The trichome-specific acetolactate synthase NtALS1 gene, is involved in acylsugar biosynthesis in tobacco (Nicotiana tabacum L.)

Chang, Ai-Xia; Chen, Biao; Yang, Aiguo; Hu, Risheng; Feng, Qi Sheng; Chen, Ming; Yang, Xiaoning; Luo, Chuping; Li, Yang-Yang; Wang, Yuanying

doi:10.1007/s00425-020-03418-x

Cited by 6 publications

(3 citation statements)

References 53 publications

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“…Many enzymes involved in acylsugar biosynthesis have been identified in recent years ( Ning et al, 2015 ; Fan et al, 2016 , 2020 ; Moghe et al, 2017 ; Nadakuduti et al, 2017 ; Chang et al, 2020 , 2022 ; Mandal et al, 2020 ; Feng et al, 2021 ; Lou et al, 2021 ; Schenck et al, 2022 ). To identify additional enzymes in this pathway, and hopefully regulatory proteins, we used a multipronged strategy to find genes whose expression and evolutionary patterns suggested they could be involved.…”

Section: Discussionmentioning

confidence: 99%

Specialized metabolism by trichome-enriched Rubisco and fatty acid synthase components

Mandal

Rezenom

et al. 2022

Plant Physiology

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Acylsugars, specialized metabolites with defense activities, are secreted by trichomes of many solanaceous plants. Several acylsugar metabolic genes (AMGs) remain unknown. We previously reported multiple candidate AMGs. Here, using multiple approaches, we characterized additional AMGs. First, we identified differentially expressed genes between high- and low-acylsugar-producing F2 plants derived from a cross between cultivated tomato (Solanum lycopersicum) and a wild relative (S. pennellii), which produce acylsugars that are ∼1% and ∼20% of leaf dry weight, respectively. Expression levels of many known and candidate AMGs positively correlated with acylsugar amounts in F2 individuals. Next, we identified lycopersicum-pennellii putative orthologs with higher nonsynonymous to synonymous substitutions. These analyses identified four candidate genes, three of which showed enriched expression in stem trichomes compared to underlying tissues (shaved stems). Virus-induced gene silencing confirmed two candidates, Sopen05g009610 [beta-ketoacyl-(acyl-carrier-protein) reductase; fatty acid synthase component] and Sopen07g006810 (Rubisco small subunit), as AMGs. Phylogenetic analysis indicated that Sopen05g009610 is distinct from specialized metabolic cytosolic reductases but closely related to two capsaicinoid biosynthetic reductases, suggesting evolutionary relationship between acylsugar and capsaicinoid biosynthesis. Analysis of publicly available datasets revealed enriched expression of Sopen05g009610 orthologs in trichomes of several acylsugar-producing species. Similarly, orthologs of Sopen07g006810 were identified as solanaceous trichome-enriched members, which form a phylogenetic clade distinct from those of mesophyll-expressed “regular” Rubisco small subunits. Furthermore, δ13C analyses indicated recycling of metabolic CO2 into acylsugars by Sopen07g006810 and showed how trichomes support high levels of specialized metabolite production. These findings have implications for genetic manipulation of trichome specialized metabolism in solanaceous crops.

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Section: Discussionmentioning

confidence: 99%

Specialized metabolism by trichome-enriched Rubisco and fatty acid synthase components

Mandal

Rezenom

et al. 2022

Plant Physiology

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“…The constructed plasmids pCAMBIA1300-CsLCYB-GFP, pCAMBIA1300-CsLCYE-GFP, and pCAMBIA1300-GFP (Table S1) were transformed into the Agrobacterium tumefaciens strain GV1301 for transient expression in the leaf epidermal cells of Nicotiana benthamiana via Agrobacterium infiltration [44]. A confocal laser microscope (Nikon, Tokyo, Japan) was used to observe the infiltrated tobacco leaves, with the GFP signal and chloroplast autofluorescence signal being captured by 488 nm and 640 nm excitation lasers, respectively [45].…”

Section: Subcellular Localizations Of Cslcyb and Cslcyementioning

confidence: 99%

Functional Characterization of Lycopene β- and ε-Cyclases from a Lutein-Enriched Green Microalga Chlorella sorokiniana FZU60

