Plants synthesize a remarkable number of lineage- and tissue-specific specialized metabolites.These compounds exhibit diverse functions for plants, e.g. communication and defense, as well as for humans, e.g. medicine and food. The anti-insect and anti-microbial acylsugars are one class of specialized metabolites and accumulate in Solanaceae species. Despite being composed of the simple building blocks of sugar cores and acyl chains, acylsugars exhibit incredible structural diversity. This variation was previously demonstrated to impact plant pest mortality and oviposition. These factors suggest that characterizing the acylsugar diversity within Solanaceae species and understanding their biosynthesis can uncover how a biologically relevant trait has evolved. While acylsucroses are the most well-characterized acylsugar type, unusual acylinositols were characterized in three species of the large and megadiverse Solanum genus. In this study, the diversity and distribution of Solanum genus acylinositols were characterized and their biosynthetic pathway was investigated. I first characterized the trichome acylsugars of Clade II species Solanum melongena (brinjal eggplant) using liquid chromatography-mass spectrometry (LC-MS), gas chromatography (GC)-MS and nuclear magnetic resonance (NMR) spectroscopy, identifying eight unusual structures with inositol cores, inositol glycoside cores, and hydroxyacyl chains. LC-MS analysis of 31 Solanum DulMo clade, VANAns clade, and Clade II species revealed striking acylsugar diversity with some traits restricted to specific clades and species. Acylinositols were found in all three major clades while acylglucoses were restricted to the DulMo and VANAns species characterized. Unusual disaccharide sugar cores and medium- length hydroxyacyl chains were found to be widespread within the surveyed species. This investigation revealed inositol sugar cores as a predominant sugar core type and prompted an investigation into their biosynthesis. Utilizing an eggplant tissue-specific transcriptome and in vitro biochemistry, an acetyltransferase ACYLSUGAR ACYLTRANSFERASE 3-LIKE 1 (SmASAT3-L1) was characterized to act upon a triacylinositol glycoside. Analysis of S. melongena triacylinositol biosynthesis uncovered an in vitro pathway producing a triacylinositol identical to a plant triacylinositol, however, production of the correct products only occurred when accompanied by nonenzymatic acyl chain rearrangement. Using this pathway knowledge and previously developed transcriptomes and gene silencing methods, I determined that two other acylinositol-producing species, Solanum quitoense and Solanum nigrum, contain an analogous acylinositol biosynthetic pathway. These results support the hypothesis that there is a conserved pathway within two major Solanum clades, DulMo and Clade II, which evolved in part due to gene duplications and altered substrate specificity. This study not only highlights the enormous amount of plant chemical diversity but also the usefulness of comparative biochemistry to uncover evolutionary mechanisms underlying metabolic novelty.
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