Elucidating the Furanocoumarin Biosynthetic Pathway in Apium graveolens L.: Uncovering the Coordination of Core Enzymes in Both Functional Activity and Gene Localization

Furanocoumarins and their derivatives are found in various plant species and have attracted considerable attention due to their diverse biological activities. By analyzing the genomes of Apium Graveolens L. and Peucedanum praeruptorum Dunn, we characterized a set of candidate genes encoding key enzymes involved in furanocoumarin biosynthesis, including one prenyltransferase (AgPT1), cyclases (AgCOC1, PpCOC1 and PpCOC2), and methyltransferases (AgOMT1 and AgOMT2). Functional validation in Saccharomyces cerevisiae demonstrated that AgCOC1 and PpCOC2 accept both linear and angular substrates, whereas PpCOC1 accepts only linear substrates. Depending on the reaction conditions, these cyclases can produce compounds with either furan or pyran scaffolds. These findings reveal a previously unappreciated catalytic versatility of cyclases involved in furanocoumarin biosynthesis. Notably, the genes encoding the prenyltransferase and cyclases were found to be co-localized in the genome, which may significantly enhance the efficiency of furanocoumarin biosynthesis. This mechanism may account for the pronounced accumulation of furanocoumarins in Apiaceae plants. Finally, we provide the first evidence that AgOMT1 functions as a multifunctional methyltransferase capable of catalyzing the O-methylation modifications observed in furanocoumarins in A. graveolens. In conclusion, this study fills a research gap in our understanding of furanocoumarin biosynthesis and reveals that genes encoding cyclases and prenyltransferases are clustered in the genome, a pattern that arose during evolution.