Characterization of an auxin-inducible 1-aminocyclopropane-1-carboxylate synthase gene, VR-ACS6, of mungbean (Vigna radiata (L.) Wilczek) and hormonal interactions on the promoter activity in transgenie tobacco

A genomic clone for VR-A CS6, an isozyme of auxin-inducible ACC synthase of mungbean, was isolated, and its promoter activity was examined in transgenic tobacco. The clone contained 1,612 bp long 5' untranscribed region and its coding sequence consisted of three exons and two introns. Genomic Southern hybridization indicated that VRA CS6 is a single copy gene. The transcription initiation site was a cytosine present at 231-base upstream the translation start codon. The VR-ACS6 promoter contained DNA sequences homologous to various functionally identified auxin-responsive elements. To demonstrate hormonal response of the promoter region, transgenic tobacco plants carrying the 1,719 bp VR-ACS6 promoter/-glucuronidase (GUS) fusion gene were generated. Strong GUS expression occurred by auxin treatment of leaves of T₀ transformants and hypocotyls of T₁ etiolated seedlings. Magnitude of the response to auxin was dose-dependent, and the increased GUS activity was detected at 0.1 μM and higher concentrations of IAA. Other plant hormones did not induce GUS activity, but greatly modified the response to auxin. Cytokinin enhanced the IAA-induced expression of GUS reporter gene, whereas ABA and ethylene suppressed the expression. These characteristics of VR-ACS6 promoter activity in transgenic tobacco are in good accordance with the expression patterns of the gene in mungbean hypocotyls. Histochemical staining showed that GUS activity was evident in both etiolated and light grown seedlings treated with IAA. Cytokinin enhanced the intensity of auxin-induced GUS stain and also expanded the stained area, whereas ABA and ethylene reduced both intensity and area of the stain.