Arabidopsis METHYLENETETRAHYDROFOLATE REDUCTASE 2 functions independently of PENETRATION 2 during primary immunity against rice blast

Non-host resistance (NHR) is the most durable and robust form of innate immunity, with a surge of interest in its role in crop improvement. Of the NHR genes identified against rice blast, a devastating disease caused by Magnaporthe oryzae, Arabidopsis PEN2 is indispensable for pre-penetration resistance to M. oryzae, while a consortium of genes orchestrates post-penetration resistance via lesser known mechanisms. We identified M. oryzae-susceptible mosA (mthfr2 pen2-3) from a randomly mutagenized Arabidopsis pen2-3 population using forward genetics. Analysis of T-DNA-inserted mthfr2 lines and pen2-3-complemented mosA lines revealed that MTHFR2-dependent resistance to M. oryzae is independent of PEN2. MTHFR2-defective plants exhibited higher accumulation of reactive oxygen species and expression of salicylic acid-dependent defense markers. MTHFR2-ligand docking revealed that A55V non-synonymous substitution in mosA altered ligand binding efficiency. This further affected the metabolomic profile of mosA, effectively allowing in vitro germination and development of M. oryzae conidia. Moreover, the loss-of-function mutation in mthfr2 (involved in the 1C metabolic pathway) potentiated mosA immunity against Pst DC3000. In conclusion, our findings showed that MTHFR2 is a positive modulator of NHR against M. oryzae. This work documents another layer of conserved yet divergent metabolomic defense in Arabidopsis regulated by folate-mediated 1C metabolism that has the potential to revolutionize crop improvement.