No Induction of β-oxidation in leaves of Arabidopsis that over-produce lauric acid

Abstract.

Leaves from transgenic Brassica napus L. plants engineered to produce lauric acid show increased levels of enzyme activities of the pathways associated with fatty acid catabolism (V.A. Eccleston and J.B. Ohlrogge, 1998, Plant Cell 10: 613–621). In order to determine if the increases in enzyme activity are mirrored by increases in the expression of genes encoding enzymes of β-oxidation, which is the major pathway of fatty acid catabolism in plants, the medium-chain acyl-acyl carrier protein (ACP) thioesterase MCTE from California bay (Umbellularia californica) was over-expressed under the control of the cauliflower mosaic virus 35S promoter in Arabidopsisthaliana (L.) Heynh. Arabidopsis was the most suitable choice for these studies since gene expression could be analyzed in a large number of independent MCTE-expressing lines using already well-characterized β-oxidation genes. Levels of MCTE transcripts in leaves varied widely over the population of plants analyzed. Furthermore, active MCTE was produced as determined by enzymatic analysis of leaf extracts of MCTE-expressing plants. These plants incorporated laurate into triacylglycerol of seeds, but not into lipids of leaves as shown by gas-chromatographic analysis of total fatty acid extracts. The expression levels of the β-oxidation and other genes that are highly expressed during developmental stages involving rapid fatty acid degradation were measured. No significant difference in gene expression was observed among MCTE-expressing plants and transgenic and non-transgenic controls. To eliminate the possibility that post-translational mechanisms are responsible for the observed increases in enzyme activity acyl-CoA oxidase activity was also measured in leaves of MCTE-expressing plants using medium and long chain acyl-CoA substrates. No significant increases in either medium- or long-chain acyl-CoA oxidase activities were detected. We conclude that endogenous β-oxidation is sufficient to account for the complete degradation of laurate produced in rosette leaves of Arabidopsis expressing MCTE.