Contribution of a novel gene to lysergic acid amide synthesis in Metarhizium brunneum.

We knocked out easP in M. brunneum via a CRISPR/Cas9-based approach, and accumulation of LAH was reduced to less than half the amount observed in the wild type. Because LAH accumulation was reduced and not eliminated, we identified and mutated the only close homolog of easP in the M. brunneum genome, a gene we named estA. An easP/estA double mutant did not differ from the easP mutant in lysergic acid amide accumulation, indicating estA had no role in the pathway. We conclude EasP contributes to LAH accumulation but is not absolutely required. Either a gene encoding redundant function and lacking sequence identity with easP resides outside the ergot alkaloid synthesis gene cluster, or EasP plays an accessory role in the synthesis of LAH.