Abstract
Tomato produces a number of terpenes in their glandular trichomes that contribute to host plant resistance against pests. While glandular trichomes of cultivated tomato Solanum lycopersicum primarily accumulate a blend of monoterpenes, those of the wild tomato species Solanum habrochaites produce various sesquiterpenes. Recently, we have identified two groups of sesquiterpenes in S. habrochaites accessions that negatively affect the performance and choice behavior of the potato aphid (Macrosiphum euphorbiae). Aphids are piercing-sucking herbivores that use their mouthpart to penetrate and probe plant tissues in order to ultimately access vascular tissue and ingest phloem sap. Because secondary metabolites produced in glandular trichomes can affect the initial steps of the aphid feeding behavior, introducing the formation of defensive terpenes into additional plant tissues via metabolic engineering has the potential to reduce tissue penetration by aphids and in consequence virus transmission. Here, we have developed two multicistronic expression constructs based on the two sesquiterpene traits with activity toward M. euphorbiae previously identified in S. habrochaites. Both constructs are composed of sequences encoding a prenyl transferase and a respective S. habrochaites terpene synthase, as well as enhanced green fluorescent protein as a visible marker. All three coding sequences were linked by short nucleotide sequences encoding the foot-and-mouth disease virus 2A self-processing oligopeptide which allows their co-expression under the control of one promoter. Transient expression of both constructs under the epidermis-specific Arabidopsis CER5-promoter in tomato leaves demonstrated that formation of the two sets of defensive sesquiterpenes, β-caryophyllene/α-humulene and (−)-endo-α-bergamotene/(+)-α-santalene/(+)-endo-β-bergamotene, can be introduced into new tissues in tomato. The epidermis-specific transgene expression and terpene formation were verified by fluorescence microscopy and tissue fractionation with subsequent analysis of terpene profiles, respectively. In addition, the longevity and fecundity of M. euphorbiae feeding on these engineered tomato leaves were significantly reduced, demonstrating the efficacy of this novel aphid control strategy.
Highlights
In nature, plants are an integral part of a complex system of antagonistic and mutualistic biotic interactions
We have developed two multicistronic expression constructs based on the two sesquiterpene traits previously identified in S. habrochaites (Wang et al, 2020) that are composed of the coding sequences for prenyl transferases and respective S. habrochaites terpene synthases, as well as enhanced green fluorescent protein as a visible marker
To engineer formation of the two groups of sesquiterpenes with activity against M. euphorbiae that we have identified in S. habrochaites (Wang et al, 2020) into the epidermis of S. lycopersicum, we have designed two multicistronic expression constructs (Figure 1) in the binary vector pMCS:GW (Michniewicz et al, 2015)
Summary
Plants are an integral part of a complex system of antagonistic and mutualistic biotic interactions Due to their sessile lifestyle, plants have adapted to the resulting challenges by evolving specific strategies for the defense against attacking as well as the attraction of beneficial organisms. While terpenes accumulated in plant tissues can be toxic to biting-chewing and piercing-sucking herbivores, their emission into the surrounding atmosphere can act repellent to these herbivores and attractive toward their natural enemies (Degenhardt et al, 2003; Gershenzon and Dudareva, 2007; Unsicker et al, 2009) To facilitate their role in the antagonistic interactions, terpenes are often produced in specific plant tissues including internal ducts, extracellular cavities, and glandular trichomes (Gershenzon and Dudareva, 2007; Zulak and Bohlmann, 2010). The formation of terpenes is catalyzed by terpene synthases (TPSs) which utilize one or several of the prenyl diphosphate substrates, and frequently have the ability to form multiple different terpene products from one prenyl diphosphate substrate (Degenhardt et al, 2009)
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