Developmental specific expression and organelle targeting of the Escherichia coli fabD gene, encoding malonyl coenzyme A-acyl carrier protein transacylase in transgenic rape and tobacco seeds.
In both plants and bacteria, de novo fatty acid biosynthesis is catalysed by a type II fatty acid synthetase (FAS) system which consists of a group of eight discrete enzyme components. The introduction of heterologous, i.e. bacterial, FAS genes in plants could provide an alternative way of modifying the plant lipid composition. In this study the Escherichia coli fabD gene, encoding malonyl CoA-ACP transacylase (MCAT), was used as a model gene to investigate the effects of over-producing a bacterial FAS component in the seeds of transgenic plants. Chimeric genes were designed, so as not to interfere with the household activities of fatty acid biosynthesis in the earlier stages of seed development, and introduced into tobacco and rapeseed using the Agrobacterium tumefaciens binary vector system. A napin promoter was used to express the E. coli MCAT in a seed-specific and developmentally specific manner. The rapeseed enoyl-ACP reductase transit peptide was used successfully, as confirmed by immunogold labelling studies, for plastid targeting of the bacterial protein. The activity of the bacterial enzyme reached its maximum (up to 55 times the maximum endogenous MCAT activity) at the end of seed development, and remained stable in mature transgenic seeds. Significant changes in fatty acid profiles of storage lipids and total seed lipid content of the transgenic plants were not found. These results are in support of the notion that MCAT does not catalyse a rate-limiting step in plant fatty acid biosynthesis.
Verwoert
I I
II
Department of Genetics, Vrije Universiteit, Amsterdam, Netherlands.
van der Linden
K H
KH
Nijkamp
H J
HJ
Stuitje
A R
AR
D051080:Acyl-Carrier Protein S-Malonyltransferase; D000217:Acyltransferases / Q000096:biosynthesis / Q000235:genetics*; D016960:Agrobacterium tumefaciens / Q000235:genetics; D001483:Base Sequence; D001937:Brassica / Q000235:genetics*; D050753:Enoyl-(Acyl-Carrier-Protein) Reductase (NADH); D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins; D054889:Fatty Acid Synthase, Type II; D064429:Fatty Acid Synthases / Q000235:genetics; D005227:Fatty Acids / Q000032:analysis; D018507:Gene Expression Regulation, Developmental / Q000502:physiology*; D018506:Gene Expression Regulation, Plant; D005822:Genetic Vectors; D008055:Lipids / Q000032:analysis; D008969:Molecular Sequence Data; D010088:Oxidoreductases / Q000235:genetics; D030821:Plants, Genetically Modified; D010947:Plants, Toxic; D018087:Plastids / Q000378:metabolism*; D021382:Protein Sorting Signals / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis; D018749:RNA, Plant / Q000032:analysis; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D012639:Seeds / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D014026:Nicotiana / Q000235:genetics*; D014170:Transformation, Genetic
D029968:Escherichia coli Proteins; D005227:Fatty Acids; D008055:Lipids; D021382:Protein Sorting Signals; D012333:RNA, Messenger; D018749:RNA, Plant; D011993:Recombinant Fusion Proteins; D010088:Oxidoreductases; D050753:Enoyl-(Acyl-Carrier-Protein) Reductase (NADH); D000217:Acyltransferases; D051080:Acyl-Carrier Protein S-Malonyltransferase; C494968:fabD protein, E coli; D064429:Fatty Acid Synthases; D054889:Fatty Acid Synthase, Type II