A metabolic regulon reveals early and late acting enzymes in neuroactive Lycopodium alkaloid biosynthesis

Abstract

Plants synthesize many diverse small molecules that affect function of the mammalian central nervous system, making them crucial sources of therapeutics for neurological disorders. A notable portion of neuroactive phytochemicals are lysine-derived alkaloids, but the mechanisms by which plants produce these compounds have remained largely unexplored. To better understand how plants synthesize these metabolites, we focused on biosynthesis of the Lycopodium alkaloids that are produced by club mosses, a clade of plants used traditionally as herbal medicines. Hundreds of Lycopodium alkaloids have been described, including huperzine A (HupA), an acetylcholine esterase inhibitor that has generated interest as a treatment for the symptoms of Alzheimer’s disease. Through combined metabolomic profiling and transcriptomics, we have identified a developmentally controlled set of biosynthetic genes, or potential regulon, for the Lycopodium alkaloids. The discovery of this putative regulon facilitated the biosynthetic reconstitution and functional characterization of six enzymes that act in the initiation and conclusion of HupA biosynthesis. This includes a type III polyketide synthase that catalyzes a crucial imine-polyketide condensation, as well as three Fe(II)/2-oxoglutarate–dependent dioxygenase (2OGD) enzymes that catalyze transformations (pyridone ring-forming desaturation, piperidine ring cleavage, and redox-neutral isomerization) within downstream HupA biosynthesis. Our results expand the diversity of known chemical transformations catalyzed by 2OGDs and provide mechanistic insight into the function of noncanonical type III PKS enzymes that generate plant alkaloid scaffolds. These data offer insight into the chemical logic of Lys-derived alkaloid biosynthesis and demonstrate the tightly coordinated coexpression of secondary metabolic genes for the biosynthesis of medicinal alkaloids.

Total Biosynthesis of the Tubulin-Binding Alkaloid Colchicine

“…When studying secondary metabolism, a common assumption is that genes from the same biosynthetic pathway will exhibit similar spatial and temporal expression patterns . While whole-pathway coordination is often observed, there are instances in which distinct portions of a given secondary metabolic pathway experience differential regulation. , In hindsight, this appears to be the case for the colchicine biosynthetic pathway.…”

Huperzine A and Its Neuroprotective Molecular Signaling in Alzheimer’s Disease

“…All the properties of H. serrata have been intensively studied in China, and most of the biological activity of H. serrata appears to be caused by the molecule Huperzine A (HupA). HupA is an unsaturated sesquiterpene alkaloid compound that effectively crosses the blood-brain barrier (BBB), acting as a mixed-competitive, reversible, and selective AChE inhibitor [4][5][6][7] with a half-life of 5 h in the bloodstream, reaching a peak concentration at approximately 60 min in humans, and its t 1 2 has been calculated to be approximately 4-5 h [8].…”

Identification of early quassinoid biosynthesis in the invasive tree of heaven (Ailanthus altissima) confirms evolutionary origin from protolimonoids

“…A key strategy to identify biosynthetic genes is to correlate metabolic profiles with gene expression across different producing and non-producing tissues ( Caputi et al, 2018 ; Dang et al, 2018 ; Nett et al, 2021 ). Despite the relevance of A. altissima as a globally invasive species, no comprehensive transcriptomic data is publicly available for this plant.…”

“…Our data show that all three early quassinoid biosynthetic pathway genes are co-expressed. It is well known that co-expression analyses are powerful tools for elucidating biosynthetic pathways in plants ( Caputi et al, 2018 ; Dang et al, 2018 ; Nett et al, 2021 ), and this also holds true for the early steps of quassinoid biosynthesis. We therefore anticipate that our data will also be useful for the identification of further pathway steps downstream of melianol.…”