Regulated expression of the rat medium chain hydrolase gene in transgenic rape seed.
Medium chain hydrolase (MCH) is an enzyme which regulates the chain length of fatty acid synthesis specifically in the mammary gland of the rat. During lactation, MCH interacts with fatty acid synthase (FAS) to cause premature release of acyl chains, thus providing medium chain fatty acids for synthesis of milk fat. In this study we have investigated the ability of rat MCH to interact with the phylogenetically more distant FAS structure present in plant systems and to cause a perturbation of fatty acid synthesis. In in vitro experiments, addition of purified MCH to rapeseed homogenates was found to cause a significant perturbation of fatty acid synthesis towards medium chain length products. The rat MCH gene was expressed in transgenic oilseed rape using a seed specific rape acyl carrier protein (ACP) promoter and a rape ACP plastid targeting sequence. Western analysis showed MCH protein to be present in transgenic seed and for its expression to be developmentally regulated in concert with storage lipid synthesis. The chimaeric preprotein was correctly processed and immunogold labelling studies confirmed MCH to be localized within plastid organelles. However, fatty acid analysis of oil from MCH-expressing rape seed showed no significant differences to that from control seed.
Safford
R
R
Dept of Biological Sciences, University of Durham, UK.
Moran
M T
MT
De Silva
J
J
Robinson
S J
SJ
Moscow
S
S
Jarman
C D
CD
Slabas
A R
AR
D000818:Animals; D015153:Blotting, Western; D001937:Brassica / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D064429:Fatty Acid Synthases / Q000235:genetics* / Q000378:metabolism / Q000648:ultrastructure; D015971:Gene Expression Regulation, Enzymologic; D016253:Microscopy, Immunoelectron; D030821:Plants, Genetically Modified / Q000235:genetics*; D051381:Rats; D012639:Seeds / Q000737:chemistry; D013869:Thiolester Hydrolases / Q000235:genetics* / Q000378:metabolism / Q000648:ultrastructure
D064429:Fatty Acid Synthases; D013869:Thiolester Hydrolases; C022738:thioesterase II