Identification of BAHD acyltransferases associated with acylinositol biosynthesis in Solanum quitoense (naranjilla)

Abstract

Plants make a variety of specialized metabolites that can mediate interactions with animals, microbes, and competitor plants. Understanding how plants synthesize these compounds enables studies of their biological roles by manipulating their synthesis in vivo as well as producing them in vitro. Acylsugars are a group of protective metabolites that accumulate in the trichomes of many Solanaceae family plants.Acylinositol biosynthesis is of interest because it appears to be restricted to a subgroup of species within the Solanum genus. Previous work characterized a triacylinositol acetyltransferase involved in acylinositol biosynthesis in the Andean fruit plant Solanum quitoense (lulo or naranjilla). We characterized three additional S. quitoense trichome expressed enzymes and found that virus-induced gene silencing of each caused changes in acylinositol accumulation. pH was shown to influence the stability and rearrangement of the product of ASAT1H and could potentially play a role in acylinositol biosynthesis. Surprisingly, the in vitro triacylinositol products of these enzymes are distinct from those that accumulate in planta. This suggests that additional enzymes are required in acylinositol biosynthesis. These characterized S. quitoense enzymes, nonetheless, provide opportunities to test the biological impact and properties of these triacylinositols in vitro.

Trading acyls and swapping sugars: metabolic innovations inSolanumtrichomes

“…In that case, a subgroup of S. habrochaites accessions possessed a functional ASAT4 copy, but its low levels of gene expression correlated with reduced accumulation of acetylated acylsugars in these 'subgroup E' accessions (Kim et al, 2012). activities were described previously (Fan et al, 2017(Fan et al, , 2015Leong et al, 2022Leong et al, , 2020Lou et al, 2021;Moghe et al, 2017;Nadakuduti et al, 2017;Schenck et al, 2022;Schilmiller et al, 2015Schilmiller et al, , 2012. Only reported, positive activities are displayed with the black squares.…”

“…Bootstrap support was calculated from 100,000 ultrafast bootstrap replicates; values >=95 indicate strong support. Other than SmASAT3-L1, enzyme activities were described previously (Fan et al, 2017, 2015; Leong et al, 2022, 2020; Lou et al, 2021; Moghe et al, 2017; Nadakuduti et al, 2017; Schenck et al, 2022; Schilmiller et al, 2015, 2012). Only reported, positive activities are displayed with the black squares.…”

“…Protein expression occurred as previously described (Leong et al, 2022; Lou et al, 2021). Briefly, 50 mL cultures of picked transformation colonies were grown overnight at 37°C, shaking at 225 rpm in Luria-Bertani (LB) media (Neogen, Lansing, MI, USA) supplemented with 1% glucose (w/v).…”

“…Rather, our LC-MS data are consistent with the hypothesis that the acyldisaccharides observed in this study consist of glycosylated inositol cores. Recent in vitro biochemistry and in vivo genetic evidence in S. quitoense suggests that acylinositols are the precursors to acylinositol disaccharides in Clade II Solanum species (Leong et al, 2022(Leong et al, , 2019.…”

“…Of the 23,251 eggplant genes expressed in at least one of the three tissues, 745 were significantly enriched (log2 fold-change >2, p-value < 0.05) in trichomes compared to hypocotyls and roots, including 20 BAHDs (Tables S37 and S38). As shown in Table S39, we selected for further testing seven BAHDs that were abundantly expressed in trichomes (TPM > 200) and homologous to characterized Solanum ASATs (sequences in colored text and denoted with "*" in Figure 6B) (D'Auria, 2006;Fan et al, 2015;Leong et al, 2022Leong et al, , 2020Lou et al, 2021;Schenck et al, 2022). We expressed each of the seven candidates in Escherichia coli and tested for acetylation of Compound 1, AI3:16(4,4,8).…”