Pest and disease protection conferred by expression of barley �-hordothionin and Nicotiana alata proteinase inhibitor genes in transgenic tobacco

Abstract

Proteinase inhibitors and thionins are among the many proteins thought to have a role in plant defence against pests and pathogens. Complementary DNA clones encoding the precursors of a multi-domain proteinase inhibitor from Nicotiana alata Link et Otto (NA-PI) (Mr approximately 43 000) and a β-hordothionin (β-HTH) (Mr approximately 13 000) from barley, were linked to constitutive promoters and subsequently transferred by Agrobacterium-mediated transformation into tobacco. The NA-PI and β-HTH precursor proteins were synthesised and post-translationally processed in transgenic tobacco and accumulated as polypeptides of apparent size Mr approximately 6000 and Mr approximately 8500, respectively. The na-pi and β-hth genes were stably inherited for at least two generations. Transgenic tobacco plants containing the highest amounts of NA-PI and β-HTH were crossed to produce plants containing both genes. Helicoverpa armigera (tobacco budworm) larvae that ingested transgenic tobacco leaves expressing both NA-PI and β-HTH, exhibited higher mortality and slower development relative to larvae fed on non-transgenic tobacco. NA-PI and β-HTH, either alone, or in combination, also conferred protection against the fungal pathogen, Botrytis cinerea (grey mould) and the bacterial pathogen, Pseudomonas solanacearum (bacterial wilt). The effect of the two proteins depended upon the organism tested and the contribution of each gene to the protective effects was not necessarily equal. The genetic engineering of plants with proteinase inhibitors or thionins, therefore, has potential for improving crop productivity by simultaneously increasing resistance to both pests and pathogens.