De novopurine nucleotide biosynthesis mediated by amidophosphoribosyl transferase is required for conidiation and essential for the successful host colonization ofMagnaporthe oryzae

Abstract

Amidophosphoribosyl transferase catalyzes the first step of the purine nucleotide biosynthesis by converting 5-phosphoribosyl-1-pyrophosphate into 5-phosphoribosyl-1-amine. In this study, we identified and characterized the functions of MoAde4, an ortholog of yeast Ade4 in the rice blast fungus. MoAde4 is a 537-amino acid protein containing the GATase_6 and pribosyltran domains. Quantitative real-time PCR analysis showed MoADE4 transcripts were highly expressed during conidiation, early-infection, and late-infection stages of the fungus. Disruption of MoADE4 gene resulted in ΔMoade4 mutant exhibiting adenine, adenosine, and hypoxanthine auxotrophy on MM. Conidia quantification assays showed ΔMoade4 mutant was significantly reduced in sporulation. The conidia of ΔMoade4 mutant could still form appressoria but mostly failed to penetrate the rice cuticle. Pathogenicity test showed ΔMoade4 was completely nonpathogenic on rice and barley leaves which was attributed by failure of its infectious hyphae to colonize the host cells. The ΔMoade4 was defective in induction of strong host immunity and had its purine transporter genes repressed during in planta infection. Addition of exogenous adenine partially rescued conidiation and pathogenicity defects of the ΔMoade4 mutant on the barley and rice leaves. Localization assays showed that MoAde4 is located in the cytoplasm. Taken together, our results demonstrate that purine biosynthesis orchestrated by MoAde4 is required for fungal development, conidiation, more importantly, we found it to be essential for fungal pathogenicity not because of the appressorial formation, but appressorium penetration and host colonization during the plant infection of M. oryzae. Thus this findings suggests that purine biosynthesis could act as an important target for combating recalcitrant plant fungal pathogens.