Stepwise artificial evolution of a plant disease resistance gene

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 110; no. 52; pp. 21189 - 21194
• Main Authors: Harris, C. Jake, Slootweg, Erik J., Goverse, Aska, Baulcombe, David C.
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences 24.12.2013; NATIONAL ACADEMY OF SCIENCES; National Acad Sciences
• Subjects: Agriculture - methods; Agrobacterium tumefaciens; Amino Acid Substitution - genetics; Amino acids; arabidopsis-thaliana; Binding sites; Biological Sciences; Blotting, Western; Capsid Proteins - genetics; Carlavirus - genetics; cell-death; coiled-coil; Disease resistance; Evolution; Gene expression; Genes, vpr - genetics; Genetic Engineering - methods; Genetic mutation; hypersensitive response; immune receptors; Immunity, Innate - genetics; Leucine-Rich Repeat Proteins; mediated resistance; Mutagenesis; Mutation; nb-lrr protein; Necrosis; Nicotiana - genetics; Nicotiana - immunology; Nicotiana - virology; Nucleotides; pathogen effectors; Pathogens; Plant Proteins - genetics; Plant Proteins - immunology; Plants; Plants, Genetically Modified; Poplar mosaic virus; Potato virus X; Proteins; Proteins - genetics; Proteins - immunology; Reverse Transcriptase Polymerase Chain Reaction; rich repeat domains; Solanum tuberosum; tobacco-mosaic-virus; Transgenic plants; arms race; plant defense; genetically modified; NLR; plant immunity
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.1311134110

Genes encoding plant nucleotide-binding leucine-rich repeat (NB-LRR) proteins confer dominant resistance to diverse pathogens. The wild-type potato NB-LRR protein Rx confers resistance against a single strain of potato virus X (PVX), whereas LRR mutants protect against both a second PVX strain and the distantly related poplar mosaic virus (PopMV). In one of the Rx mutants there was a cost to the broad-spectrum resistance because the response to PopMV was transformed from a mild disease on plants carrying wild-type Rx to a trailing necrosis that killed the plant. To explore the use of secondary mutagenesis to eliminate this cost of broad-spectrum resistance, we performed random mutagenesis of the N-terminal domains of this broad-recognition version of Rx and isolated four mutants with a stronger response against the PopMV coat protein due to enhanced activation sensitivity. These mutations are located close to the nucleotide-binding pocket, a highly conserved structure that likely controls the “switch” between active and inactive NB-LRR conformations. Stable transgenic plants expressing one of these versions of Rx are resistant to the strains of PVX and the PopMV that previously caused trailing necrosis. We conclude from this work that artificial evolution of NB-LRR disease resistance genes in crops can be enhanced by modification of both activation and recognition phases, to both accentuate the positive and eliminate the negative aspects of disease resistance.