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Lipidomic analysis of Arabidopsis seed genetically engineered to contain DHA.

Zhou XR, Callahan DL, Shrestha P, Liu Q, Petrie JR, Singh SP - Front Plant Sci (2014)

Bottom Line: Metabolic engineering of omega-3 long-chain (≥C20) polyunsaturated fatty acids (ω3 LC-PUFA) in oilseeds has been one of the key targets in recent years.Liquid chromatography-mass spectrometry (LC-MS) was used to characterize the triacylglycerol (TAG), diacylglycerol (DAG) and phospholipid (PL) lipid classes in the transgenic and wild type Arabidopsis seeds at both developing and mature stages.Trace amounts of di-DHA PC and tri-DHA TAG were identified and confirmed by high resolution MS/MS.

View Article: PubMed Central - PubMed

Affiliation: Food Futures National Research Flagship, Commonwealth Scientific and Industrial Research Organisation Canberra, ACT, Australia ; Plant Industry, Commonwealth Scientific and Industrial Research Organisation Canberra, ACT, Australia.

ABSTRACT
Metabolic engineering of omega-3 long-chain (≥C20) polyunsaturated fatty acids (ω3 LC-PUFA) in oilseeds has been one of the key targets in recent years. By expressing a transgenic pathway for enhancing the synthesis of the ω3 LC-PUFA docosahexaenoic acid (DHA) from endogenous α-linolenic acid (ALA), we obtained the production of fish oil-like proportions of DHA in Arabidopsis seed oil. Liquid chromatography-mass spectrometry (LC-MS) was used to characterize the triacylglycerol (TAG), diacylglycerol (DAG) and phospholipid (PL) lipid classes in the transgenic and wild type Arabidopsis seeds at both developing and mature stages. The analysis identified the appearance of several abundant DHA-containing phosphatidylcholine (PC), DAG and TAG molecular species in mature seeds. The relative abundances of PL, DAG, and TAG species showed a preferred combination of LC-PUFA with ALA in the transgenic seeds, where LC-PUFA were esterified in positions usually occupied by 20:1ω9. Trace amounts of di-DHA PC and tri-DHA TAG were identified and confirmed by high resolution MS/MS. Studying the lipidome in transgenic seeds provided insights into where DHA accumulated and combined with other fatty acids of neutral and phospholipids from the developing and mature seeds.

No MeSH data available.


Distribution of phosphatidylcholine (PC) and phosphatidic acid (PA) species in the developing and mature seeds from the WT or GA7 Arabidopsis seeds. The PC species (A) and PA species (B) are annotated as X:Y where X is the total carbon number and Y is the total double bond number in the two acyl chains. Data are shown as the mean of triplicate analysis with the error bars representing the standard deviations.
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Figure 4: Distribution of phosphatidylcholine (PC) and phosphatidic acid (PA) species in the developing and mature seeds from the WT or GA7 Arabidopsis seeds. The PC species (A) and PA species (B) are annotated as X:Y where X is the total carbon number and Y is the total double bond number in the two acyl chains. Data are shown as the mean of triplicate analysis with the error bars representing the standard deviations.

Mentions: Both the developing and mature seeds of GA7 produced new LC-PUFA in phospholipids. PC precursor total ion scan showed that the WT seeds contained PC with total acyl chain lengths only up to C38 (PC 38:Y, in which a total of 38 carbons occurs in the two acyl chains on PC with a total double bond number of Y). The most likely fatty acid combination of being C18 and C20 (Figure 3). The WT only had low amounts of C20 fatty acids in PC (Table 1). The developing GA7 seeds produced PC 40:Y, indicating the esterification of two C20 fatty acids. In the mature GA7 seeds, there were low yet detectable amounts of PC 42:Y and PC 44:Y, i.e., PC C20/C22 or PC C22/C22, indicating the accumulation of LC-PUFA in the PC pool. Furthermore, precursor ion scan showed higher abundance of the earlier eluted part of the each PC chain length group in GA7 when compared to WT. The molecular species with more double bonds eluted slightly earlier than the molecular species with fewer double bonds in the group with same chain length. This indicated that the PC clusters in GA7 contain more double bonds. This was further supported by the quantification of the different PC species as shown in Figure 4A.


