Regiobiochemical analysis reveals the role of castor LPAT2 in the accumulation of hydroxy fatty acids in transgenic lesquerella seeds

Chen, Grace Qianhong; Lin, Jiann‐Tsyh; Erp, Harrie van; Johnson, Kumiko; Lu, Chaofu

doi:10.1016/j.bcab.2020.101617

Cited by 5 publications

(5 citation statements)

References 37 publications

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“…In vitro enzyme assays have indicated that RcLPAT2 has a strong preference for 18:1OH-CoA when sn -1-18:1OH-LPA was used as an acyl acceptor; however, when sn -1-18:1-LPA was used as an acyl acceptor, RcLPAT2 selected 18:1-CoA [ 90 ]. The phenomenon of RcLPAT2 having different specificities toward acyl-donor and acyl-acceptor is supported by in vivo analysis of TAG species in transgenic lesquerella lines expressing RcLPAT2 [ 55 ]. Interestingly, RcLPAT-B is expressed in diverse organs [ 49 , 90 ], and exhibits broad substrate specificity, with saturated CoAs (12:0–16:0) being the preferred substrates [ 90 ].…”

Section: Resultsmentioning

confidence: 99%

“…To increase the HFA content at the sn -2 position of lesquerella TAGs, we have overexpressed RcLPAT2 in lesquerella seeds, and the resulting transgenic seed oil increases 3-HFA-TAGs (TAGs with all three sn positions acylated with HFAs) from 5% to 13–14% [ 18 ]. Regiochemical analysis reveals that RcLPAT2 increases the 3-HFA-TAGs content by enhancing the acylation of 18:1OH at the sn -2 position of 20:1OH-LPA for the subsequent generation of sn -1/3-20:1OH- sn -2-18:1OH-TAG [ 55 ]. The limited accumulation of HFAs in transgenics indicates that additional genes are required to implement new strategies for increased HFA production in lesquerella.…”

Section: Introductionmentioning

confidence: 99%

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Transcriptome Analysis and Identification of Lipid Genes in Physaria lindheimeri, a Genetic Resource for Hydroxy Fatty Acids in Seed Oil

Chen

Kim

Johnson

et al. 2021

IJMS

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Hydroxy fatty acids (HFAs) have numerous industrial applications but are absent in most vegetable oils. Physaria lindheimeri accumulating 85% HFA in its seed oil makes it a valuable resource for engineering oilseed crops for HFA production. To discover lipid genes involved in HFA synthesis in P. lindheimeri, transcripts from developing seeds at various stages, as well as leaf and flower buds, were sequenced. Ninety-seven percent clean reads from 552,614,582 raw reads were assembled to 129,633 contigs (or transcripts) which represented 85,948 unique genes. Gene Ontology analysis indicated that 60% of the contigs matched proteins involved in biological process, cellular component or molecular function, while the remaining matched unknown proteins. We identified 42 P. lindheimeri genes involved in fatty acid and seed oil biosynthesis, and 39 of them shared 78–100% nucleotide identity with Arabidopsis orthologs. We manually annotated 16 key genes and 14 of them contained full-length protein sequences, indicating high coverage of clean reads to the assembled contigs. A detailed profiling of the 16 genes revealed various spatial and temporal expression patterns. The further comparison of their protein sequences uncovered amino acids conserved among HFA-producing species, but these varied among non-HFA-producing species. Our findings provide essential information for basic and applied research on HFA biosynthesis.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Transcriptome Analysis and Identification of Lipid Genes in Physaria lindheimeri, a Genetic Resource for Hydroxy Fatty Acids in Seed Oil

Chen

Kim

Johnson

et al. 2021

IJMS

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“…During seed development, a lesquerella LPAT acts like a typical plant LPAT that has substrate preference for unsaturated FAs including18:1-CoA, resulting in efficient incorporation of 18:1-CoA into TAG through the Kennedy pathway. Castor RcLPAT2 is useful for increasing 18:1OH at the sn-2 of TAGs in lesquerella [15,37]. Additional isoforms, RcLPAT3B and RcLPATB, have also been shown to increase 18:1OH in arabidopsis seed TAGs [52].…”

