A strong constitutive positive element is essential for the ammonium-regulated expression of a soybean gene encoding cytosolic glutamine synthetase

• Published in: Plant molecular biology 1999-02, Vol.39 (3), p.551-564
• Main Authors: Terce-Laforgue, T. (Laboratoire du Metabolisme et de la Nutrition des Plantes INRA, Versailles (France).), Carrayol, E, Cren, M, Desbrosses, G, Hecht, V, Hirel, B
• Format: Article
• Language: English
• Subjects: ACTIVADOR GENICO; ACTIVATEUR GENIQUE; Agrobacterium rhizogenes; AMMONIA; AMMONIAC; Ammonium; AMONIACO; Bacteria; Base Sequence; Cytosol - enzymology; DNA, Plant - chemistry; DNA, Plant - genetics; EXPRESION GENICA; EXPRESSION DES GENES; Fabaceae - enzymology; Fabaceae - genetics; GENE ACTIVATORS; GENE EXPRESSION; Gene Expression - drug effects; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Plant; Genes, Plant - genetics; GLUTAMATE AMMONIA LIGASE; Glutamate-Ammonia Ligase - genetics; Glutamate-Ammonia Ligase - metabolism; GLUTAMINA SINTETASA; GLUTAMINE SYNTHETASE; GLYCINE MAX; Glycine max - chemistry; Glycine max - enzymology; Glycine max - genetics; LOTUS CORNICULATUS; Molecular Sequence Data; NODOSITE RACINAIRE; NUCLEOTIDE SEQUENCE; NUDOSIDADES RADICULARES; PLANTAS TRANSGENICAS; PLANTE TRANSGENIQUE; Plants, Genetically Modified; Plants, Medicinal; Promoter Regions, Genetic; Quaternary Ammonium Compounds - pharmacology; Regulatory Sequences, Nucleic Acid; ROOT NODULES; Roots; SECUENCIA NUCLEOTIDICA; Sequence Alignment; Sequence Analysis, DNA; Sequence Deletion; SEQUENCE NUCLEOTIDIQUE; Soybeans; TRANSGENIC PLANTS
• ISSN: 0167-4412; 1573-5028
• DOI: 10.1023/A:1006169018296

In order to identify important promoter elements controlling the ammonium-regulated expression of the soybean gene GS15 encoding cytosolic glutamine synthetase, a series of 5' promoter deletions were fused to the GUS reporter gene. To allow the detection of positive and negative regulatory elements, a series of 3' deletions were fused to a -90 CaMV 35S promoter fragment placed upstream of the GUS gene. Both types of construct were introduced into Lotus corniculatus plants and soybean roots via Agrobacterium rhizogenes-mediated transformation. Both spectrophotometric enzymatic analysis and histochemical localization of GUS activity in roots, root nodules and shoots of transgenic plants revealed that a strong constitutive positive element (SCPE) of 400 bp, located in the promoter distal region is indispensable for the ammonium-regulated expression of GS15. Interestingly, this SCPE was able to direct constitutive expression in both a legume and non-legume background to a level similar to that driven by the CaMV 35S full-length promoter. In addition, results showed that separate proximal elements, located in the first 727 bp relative to the transcription start site, are essential for root- and root nodule-specific expression. This proximal region contains an AAAGAT and two TATTTAT consensus sequences characteristic of nodulin or nodule-enhanced gene promoters. A putative silencer region containing the same TATTTAT consensus sequence was identified between the SCPE and the organ-specific elements. The presence of positive, negative and organ-specific elements together with the three TATTTAT consensus sequences within the promoter strongly suggest that these multiple promoter fragments act in a cooperative manner, depending on the spatial conformation of the DNA for trans-acting factor accessibility.