Lipidomic analysis of Arabidopsis seed genetically engineered to contain DHA.

Zhou XR, Callahan DL, Shrestha P, Liu Q, Petrie JR, Singh SP - Front Plant Sci (2014)

Distribution of phosphatidylcholine (PC) and phosphatidic acid (PA) species in the developing and mature seeds from the WT or GA7 Arabidopsis seeds. The PC species (A) and PA species (B) are annotated as X:Y where X is the total carbon number and Y is the total double bond number in the two acyl chains. Data are shown as the mean of triplicate analysis with the error bars representing the standard deviations.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4150447&req=5

Figure 4: Distribution of phosphatidylcholine (PC) and phosphatidic acid (PA) species in the developing and mature seeds from the WT or GA7 Arabidopsis seeds. The PC species (A) and PA species (B) are annotated as X:Y where X is the total carbon number and Y is the total double bond number in the two acyl chains. Data are shown as the mean of triplicate analysis with the error bars representing the standard deviations.
Mentions: Both the developing and mature seeds of GA7 produced new LC-PUFA in phospholipids. PC precursor total ion scan showed that the WT seeds contained PC with total acyl chain lengths only up to C38 (PC 38:Y, in which a total of 38 carbons occurs in the two acyl chains on PC with a total double bond number of Y). The most likely fatty acid combination of being C18 and C20 (Figure 3). The WT only had low amounts of C20 fatty acids in PC (Table 1). The developing GA7 seeds produced PC 40:Y, indicating the esterification of two C20 fatty acids. In the mature GA7 seeds, there were low yet detectable amounts of PC 42:Y and PC 44:Y, i.e., PC C20/C22 or PC C22/C22, indicating the accumulation of LC-PUFA in the PC pool. Furthermore, precursor ion scan showed higher abundance of the earlier eluted part of the each PC chain length group in GA7 when compared to WT. The molecular species with more double bonds eluted slightly earlier than the molecular species with fewer double bonds in the group with same chain length. This indicated that the PC clusters in GA7 contain more double bonds. This was further supported by the quantification of the different PC species as shown in Figure 4A.

Bottom Line: Metabolic engineering of omega-3 long-chain (≥C20) polyunsaturated fatty acids (ω3 LC-PUFA) in oilseeds has been one of the key targets in recent years.Liquid chromatography-mass spectrometry (LC-MS) was used to characterize the triacylglycerol (TAG), diacylglycerol (DAG) and phospholipid (PL) lipid classes in the transgenic and wild type Arabidopsis seeds at both developing and mature stages.Trace amounts of di-DHA PC and tri-DHA TAG were identified and confirmed by high resolution MS/MS.

View Article: PubMed Central - PubMed

Affiliation: Food Futures National Research Flagship, Commonwealth Scientific and Industrial Research Organisation Canberra, ACT, Australia ; Plant Industry, Commonwealth Scientific and Industrial Research Organisation Canberra, ACT, Australia.

ABSTRACT
Metabolic engineering of omega-3 long-chain (≥C20) polyunsaturated fatty acids (ω3 LC-PUFA) in oilseeds has been one of the key targets in recent years. By expressing a transgenic pathway for enhancing the synthesis of the ω3 LC-PUFA docosahexaenoic acid (DHA) from endogenous α-linolenic acid (ALA), we obtained the production of fish oil-like proportions of DHA in Arabidopsis seed oil. Liquid chromatography-mass spectrometry (LC-MS) was used to characterize the triacylglycerol (TAG), diacylglycerol (DAG) and phospholipid (PL) lipid classes in the transgenic and wild type Arabidopsis seeds at both developing and mature stages. The analysis identified the appearance of several abundant DHA-containing phosphatidylcholine (PC), DAG and TAG molecular species in mature seeds. The relative abundances of PL, DAG, and TAG species showed a preferred combination of LC-PUFA with ALA in the transgenic seeds, where LC-PUFA were esterified in positions usually occupied by 20:1ω9. Trace amounts of di-DHA PC and tri-DHA TAG were identified and confirmed by high resolution MS/MS. Studying the lipidome in transgenic seeds provided insights into where DHA accumulated and combined with other fatty acids of neutral and phospholipids from the developing and mature seeds.

No MeSH data available.