Metabolic evidence for the involvement of a Δ4-palmitoyl-acyl carrier protein desaturase in petroselinic acid synthesis in coriander endosperm and transgenic tobacco cells

Edgar B. Cahoon, John B. Ohlrogge

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Abstract

We have previously demonstrated that the double bond of petroselinic acid (18:1 Δ6cis) in coriander (Coriandrum sativum L.) seed results from the activity of a 36-kD desaturase that is structurally related to the Δ9-stearoyl-acyl carrier protein (ACP) desaturase (E.B. Cahoon, J. Shanklin, J.B. Ohlrogge [1992] Proc Natl Acad Sci USA 89: 11184-11188). To further characterize the biosynthetic pathway of this unusual fatty acid, 14C-labeling experiments were conducted using developing endosperm of coriander. Studies were also performed using suspension cultures of transgenic tobacco (Nicotiana tabacum L.) that express the coriander 36-kD desaturase, and as a result produce petroselinic acid and Δ4-hexadecenoic acid. When supplied exogenously to coriander endosperm slices, [1-14C]palmitic acid and stearic acid were incorporated into glycerolipids but were not converted to petroselinic acid. This suggested that petroselinic acid is not formed by the desaturation of a fatty acid bound to a glycerolipid or by reactions involving acyl-coenzyme As (CoA). Instead, evidence was most consistent with an acyl-ACP route of petroselinic acid synthesis. For example, the exogenous feeding of [1-14C]lauric acid and myristic acid to coriander endosperm slices resulted in the incorporation of the radiolabels into long-chain fatty acids, including primarily petroselinic acid, presumably through acyl-ACP-associated reactions. In addition, using an in vitro fatty acid biosynthetic system, homogenates of coriander endosperm incorporated [2-14C]malonyl-CoA into petroselinic acid, of which a portion was detected in a putative acyl-ACP fraction. Furthermore, analysis of transgenic tobacco suspension cultures expressing the coriander 36-kD desaturase revealed significant amounts of petroselinic acid and Δ4-hexadecenoic acid in the acyl-ACP pool of these cells. Also presented is evidence derived from [U-14C]nonanoic acid labeling of coriander endosperm, which demonstrates that the coriander 36-kD desaturase positions double bonds relative to the carboxyl end of acyl-ACP substrates. The data obtained in these studies are rationalized in terms of a biosynthetic pathway of petroselinic acid involving the Δ4 desaturation of palmitoyl-ACP by the 36-kD desaturase followed by two-carbon elongation of the resulting Δ4-hexadecenoyl-ACP.

Original languageEnglish (US)
Pages (from-to)827-837
Number of pages11
JournalPlant physiology
Volume104
Issue number3
DOIs
StatePublished - Mar 1994

Fingerprint

Coriandrum
Acyl Carrier Protein
acyl carrier protein
Endosperm
Coriandrum sativum
endosperm
Tobacco
tobacco
genetically modified organisms
synthesis
acids
cells
Fatty Acids
lauric acid
acyl-(acyl-carrier-protein)desaturase
Coenzymes
Biosynthetic Pathways
coenzymes
fatty acids
Suspensions

ASJC Scopus subject areas

  • Physiology
  • Genetics
  • Plant Science

Cite this

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title = "Metabolic evidence for the involvement of a Δ4-palmitoyl-acyl carrier protein desaturase in petroselinic acid synthesis in coriander endosperm and transgenic tobacco cells",
abstract = "We have previously demonstrated that the double bond of petroselinic acid (18:1 Δ6cis) in coriander (Coriandrum sativum L.) seed results from the activity of a 36-kD desaturase that is structurally related to the Δ9-stearoyl-acyl carrier protein (ACP) desaturase (E.B. Cahoon, J. Shanklin, J.B. Ohlrogge [1992] Proc Natl Acad Sci USA 89: 11184-11188). To further characterize the biosynthetic pathway of this unusual fatty acid, 14C-labeling experiments were conducted using developing endosperm of coriander. Studies were also performed using suspension cultures of transgenic tobacco (Nicotiana tabacum L.) that express the coriander 36-kD desaturase, and as a result produce petroselinic acid and Δ4-hexadecenoic acid. When supplied exogenously to coriander endosperm slices, [1-14C]palmitic acid and stearic acid were incorporated into glycerolipids but were not converted to petroselinic acid. This suggested that petroselinic acid is not formed by the desaturation of a fatty acid bound to a glycerolipid or by reactions involving acyl-coenzyme As (CoA). Instead, evidence was most consistent with an acyl-ACP route of petroselinic acid synthesis. For example, the exogenous feeding of [1-14C]lauric acid and myristic acid to coriander endosperm slices resulted in the incorporation of the radiolabels into long-chain fatty acids, including primarily petroselinic acid, presumably through acyl-ACP-associated reactions. In addition, using an in vitro fatty acid biosynthetic system, homogenates of coriander endosperm incorporated [2-14C]malonyl-CoA into petroselinic acid, of which a portion was detected in a putative acyl-ACP fraction. Furthermore, analysis of transgenic tobacco suspension cultures expressing the coriander 36-kD desaturase revealed significant amounts of petroselinic acid and Δ4-hexadecenoic acid in the acyl-ACP pool of these cells. Also presented is evidence derived from [U-14C]nonanoic acid labeling of coriander endosperm, which demonstrates that the coriander 36-kD desaturase positions double bonds relative to the carboxyl end of acyl-ACP substrates. The data obtained in these studies are rationalized in terms of a biosynthetic pathway of petroselinic acid involving the Δ4 desaturation of palmitoyl-ACP by the 36-kD desaturase followed by two-carbon elongation of the resulting Δ4-hexadecenoyl-ACP.",
author = "Cahoon, {Edgar B.} and Ohlrogge, {John B.}",
year = "1994",
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journal = "Plant Physiology",
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T1 - Metabolic evidence for the involvement of a Δ4-palmitoyl-acyl carrier protein desaturase in petroselinic acid synthesis in coriander endosperm and transgenic tobacco cells

AU - Cahoon, Edgar B.

