Transcriptome analysis and identification of lipid genes in Physaria lindheimeri, a genetic resource for hydroxy fatty acids in seed oil

Authors: Chen, Grace; KIM, WON NYEONG - University Of Seoul; Johnson, Kumiko; PARK, MID-EUM - University Of Seoul; LEE, KYEONG-RYEOL - Jeonju University; KIM, HYUN UK - University Of Seoul

Submitted to: International Journal of Molecular Sciences
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/4/2021
Publication Date: 1/6/2021
Citation: Chen, G.Q., Kim, W., Johnson, K., Park, M., Lee, K., Kim, H. 2021. Transcriptome analysis and identification of lipid genes in Physaria lindheimeri, a genetic resource for hydroxy fatty acids in seed oil. International Journal of Molecular Sciences. 22(2). Article 514. https://doi.org/10.3390/ijms22020514.
DOI: https://doi.org/10.3390/ijms22020514

Interpretive Summary: Hydroxy fatty acid (HFA) and its derivatives are used as raw materials for numerous industrial products, such as lubricants, plastics and surfactants. The conventional source of HFA is from castor (Ricinus communis) which contains 90% ricinoleic acid (12-hydroxy 9-octadecenoic acid, 18:1OH) in its seed oil. The production of castor oil, however, is hampered by the presence of the toxin ricin and hyper-allergic 2S albumins in its seeds. The wild Brassicaceae species Physaria lindheimeri accumulates up to 85% HFA in its seed oil, therefore is a valuable genetic resource for engineering oilseed crops for HFA production. In this study, we describe P. lindheimeri transcriptomes constructed from leaf, flower bud, and developing seeds at various stages. Lipid genes involved in fatty acid and TAG biosynthesis were identified by deep mining of the transcriptomes. Detailed spatial and temporal expression profiles of key genes were characterized by quantitative polymerase chain reaction (qPCR). P. lindheimeri sequences were further compared with orthologs from HFA-producing castor and lesquerella, as well as orthologs from non-HFA-producing species including Arabidopsis and Camelina. Our findings provide essential information for future basic and applied research on HFA biosynthesis in oilseeds.

Technical Abstract: Hydroxy fatty acids (HFA) have numerous industrial applications but are absent in most commercial vegetable oils. The wild Brassicaceae species Physaria lindheimeri accumulates up to 85% HFA in its seed oil, therefore is a valuable genetic resource for engineering oilseed crops for HFA production. To discover lipid genes involved in HFA synthesis in P. lindheimeri, transcripts from developing seeds at early, middle and late stages, as well as leaf and flower bud were sequenced. A total number of 552,614,582 raw reads were obtained and 97% clean reads were used for de novo assembly, generating 129,633 contigs (or transcripts) for 85,948 unique genes. Gene Ontology analysis indicated that 24%, 19% and 17% of the contigs matched proteins involved in biological process, cellular component and molecular function, respectively. The remaining 40% of the contigs did not match proteins with known functions. Compared with leaf sample, developing seeds had a total of 5,309 differentially expressed genes (DEG) whereas flower bud had only 219 DEG. Based on known Arabidopsis genes, we identified 38 P. lindheimeri orthologs involved in fatty acid and seed oil biosynthesis, most (95%) of them shared high (85–97%) nucleotide identity with Arabidopsis genes. We manually annotated 16 key P. lindheimeri genes and 14 of them contained full-length protein sequences, indicating high coverage of clean reads to the assembled contigs. Detailed profiling of the 16 genes revealed various spatial and temporal expression patterns. Further comparison of their protein sequences uncovered amino acids conserved among HFA-producing species but varied among non-HFA-producing species. Our findings provide essential information for future basic and applied research on HFA biosynthesis in oilseeds.