Discovery of functional NLRs using expression level, high-throughput transformation and large-scale phenotyping

Authors: BRABHAM, HELEN - Sainsbury Laboratory; HERNANDEZ-PINZON, INMACULADA - Sainsbury Laboratory; YANAGIHARA, CHIZU - Kaneka Corporation; ISHIKAWA, NORIKO - Kaneka Corporation; KOMORI, TOSHIYUKI - Kaneka Corporation; MATNY, OADI - University Of Minnesota; HUBBARD, AMELIA - National Institute Of Agricultural Botany (NIAB); WITEK, KAMIL - Sainsbury Laboratory; FEIST, ALEXIS - University Of Minnesota; NUMAZAWA, HIRONOBU - Japan Tobacco International; GREEN, PHON - Sainsbury Laboratory; DREISEITL, ANTONIN - Agricultural Research Institute Kromeríž, Ltd; TAKEMORI, NOAKI - Sainsbury Laboratory; KOMARI, TOSHIHIKO - Kaneka Corporation; FREEDMAN, ROGER - The Gatsby Charitable Foundation; STEFFENSON, BRIAN - University Of Minnesota; VAN ESSE, H. PETER - University Of East Anglia; Moscou, Matthew

Submitted to: Nature Plants
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/20/2025
Publication Date: 9/23/2025
Citation: Brabham, H.J., Hernandez-Pinzon, I., Yanagihara, C., Ishikawa, N., Komori, T., Matny, O.N., Hubbard, A., Witek, K., Feist, A., Numazawa, H., Green, P., Dreiseitl, A., Takemori, N., Komari, T., Freedman, R.P., Steffenson, B.J., Van Esse, H., Moscou, M.J. 2025. Discovery of functional NLRs using expression level, high-throughput transformation and large-scale phenotyping. Nature Plants. 15. Article 2110. https://doi.org/10.1038/s41477-025-02110-w.
DOI: https://doi.org/10.1038/s41477-025-02110-w

Interpretive Summary: In agriculture, integrated disease/pest management is built on three pillars: (1) agricultural practice (i.e. crop rotation, planting density, mixed cropping), (2) chemical control (e.g. pesticides and fungicides), and (3) genetic resistance. The use of genetic resistance requires that plant breeders continuously integrate diverse sources of resistance, as plant pathogens can break resistance in the plants. The process of identifying new disease resistance genes typically involves screening diverse domesticated or wild species, generating crosses with elite crop accessions, identifying the location of resistance in the genome, and lastly, breeding this resistance into elite accessions. To accelerate this pipeline, we leveraged a high-throughput transformation system with a signature of functional resistance genes. Using this approach, we have mined resistance genes from 69 accessions belonging to 18 Pooideae species and identified 19 stem rust resistance genes.

Technical Abstract: Breeding crop species that are safe from pests and diseases is vital to build the sustainable food systems crucial for food security. An effective and environmentally friendly method of disease control is to enhance the plant immune system by introducing functional resistance genes. A major class of plant immune receptors are nucleotide binding, leucine-rich repeat receptors (NLRs), however identifying functional NLRs for use in elite crop varieties is time-consuming and resource intensive. Through analyses of expression data, we identify high expression as an overlooked molecular signature of functional NLRs. Combining this signature with high-throughput crop transformation, we developed NLRseek™ which enables rapid identification and in planta validation of functional NLRs from non-domesticated germplasm. Using this approach, we double the amount of known NLRs that provide stem rust resistance in wheat, a critical threat to food security. The NLRseek™ pipeline facilitates rapid resistance gene discovery for use in crop species.