AU - Ohlrogge, John B.

PY - 1994/3

Y1 - 1994/3

N2 - We have previously demonstrated that the double bond of petroselinic acid (18:1 Δ6cis) in coriander (Coriandrum sativum L.) seed results from the activity of a 36-kD desaturase that is structurally related to the Δ9-stearoyl-acyl carrier protein (ACP) desaturase (E.B. Cahoon, J. Shanklin, J.B. Ohlrogge [1992] Proc Natl Acad Sci USA 89: 11184-11188). To further characterize the biosynthetic pathway of this unusual fatty acid, 14C-labeling experiments were conducted using developing endosperm of coriander. Studies were also performed using suspension cultures of transgenic tobacco (Nicotiana tabacum L.) that express the coriander 36-kD desaturase, and as a result produce petroselinic acid and Δ4-hexadecenoic acid. When supplied exogenously to coriander endosperm slices, [1-14C]palmitic acid and stearic acid were incorporated into glycerolipids but were not converted to petroselinic acid. This suggested that petroselinic acid is not formed by the desaturation of a fatty acid bound to a glycerolipid or by reactions involving acyl-coenzyme As (CoA). Instead, evidence was most consistent with an acyl-ACP route of petroselinic acid synthesis. For example, the exogenous feeding of [1-14C]lauric acid and myristic acid to coriander endosperm slices resulted in the incorporation of the radiolabels into long-chain fatty acids, including primarily petroselinic acid, presumably through acyl-ACP-associated reactions. In addition, using an in vitro fatty acid biosynthetic system, homogenates of coriander endosperm incorporated [2-14C]malonyl-CoA into petroselinic acid, of which a portion was detected in a putative acyl-ACP fraction. Furthermore, analysis of transgenic tobacco suspension cultures expressing the coriander 36-kD desaturase revealed significant amounts of petroselinic acid and Δ4-hexadecenoic acid in the acyl-ACP pool of these cells. Also presented is evidence derived from [U-14C]nonanoic acid labeling of coriander endosperm, which demonstrates that the coriander 36-kD desaturase positions double bonds relative to the carboxyl end of acyl-ACP substrates. The data obtained in these studies are rationalized in terms of a biosynthetic pathway of petroselinic acid involving the Δ4 desaturation of palmitoyl-ACP by the 36-kD desaturase followed by two-carbon elongation of the resulting Δ4-hexadecenoyl-ACP.

AB - We have previously demonstrated that the double bond of petroselinic acid (18:1 Δ6cis) in coriander (Coriandrum sativum L.) seed results from the activity of a 36-kD desaturase that is structurally related to the Δ9-stearoyl-acyl carrier protein (ACP) desaturase (E.B. Cahoon, J. Shanklin, J.B. Ohlrogge [1992] Proc Natl Acad Sci USA 89: 11184-11188). To further characterize the biosynthetic pathway of this unusual fatty acid, 14C-labeling experiments were conducted using developing endosperm of coriander. Studies were also performed using suspension cultures of transgenic tobacco (Nicotiana tabacum L.) that express the coriander 36-kD desaturase, and as a result produce petroselinic acid and Δ4-hexadecenoic acid. When supplied exogenously to coriander endosperm slices, [1-14C]palmitic acid and stearic acid were incorporated into glycerolipids but were not converted to petroselinic acid. This suggested that petroselinic acid is not formed by the desaturation of a fatty acid bound to a glycerolipid or by reactions involving acyl-coenzyme As (CoA). Instead, evidence was most consistent with an acyl-ACP route of petroselinic acid synthesis. For example, the exogenous feeding of [1-14C]lauric acid and myristic acid to coriander endosperm slices resulted in the incorporation of the radiolabels into long-chain fatty acids, including primarily petroselinic acid, presumably through acyl-ACP-associated reactions. In addition, using an in vitro fatty acid biosynthetic system, homogenates of coriander endosperm incorporated [2-14C]malonyl-CoA into petroselinic acid, of which a portion was detected in a putative acyl-ACP fraction. Furthermore, analysis of transgenic tobacco suspension cultures expressing the coriander 36-kD desaturase revealed significant amounts of petroselinic acid and Δ4-hexadecenoic acid in the acyl-ACP pool of these cells. Also presented is evidence derived from [U-14C]nonanoic acid labeling of coriander endosperm, which demonstrates that the coriander 36-kD desaturase positions double bonds relative to the carboxyl end of acyl-ACP substrates. The data obtained in these studies are rationalized in terms of a biosynthetic pathway of petroselinic acid involving the Δ4 desaturation of palmitoyl-ACP by the 36-kD desaturase followed by two-carbon elongation of the resulting Δ4-hexadecenoyl-ACP.

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