 previously reported effector candidates, including BAS3 , BAS113 , MEP19 , and MEP27 (8, 25), and the necrosis and ethylene-inducing (Nep-like) peptide effector NLP4 which is involved in programmed cell death in plant tissue (34) (Fig. 4 D ).…”

“…5 C ). We did not find a clear reduction in disease symptoms in lesion density so we decided to use a recently described relative fitness assay to evaluate the effect of over-expressing Rgs1 (25). For this, we used a mixed spore inoculum of the ToxAp:Rgs1-GFP strain and a wild type Guy11 strain expressing H1-RFP.…”

“…Indeed, one of the main strategies for identifying putative effector candidates has been to use transcriptional profiling to identify differentially expressed pathogen genes. This has revealed the presence of temporally co-expressed families of effector genes in many fungal pathogens including Ustilago maydis (37) and M. oryzae (25). In rice blast, a very large repertoire of 546 effector genes has been reported, which are temporally co-expressed throughout invasive growth between 8h and 144h after infection, with large families of structurally-related effectors co-expressed between 24h and 48h after infection during biotrophic proliferation of the fungus in rice tissue (25).…”