The cytochrome P450 CYP86A22 is a fatty acyl-CoA omega-hydroxylase essential for Estolide synthesis in the stigma of Petunia hybrida.
The stigmatic estolide is a lipid-based polyester constituting the major component of exudate in solanaceous plants. Although the exudate is believed to play important roles in the pollination process, the biosynthetic pathway of stigmatic estolide, including genes encoding the key enzymes, remains unknown. Here we report the cloning and characterization of the cytochrome P450 gene CYP86A22, which encodes a fatty acyl-CoA omega-hydroxylase involved in estolide biosynthesis in the stigma of Petunia hybrida. A CYP86A22 cDNA was isolated from a developing stigma cDNA library, and the corresponding gene was shown to express predominantly in the developing stigma. Among six P450 genes isolated from this library, only CYP86A22 was implicated in omega-hydroxylation following RNA interference (RNAi)-mediated suppression. Unlike wild-type plants in which omega-hydroxy fatty acids (mainly in the form of 18-hydroxy oleic acid and 18-hydroxy linoleic acid) compose 96% of total stigma fatty acids, the omega-hydroxy fatty acids were essentially absent in the stigmas from 18 of 46 CYP86A22-RNAi transgenic plants and had varying levels of suppression in the remaining 28 plants. Furthermore, lipids in the 18 CYP86A22-RNAi stigmas were predominantly triacylglycerols and diacylglycerols instead of the estolides, which characterize the wild-type stigma. Analyses of recombinant CYP86A22 conclusively demonstrated that this P450 is a omega-hydroxylase with a substrate preference for both saturated and unsaturated acyl-CoAs rather than free fatty acids. We conclude that the cytochrome P450 enzyme CYP86A22 is the key fatty acyl-CoA omega-hydroxylase essential for the production of omega-hydroxy fatty acids and the biosynthesis of triacylglycerol-/diacylglycerol-based estolide polyesters in the petunia stigma.
Han
Jixiang
J
From the Donald Danforth Plant Science Center, St. Louis, Missouri 63132.
Clement
Joel M
JM
From the Donald Danforth Plant Science Center, St. Louis, Missouri 63132.
Li
Jia
J
From the Donald Danforth Plant Science Center, St. Louis, Missouri 63132.
King
Andrew
A
From the Donald Danforth Plant Science Center, St. Louis, Missouri 63132.
Ng
Shirley
S
From the Donald Danforth Plant Science Center, St. Louis, Missouri 63132.
Jaworski
Jan G
JG
From the Donald Danforth Plant Science Center, St. Louis, Missouri 63132. Electronic address: jjaworski@danforthcenter.org.
D000214:Acyl Coenzyme A / Q000378:metabolism; D000595:Amino Acid Sequence; D015152:Blotting, Northern; D003001:Cloning, Molecular; D042926:Cytochrome P-450 CYP4A / Q000235:genetics / Q000378:metabolism; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics* / Q000378:metabolism; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D005227:Fatty Acids / Q000378:metabolism; D035264:Flowers / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D018506:Gene Expression Regulation, Plant; D015723:Gene Library; D008055:Lipids / Q000737:chemistry; D008969:Molecular Sequence Data; D032306:Petunia / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D010940:Plant Proteins / Q000235:genetics* / Q000378:metabolism; D030821:Plants, Genetically Modified; D011091:Polyesters / Q000737:chemistry / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics; D017422:Sequence Analysis, DNA; D013379:Substrate Specificity
D000214:Acyl Coenzyme A; D018076:DNA, Complementary; D005227:Fatty Acids; D008055:Lipids; D010940:Plant Proteins; D011091:Polyesters; D034741:RNA, Small Interfering; D003577:Cytochrome P-450 Enzyme System; D042926:Cytochrome P-450 CYP4A