insect peptide engineered into the tomato prosystemin gene is released in transgenic tobacco plants and exerts biological activity

• Published in: Plant molecular biology 2003-12, Vol.53 (6), p.891-902
• Main Authors: Tortiglione, C, Fogliano, V, Ferracane, R, Fanti, P, Pennacchio, F, Monti, L.M, Rao, R
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Amino acids; Animals; Chromatography, High Pressure Liquid; Gene Expression; Genes; Genetic engineering; Heliothis virescens; insect control; insect development; insect hormones; insect pests; insecticidal properties; insecticidal proteins; Insects; Larva - drug effects; Larva - growth & development; Larvae; leaves; Lycopersicon esculentum; Molecular Sequence Data; Moths - drug effects; Moths - growth & development; Nicotiana - genetics; Nicotiana - metabolism; Nicotiana tabacum; oligopeptides; Oligopeptides - analysis; Oligopeptides - genetics; peptide hormones; Peptide Hormones - analysis; Peptide Hormones - genetics; Peptides; Peptides - genetics; plant hormones; Plant Leaves - genetics; Plant Leaves - metabolism; Plant Proteins - genetics; Plants, Genetically Modified; recombinant DNA; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - metabolism; Recombinant Fusion Proteins - pharmacology; RNA, Messenger - genetics; RNA, Messenger - metabolism; Solanum lycopersicum var. lycopersicum; Spectrometry, Mass, Electrospray Ionization; synthetic genes; Tobacco; Tomatoes; transgenes; transgenic plants; trypsin-modulating oostatic factor
• ISSN: 0167-4412; 1573-5028
• DOI: 10.1023/B:PLAN.0000023667.62501.ef

Tomato systemin is a signalling peptide produced in response to wounding that locally and systemically activates several defence genes. The peptide is released from the C-terminus of prosystemin, the 200 amino acid precursor, following post-translational modifications involving unknown events and enzymes. In tobacco, two systemin molecules have been recently isolated, neither sharing any sequence homologies with the tomato prosystemin gene/protein, but performing similar functions. We modified the tomato prosystemin gene by replacing the systemin-encoding region with a synthetic sequence encoding TMOF (trypsin-modulating oostatic factor), a 10 amino acid insect peptide hormone toxic to Heliothis virescens larvae, and expressed the chimeric gene in tobacco. The results reported here show that transformed leaves contain the TMOF peptide and exert toxic activity against insect larvae reared on them. In addition, subcellular localization studies showed the cytoplasmic location of the released TMOF, suggesting that in tobacco the enzymes responsible for the post-translational modifications of the tomato precursor protein are present and act in the cytoplasm to recognise the modified prohormone. The molecular engineering of the precursor, beside supplying new clues towards the understanding of prosystemin processing, constitutes an useful tool for plant genetic manipulation, by enabling the delivery of short biological active peptides.