A novel gene from a secondary metabolism gene cluster is required for microsclerotia formation and virulence in Verticillium dahliae

Verticillum dahliae is a notorious plant pathogenic fungus, which can cause wilt disease on hundreds of plant species. The genome of V. dahliae has been sequenced and annotated, but incorrect gene annotations and unidentified transcribed regions persist. Using RNA-Seq combined with reverse-transcription PCR, we identified a novel transcribed gene, Nag1, located in a cluster of putative secondary metabolic genes whose roles remain unknown. Functional analysis of Nag1 by dsRNA-mediated gene silencing revealed that loss of Nag1 significantly decreased fungal growth and conidial production. In addition, Nag1-silenced mutants exhibited obvious defects in microsclerotia formation and fungal virulence. Consistent with the reduction in microsclerotia formation, melanin production and expression of genes involved in melanin biosynthesis were markedly reduced in Nag1-silenced mutants. Overall, our data suggest that Nag1 acts as an important regulator of fungal development, microsclerotia formation and secondary metabolism in V. dahliae.