Ammonia-regulated expression of a soybean gene encoding cytosolic glutamine synthetase in transgenic Lotus corniculatus.
A full-length cDNA clone encoding cytosolic glutamine synthetase (GS), expressed in roots and root nodules of soybean, was isolated by direct complementation of an Escherichia coli gln A- mutant. This sequence is induced in roots by the availability of ammonia. A 3.5-kilobase promoter fragment of a genomic clone (lambda GS15) corresponding to this cDNA was isolated and fused with a reporter [beta-glucuronidase (GUS)] gene. The GS-GUS fusion was introduced into a legume (Lotus corniculatus) and a nonlegume (tobacco) plant by way of Agrobacterium-mediated transformations. This chimeric gene was found to be expressed in a root-specific manner in both tobacco and L. corniculatus, the expression being restricted to the growing root apices and the vascular bundles of the mature root. Treatment with ammonia increased the expression of this chimeric gene in the legume background (i.e., L. corniculatus); however, no induction was observed in tobacco roots. Histochemical localization of GUS activity in ammonia-treated transgenic L. corniculatus roots showed a uniform distribution across all cell types. These data suggest that the tissue specificity of the soybean cytosolic GS gene is conserved in both tobacco and L. corniculatus; however, in the latter case, this gene is ammonia inducible. Furthermore, the ammonia-enhanced GS gene expression in L. corniculatus is due to an increase in transcription. That this gene is directly regulated by externally supplied or symbiotically fixed nitrogen is also evident from the expression of GS-GUS in the infection zone, including the uninfected cells, and the inner cortex of transgenic L. corniculatus nodules, where a flux of ammonia is encountered by this tissue. The lack of expression of GS-GUS in the outer cortex of the nodules suggests that ammonia may not be able to diffuse outside the endodermis.
Department of Molecular Genetics and Biotechnology Center, Ohio State University, Columbus 43210.
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Miao
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Hirel
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Marsolier
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Ridge
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Verma
D000595:Amino Acid Sequence; D000641:Ammonia / Q000494:pharmacology*; D001483:Base Sequence; D003600:Cytosol / Q000201:enzymology; D007887:Fabaceae / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D005974:Glutamate-Ammonia Ligase / Q000096:biosynthesis / Q000235:genetics*; D008969:Molecular Sequence Data; D030821:Plants, Genetically Modified / Q000235:genetics*; D010946:Plants, Medicinal; D010947:Plants, Toxic; D013025:Glycine max / Q000201:enzymology / Q000235:genetics*; D014026:Nicotiana / Q000235:genetics
D000641:Ammonia; D005974:Glutamate-Ammonia Ligase