Production of mono- and sesquiterpenes in Camelina sativa oilseed

Main conclusion: Camelina was bioengineered to accumulate (4S)-limonene and (+)-δ-cadinene in seed. Plastidic localization of the recombinant enzymes resulted in higher yields than cytosolic localization. Overexpressing 1-deoxy-d-xylulose-5-phosphate synthase (DXS) further increased terpene accumulation. Many plant-derived compounds of high value for industrial or pharmaceutical applications originate from plant species that are not amenable to cultivation. Biotechnological production in low-input organisms is an attractive alternative. Several microbes are well established as biotechnological production platforms; however, their growth requires fermentation units, energy input, and nutrients. Plant-based production systems potentially allow the generation of high-value compounds on arable land with minimal input. Here we explore whether Camelina sativa (camelina), an emerging low-input non-foodstuff Brassicaceae oilseed crop grown on marginal lands or as a rotation crop on fallow land, can successfully be refactored to produce and store novel compounds in seed. As proof-of-concept, we use the cyclic monoterpene hydrocarbon (4S)-limonene and the bicyclic sesquiterpene hydrocarbon (+)-δ-cadinene, which have potential biofuel and industrial solvent applications. Post-translational translocation of the recombinant enzymes to the plastid with concurrent overexpression of 1-deoxy-d-xylulose-5-phosphate synthase (DXS) resulted in the accumulation of (4S)-limonene and (+)-δ-cadinene up to 7 mg g⁻¹ seed and 5 mg g⁻¹ seed, respectively. This study presents the framework for rapid engineering of camelina oilseed production platforms for terpene-based high-value compounds.