Two Novel Transcriptional Regulators Are Essential for Infection-related Morphogenesis and Pathogenicity of the Rice Blast Fungus Magnaporthe oryzae

Abstract

The cyclic AMP-dependent protein kinase A signaling pathway plays a major role in regulating plant infection by the rice blast fungus Magnaporthe oryzae. Here, we report the identification of two novel genes, MoSOM1 and MoCDTF1, which were discovered in an insertional mutagenesis screen for non-pathogenic mutants of M. oryzae. MoSOM1 or MoCDTF1 are both necessary for development of spores and appressoria by M. oryzae and play roles in cell wall differentiation, regulating melanin pigmentation and cell surface hydrophobicity during spore formation. MoSom1 strongly interacts with MoStu1 (Mstu1), an APSES transcription factor protein, and with MoCdtf1, while also interacting more weakly with the catalytic subunit of protein kinase A (CpkA) in yeast two hybrid assays. Furthermore, the expression levels of MoSOM1 and MoCDTF1 were significantly reduced in both Δmac1 and ΔcpkA mutants, consistent with regulation by the cAMP/PKA signaling pathway. MoSom1-GFP and MoCdtf1-GFP fusion proteins localized to the nucleus of fungal cells. Site-directed mutagenesis confirmed that nuclear localization signal sequences in MoSom1 and MoCdtf1 are essential for their sub-cellular localization and biological functions. Transcriptional profiling revealed major changes in gene expression associated with loss of MoSOM1 during infection-related development. We conclude that MoSom1 and MoCdtf1 functions downstream of the cAMP/PKA signaling pathway and are novel transcriptional regulators associated with cellular differentiation during plant infection by the rice blast fungus.

Author Summary

Magnaporthe oryzae, the causal agent of rice blast disease, is an important model fungal pathogen for understanding the molecular basis of plant-fungus interactions. In M. oryzae, the conserved cAMP/PKA signaling pathway has been demonstrated to be crucial for regulating infection-related morphogenesis and pathogenicity, including the control of sporulation and appressorium formation. In this study, we report the identification of two novel pathogenicity-related genes, MoSOM1 and MoCDTF1, by T-DNA insertional mutagenesis. Our results show that MoSOM1 or MoCDTF1 are essential for sporulation, appressorium formatiom and pathogenicity, and also play a key role in hyphal growth, melanin pigmentation and cell surface hydrophobicity. Nuclear localization sequences and conserved domains of the MoSom1 and MoCdtf1 proteins are crucial for their biological function. MoSom1 interacts physically with the transcription factors MoCdtf1 and MoStu1. We also show evidence that MoSom1 has the capacity to interact with CpkA, suggesting that MoSom1 may act downstream of the cAMP/PKA signaling pathway to regulate infection-related morphogenesis and pathogenicity in M. oryzae. Our studies extend the current understanding of downstream components of the conserved cAMP/PKA pathway and its precise role in regulating infection-related development and cellular differentiation by M. oryzae.