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  • 1
    ISSN: 1432-2048
    Keywords: Key words: Acyl-CoA oxidase ; Arabidopsis (β-oxidation) ; Beta-oxidation ; Gene expression ; Lauric acid ; Medium-chain thioesterase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: 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.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1617-4623
    Keywords: Cucumber (Cucumis sativus L.) ; Malate synthase ; Glyoxylate cycle ; Gene transcription ; Metabolic regulation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The malate synthase gene (ms) promoter in cucumber (Cucumis sativus L.) was investigated with the aim of distinguishing DNA sequences mediating regulation of gene expression by sugar, and expression following seed germination. Promoter deletions were constructed and their ability to direct expression of theβ-glucuronidase (gus) reporter gene was investigated in transgenicNicotiana plumbaginifolia. Gene expression was assayed in germinating seeds and developing seedlings (the germination response) and in seedlings transferred from light into darkness with and without sucrose (the sugar response). As progressively more of the promoter was deleted from the 5′ end, first the sugar response and then the germination response was lost. Thus, distinct regions of the promoter are required for carbohydrate control and for regulation of gene expression in response to germination. Sequence comparisons of thems promoter with that of the isocitrate lyase gene (icl) of cucumber have previously identified four IMH (ICL-MS Homology) sequences. One such sequence, IMH2, is shown here to be implicated in the sugar response of thems gene. The 17 bp sequence, which when deleted from thems gene results in loss of the germination response, contains a 14 bp sequence which is similar to a sequence in theicl promoter, which we refer to as IMH5. Furthermore, this sequence has similarity withamdI9-like sequences in filamentous fungi, which conferfacB-mediated acetate inducibility on several genes, including those encoding ICL and MS.
    Type of Medium: Electronic Resource
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