Genetic engineering of lesquerella with increased ricinoleic acid content in seed oil

Authors: Chen, Grace; Johnson, Kumiko; NAZARENUS, TARA - University Of Nebraska; Ponciano, Grisel; MORALE, EVA - US Department Of Agriculture (USDA); CAHOON, EDGAR - University Of Nebraska

Submitted to: Plants
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/27/2021
Publication Date: 5/29/2021
Citation: Chen, G.Q., Johnson, K., Nazarenus, T.J., Ponciano, G.P., Morales, E., Cahoon, E.B. 2021. Genetic engineering of lesquerella with increased ricinoleic acid content in seed oil. Plants. 10(6). Article 1093. https://doi.org/10.3390/plants10061093.
DOI: https://doi.org/10.3390/plants10061093

Interpretive Summary: The conventional source of hydroxy fatty acid (HFA) is from castor (Ricinus communis), which contains 90% ricinoleic acid (18:1OH) in its seed oil. The fatty acid 18:1OH and its derivatives are used as raw materials for numerous industrial products, such as lubricants, plastics and surfactants. The production of castor oil is hampered by the presence of the toxin ricin and hyper-allergenic 2S albumins in its seeds. Lesquerella seed oil contains a major HFA, lesquerolic acid at 55%–60%, therefore, efforts have been made through plant breeding to develop lesquerella as a new oilseed crop for a safe source of HFA. In this study, we demonstrated that significant high levels of 18:1OH can be achieved by blocking the desaturation and elongation steps in lesquerella. Our results not only provide tools for engineering castor oil-producing lesquerella, but also enhance our understanding of the mechanisms of HFA synthesis.

Technical Abstract: Castor oil contains ricinoleic acid (18:1OH) with numerous industrial uses, but its seeds contain toxin ricin hampering its production. Lesquerella (Physaria fendleri) accumulates lesquerolic acid (20:1OH) in seed oil and is being developed as a safe industrial crop. Synthesis of 20:1OH is through elongation of ricinoleic acid (18:1OH) by a lesquerella elongase, PfKCS18. Oleic acid (18:1) is the substrate for 18:1OH synthesis, but it is also used by fatty acid desaturase 2 (FAD2) and FAD3 to produce linoleic and linolenic acids, respectively. To develop lesquerlla producing 18:1OH, RNA interference sequences targeting KCS18, FAD2 and FAD3 were introduced to lesquerella to suppress the elongation and desaturation steps. Seeds from transgenic lines had increased 18:1OH to 1.1–26.6% compared with that of 0.4–0.6% in wild type (WT) seeds. Multiple lines had reduced 18:1OH levels at second generation including the top line which dropped from 26.7% to 19%. Transgenic lines also accumulated more 18:1 than that of WT, indicating that 18:1 is not efficiently used for 18:1OH synthesis and accumulation. Factors limiting 18:1OH accumulation and new targets for further increasing 18:1OH production are discussed. Our results provide insights into complex mechanisms of oil biosynthesis in lesquerella and show the potential of biotechnology to tailor the oil functionality of this industrial oil feedstock.