Fang

Liu

et al. 2023

Marine Drugs

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Lutein is a high-value carotenoid with many human health benefits. Lycopene β- and ε-cyclases (LCYB and LCYE, respectively) catalyze the cyclization of lycopene into distinct downstream branches, one of which is the lutein biosynthesis pathway, via α-carotene. Hence, LCYB and LCYE are key enzymes in lutein biosynthesis. In this study, the coding genes of two lycopene cyclases (CsLCYB and CsLCYE) of a lutein-enriched marine green microalga, Chlorella sorokiniana FZU60, were isolated and identified. A sequence analysis and computational modeling of CsLCYB and CsLCYE were performed using bioinformatics to identify the key structural domains. Further, a phylogenetic analysis revealed that CsLCYB and CsLCYE were homogeneous to the proteins of other green microalgae. Subcellular localization tests in Nicotiana benthamiana showed that CsLCYB and CsLCYE localized in chloroplasts. A pigment complementation assay in Escherichia coli revealed that CsLCYB could efficiently β-cyclize both ends of lycopene to produce β-carotene. On the other hand, CsLCYE possessed a strong ε-monocyclase activity for the production of δ-carotene and a weak ε-bicyclic activity for the production of ε-carotene. In addition, CsLCYE was able to catalyze lycopene into β-monocyclic γ-carotene and ultimately produced α-carotene with a β-ring and an ε-ring via γ-carotene or δ-carotene. Moreover, the co-expression of CsLCYB and CsLCYE in E. coli revealed that α-carotene was a major product, which might lead to the production of a high level of lutein in C. sorokiniana FZU60. The findings provide a theoretical foundation for performing metabolic engineering to improve lutein biosynthesis and accumulation in C. sorokiniana FZU60.

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“…Within the Solanaceae, acylsugars accumulate in glandular trichomes and are involved in defense against an array of biological pests (Leckie et al, 2016; Ghosh and Jones, 2017; Luu et al, 2017; Smeda et al, 2017; Feng et al, 2021; Kortbeek et al, 2021). Sugars and acyl-CoAs from core metabolism are acylsugar precursors, with the acyl-CoAs derived from branched chain amino acid (BCAA) or fatty acid metabolism (Kandra et al, 1990; Slocombe et al, 2008; Chang et al, 2020; Fan et al, 2020; Mandal et al, 2020). While acylsugars with sucrose cores are common across the Solanaceae, other sugar core types are found.…”

Section: Introductionmentioning

confidence: 99%

Natural Variation Meets Synthetic Biology: Promiscuous Trichome Expressed Acyltransferases from Nicotiana acuminata

Schenck

Anthony

Jacobs

et al. 2022

Preprint

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Acylsugars are defensive, trichome-synthesized sugar esters produced in plants across the Solanaceae (nightshade) family. Although assembled from simple metabolites and synthesized by a relatively short core biosynthetic pathway, tremendous within- and across- species acylsugar structural variation is documented across the family. To advance our understanding of the diversity and the synthesis of acylsugars within the Nicotiana genus, trichome extracts were profiled across the genus coupled with transcriptomics-guided enzyme discovery and in vivo and in vitro analysis. Differences in the types of sugar cores, numbers of acylations, and acyl chain structures contributed to over 300 unique annotated acylsugars throughout Nicotiana. Placement of acyl chain length into a phylogenetic context revealed that an unsaturated acyl chain type was detected in a few closely-related species. A comparative transcriptomics approach identified trichome-enriched Nicotiana acuminata acylsugar biosynthetic candidate enzymes. > 25 acylsugar variants could be produced in a single enzyme assay with four acylsugar acyltransferases (NacASAT1-4) together with structurally diverse acyl-CoAs and sucrose. Liquid chromatography coupled with mass spectrometry screening of in vitro products revealed the ability of these enzymes to make acylsugars not present in Nicotiana plant extracts. In vitro acylsugar production also provided insights into acyltransferase acyl donor promiscuity and acyl acceptor specificity as well as regiospecificity of some ASATs. This study suggests that promiscuous Nicotiana acyltransferases can be used as synthetic biology tools to produce novel and potentially useful metabolites.

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The trichome-specific acetolactate synthase NtALS1 gene, is involved in acylsugar biosynthesis in tobacco (Nicotiana tabacum L.)

Cited by 6 publications

References 53 publications

Specialized metabolism by trichome-enriched Rubisco and fatty acid synthase components

Specialized metabolism by trichome-enriched Rubisco and fatty acid synthase components

Functional Characterization of Lycopene β- and ε-Cyclases from a Lutein-Enriched Green Microalga Chlorella sorokiniana FZU60

Natural Variation Meets Synthetic Biology: Promiscuous Trichome Expressed Acyltransferases from Nicotiana acuminata

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