A bacterial-like Pictet-Spenglerase drives the evolution of fungi to produce β-carboline glycosides together with separate genes.

Abstract

Diverse β-carboline (βC) alkaloids are produced by microbes, plants, and animals with myriad bioactivities and drug potentials. However, the biosynthetic mechanism of βCs remains largely elusive, especially regarding the hydroxyl and glucosyl modifications of βCs. Here, we report the presence of the bacterial-like Pictet-Spenglerase gene <i>Fcs1</i> in the entomopathogenic <i>Beauveria</i> fungi that can catalyze the biosynthesis of the βC skeleton. The overexpression of <i>Fcs1</i> in <i>Beauveria bassiana</i> led to the identification of six βC methyl glycosides, termed bassicarbosides (BCSs) A-F. We verified that the cytochrome P450 (CYP) genes adjacent to <i>Fcs1</i> cannot oxidize βCs. Alternatively, the separated CYP684B2 family gene <i>Fcs2</i> was identified to catalyze βC hydroxylation together with its cofactor gene <i>Fcs3</i>. The functional homologue of <i>Fcs2</i> is only present in the <i>Fcs1-</i>containing fungi and highly similar to the <i>Fcs1-</i>connected yet nonfunctional CYP. Both evolved quicker than those from fungi without <i>Fcs1</i> homologues. Finally, the paired methyl/glucosyl transferase genes were verified to mediate the production of BCSs from hydroxy-βCs. All these functionally verified genes are located on different chromosomes of <i>Beauveria</i>, which is in contrast to the typical content-clustered feature of fungal biosynthetic gene clusters (BGCs). We also found that the production of BCSs selectively contributed to fungal infection of different insect species. Our findings shed light on the biosynthetic mechanism of βC glycosides, including the identification of a βC hydroxylase. The results of this study also propose an evolving process of fungal BGC formation following the horizontal transfer of a bacterial gene to fungi.