Variant castor lysophosphatidic acid acyltransferases acylate ricinoleic acid in seed oil

Authors: KIM, HYUN UK - Sejong University; PARK, MID-EUM - Sejong University; LEE, KYEONG-RYEOL - Korean Rural Development Administration; SUH, MI-CHUNG - Sogang University; Chen, Grace

Submitted to: Industrial Crops and Products
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/16/2020
Publication Date: 4/2/2020
Citation: Kim, H., Park, M., Lee, K., Suh, M., Chen, G.Q. 2020. Variant castor lysophosphatidic acid acyltransferases acylate ricinoleic acid in seed oil. Industrial Crops and Products. 150:112245. https://doi.org/10.1016/j.indcrop.2020.112245.
DOI: https://doi.org/10.1016/j.indcrop.2020.112245

Interpretive Summary: Castor seed oil (triacylglycerols, TAG) contains 90% ricinoleic acid (18:1OH), a hydroxy fatty acid (HFA) with numerous industrial applications. Castor lysophosphatidic acid acyltransferases (RcLPATs) are key enzymes contributing to the accumulation of 18:1OH in TAG. To identify isoforms of RcLPATs capable of acylating 18:1OH, we characterized the entire family of seven RcLPATs. We discover that all of the seven RcLPATs encode functional enzyme. We also reveal that RcLPATs are expressed variably among castor organs and during seed development. Based on spatial and temporal expression patterns, we suggest that RcLPATs play distinct and redundant roles in castor organs and developing seeds. Moreover, we found that three RcLPATs increased hydroxy fatty acids from 16.4% to 18.2–21.9% in Arabidopsis. These new RcLPAT genes are new targets for genetic engineering of oilseeds for hydroxy fatty acid production.

Technical Abstract: Castor seed oil (triacylglycerols, TAG) contains 90% ricinoleic acid (18:1OH), a hydroxy fatty acid (HFA) with numerous industrial applications. Castor lysophosphatidic acid acyltransferases (RcLPATs) are key enzymes contributing to the accumulation of 18:1OH in TAG. To identify isoforms of RcLPATs capable of acylating 18:1OH, we characterized the entire family of seven RcLPATs. Expression of each RcLPATs in E. coli JC201 lpat mutant restored the LPAT activity indicating that all RcLPATs encode active enzymes. Based on spatial and temporal gene expression profiles, RcLPAT1 and RcLPAT2 may perform house-keeping function in plastid and ER, respectively. RcLPAT3A and RcLPAT3B may have a critical role in male flower development, but not in leaf. Besides, RcLPTA3B is not active in stem. RcLPAT3A and RcLPAT3B may also function as a house-keeping gene in unopened-flower buds and female flower. RcLPAT4 is the only isoform of class-A microsomal RcLPATs showed a high transcript level in leaf indicating its importance in maintaining leaf function. RcLPAT4 may play a house-keeping role in various other organs, but during seed development its expression might associate with late embryo development and/or seed coat selerification process. RcLPAT5 exhibited a constitutive expression pattern typical to a house-keeping gene during seed development, but its expression was low in the other organs. Castor has a class-B type RcLPATB gene, its transcript level was about the same high as that of RcLPAT4 in leaf, indicating that RcLPATB together with RcLPAT4 are two major microsomal RcLPATs functioning in leaf. When individual RcLPATs was expressed in CL37 background, an Arabidopsis line contains about 17% HFA in seed TAG, three out of seven RcLPATs (RcLPAT2, RcLPAT3B and RcLPATB) increased total HFA (18:1OH and densipolic acid 18:2OH) from an average of 16.6% in CL37 to averages of 18.2%, 21.9% and 20.3% in RcLPAT2-CL37, RcLPAT3B-CL27, and RcLPATB-CL37, respectively. Furthermore, we observed different accumulation levels between 18:1OH and 18:2OH among these three RcLPATs-CL37 transgenics. The mechanisms on substrate selectivity among RcLPAT2, RcLPAT3B and RcLPATB are discussed.