Section: Constrains and Potential For Production Of A High 18:1oh-containing Oil In Lesquerellamentioning

confidence: 97%

“…Finally, the sn-3 position of DAG is acylated by 1,2-sn-diacylglycerol acyltransferase (DGAT) to produce TAG. Lesquerella TAGs contain~60% 20:1OH, and almost all of 20:1OH are acylated to the sn-1 and sn-3 positions, and the sn-2 positions of lesquerella TAGs are exclusively occupied by unsaturated FAs, i.e., 18:1, 18:2 and 18:3 [34][35][36][37]. The reason for lack of HFA at the sn-2 position of TAG has been suggested, in part, by the selectivity of lesquerella LPAT (PfLPAT) for unsaturated FA [15], which is a common feature for most plant microsomal LPAT [38].…”

Section: Introductionmentioning

confidence: 99%

Genetic Engineering of Lesquerella with Increased Ricinoleic Acid Content in Seed Oil

Chen

Johnson

Nazarenus

et al. 2021

Plants

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Seeds of castor (Ricinus communis) are enriched in oil with high levels of the industrially valuable fatty acid ricinoleic acid (18:1OH), but production of this plant is limited because of the cooccurrence of the ricin toxin in its seeds. Lesquerella (Physaria fendleri) is being developed as an alternative industrial oilseed because its seeds accumulate lesquerolic acid (20:1OH), an elongated form of 18:1OH in seed oil which lacks toxins. Synthesis of 20:1OH is through elongation of 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 sequentially produce linoleic and linolenic acids. To develop lesquerella that produces 18:1OH-rich seed oils such as castor, 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 in the T2 generation, including a top line with 18:1OH reduced 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 biotechnological potential to tailor lesquerella seeds to produce castor-like industrial oil functionality.

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“…Li et al (2020) found that the content of TAG in double-mutant Arabidopsis seeds was 30% lower than that in conventional seeds by HPLC-ESI-MS. Similarly, HPLC-ESI-MS can also characterize the molecular species of TAG in transgenic Lesquerella oils and their regioisomers in the oil, providing technical support for determining transgenic oils (Chen et al, 2020). In addition, ESI-MS has further enabled the detection of TAG in transgenic oilseeds alone.…”

Section: Tag Characterizationmentioning

confidence: 99%

A review on the identification of transgenic oilseeds and oils

Pan

Xing

Zhang

et al. 2023

Journal of Food Science

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Transgenic technology can increase the quantity and quality of vegetable oils worldwide. However, people are skeptical about the safety of transgenic oil‐bearing crops and the oils they produce. In order to protect consumers’ rights and avoid transgenic oils being adulterated or labeled as nontransgenic oils, the transgenic detection technology of oilseeds and oils needs careful consideration. This paper first summarized the current research status of transgenic technologies implemented at oil‐bearing crops. Then, an inspection process was proposed to detect a large number of samples to be the subject rapidly, and various inspection strategies for transgenic oilseeds and oils were summarized according to the process sequence. The detection indicators included oil content, fatty acid, triglyceride, tocopherol, and nucleic acid. The detection technologies involved chromatography, spectroscopy, nuclear magnetic resonance, and polymerase chain reaction. It is hoped that this article can provide crucial technical reference and support for staff engaging in the supervision of transgenic food and for researchers developing fast and efficient monitoring methods in the future.

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Regiobiochemical analysis reveals the role of castor LPAT2 in the accumulation of hydroxy fatty acids in transgenic lesquerella seeds

Cited by 5 publications

References 37 publications

Transcriptome Analysis and Identification of Lipid Genes in Physaria lindheimeri, a Genetic Resource for Hydroxy Fatty Acids in Seed Oil

Transcriptome Analysis and Identification of Lipid Genes in Physaria lindheimeri, a Genetic Resource for Hydroxy Fatty Acids in Seed Oil

Genetic Engineering of Lesquerella with Increased Ricinoleic Acid Content in Seed Oil

A review on the identification of transgenic oilseeds and oils

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