Physaria fendleri and Ricinus communis lecithin: cholesterol acyltransferase-like phospholipases selectively cleave hydroxy acyl chains from phosphatidylcholine

Authors: XU, YANG - University Of Alberta; CALDO, KRISTIAN MARK - University Of Alberta; SINGER, STACY - University Of Alberta; MIETKIEWSKA, ELZBIETA - University Of Alberta; GREER, MICHAEL - University Of Alberta; TIAN, BO - University Of Alberta; Dyer, John; SMITH, MARK - Agriculture And Agri-Food Canada; ZHOU, XUE-RONG - Commonwealth Scientific And Industrial Research Organisation (CSIRO); QIU, XIAO - University Of Saskatchewan; WESELAKE, RANDALL - University Of Alberta; CHEN, GUANQUN - University Of Alberta

Submitted to: The Plant Journal
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/21/2020
Publication Date: 10/27/2020
Citation: Xu, Y., Caldo, K.P., Singer, S.D., Mietkiewska, E., Greer, M.S., Tian, B., Dyer, J.M., Smith, M., Zhou, X., Qiu, X., Weselake, R.J., Chen, G. 2020. Physaria fendleri and Ricinus communis lecithin: cholesterol acyltransferase-like phospholipases selectively cleave hydroxy acyl chains from phosphatidylcholine. The Plant Journal. 105(1):182-196. https://doi.org/10.1111/tpj.15050.
DOI: https://doi.org/10.1111/tpj.15050

Interpretive Summary: The majority of oilseed crops produce oil containing different ratios of just five basic fatty acids. However, there are hundreds of structurally diverse fatty acids produced in nature, and many non-domesticated plant species accumulate high amounts of a single "unusual" fatty acid in their seed oil. Examples include the hydroxy fatty acids present in the seed oils of castor bean or Physaria fendleri, which have potential usage in formulations of varnishes, cosmetics, surfactants, and engine oil lubricants. Castor bean and P. fendleri, however, have poor agronomic traits, which limits their utilization for production of these valuable feedstocks. Metabolic engineering offers an attractive approach for resolving this problem by transferring genes for hydroxy fatty acid synthesis to other crops that have better agronomic traits. Experiments to date have shown however, that relatively low amounts of hydroxy fatty acids are produced in engineered crops due to inefficiencies in transfer of these fatty acids into the oil of developing seeds. In the current study, an international group of scientists from the University of Alberta, Agriculture and Agri-Food Canada, the National Research Council of Canada, CSIRO in Australia, the Swedish University of Agicultural Sciences, and the ARS labs in Maricopa, Arizona, and Albany, California, identified genes in castor and Physaria fendleri that help increase the accumulation of hydroxy fatty acids in engineered plants. These results will be of greatest interest to other scientists interested in understanding the genes and enzymes required for production of high value oils in crop plants, as well as companies involved in the production and distribution of these oils for industrial partners and other end users.

Technical Abstract: Production of hydroxy fatty acids (HFAs) in transgenic crops represents a promising strategy to meet our demands for specialized plant oils with industrial applications. The expression of Ricinus communis (castor) OLEATE 12-HYDROXYLASE (RcFAH12) in Arabidopsis has resulted in only limited accumulation of HFAs in seeds, which probably results from inefficient transfer of HFAs from their site of synthesis (phosphatidylcholine; PC) to triacylglycerol (TAG), especially at the sn-1/3 positions of TAG. Phospholipase As (PLAs) may be directly involved in the liberation of HFAs from PC, but the functions of their over-expression in HFA accumulation and distribution at TAG in transgenic plants have not been well studied. In this work, the functions of lecithin:cholesterol acyltransferase-like PLAs (LCAT-PLAs) in HFA biosynthesis were characterized. The LCAT-PLAs were shown to exhibit homology to LCAT and mammalian lysosomal PLA2, and to contain a conserved and functional Ser/His/Asp catalytic triad. In vitro assays revealed that LCAT-PLAs from the HFA-accumulating plant species Physaria fendleri (PfLCAT-PLA) and castor (RcLCAT-PLA) could cleave acyl chains at both the sn-1 and sn-2 positions of PC, and displayed substrate selectivity towards sn-2-ricinoleoyl- PC over sn-2-oleoyl-PC. Furthermore, co-expression of RcFAH12 with PfLCAT-PLA or RcLCAT-PLA, but not Arabidopsis AtLCAT-PLA, resulted in increased occupation of HFA at the sn-1/3 positions of TAG as well as small but insignificant increases in HFA levels in Arabidopsis seeds compared with RcFAH12 expression alone. Therefore, PfLCAT-PLA and RcLCAT-PLA may contribute to HFA turnover on PC, and represent potential candidates for engineering the production of unusual fatty acids in crops.