Elucidating the Fundamental Process of Methyl-(5hydroxymethyl) Furan-2-Carboxylate Toxin Biosynthesis in Curvularia lunata Causing Maize Leaf Spot

Maize leaf spot, which is caused by <i>Curvularia lunata</i> (Wakkre) Boedijn, was epidemic in the maize-growing regions of northeastern and northern China in the mid-1990s, where it led to large yield losses. Since then, the epidemic has evolved into a kind of common disease. In recent years, however, a tendency of becoming an epidemic disease again has been observed in some areas in China due to significant changes in climate, farming, systems and crop varieties. The significance of methyl-(5hydroxymethyl) furan-2-carboxylate (M5HF2C) as a nonspecific host toxin in causing maize leaf spot disease has been demonstrated in previous research. However, the key enzymes involved in M5HF2C toxin synthesis remain unclear. In our study, we demonstrate that the synthesis of M5HF2C toxin starts from a precursor substrate in the pathogen, furfural, which is then catalytically dehydrogenated into furoic acid via an alcohol dehydrogenase (CLADH6). The furoic acid was further confirmed as one of the raw materials for the biosynthesis of M5HF2C toxin based on deletion mutants of the alcohol dehydrogenase gene (<i>Cladh6</i>) in <i>C</i>. <i>lunata</i>, which had reduced M5HF2C toxin-producing ability; however, this ability could be restored in all deletion mutants through complementation with furoic acid, thereby confirming that furoic acid is an intermediate in the biosynthesis of M5HF2C toxin. In summary, the biosynthesis process of M5HF2C toxin in <i>C</i>. <i>lunata</i> involves three transformation steps: (1) from xylose to furfural; (2) then from furfural to furoic acid; and (3) eventually from furoic acid to M5HF2C toxin. Our research findings provide new clues in elucidating the major steps in the process of M5HF2C toxin biosynthesis in <i>C</i>. <i>lunata</i>.