Functional diversification of a wild potato immune receptor at its center of origin.

Plant cell surface pattern recognition receptors (PRRs) and intracellular immune receptors cooperate to provide immunity to microbial infection. Both receptor families have coevolved at an accelerated rate, but the evolution and diversification of PRRs is poorly understood. We have isolated potato surface receptor Pep-13 receptor unit (PERU) that senses Pep-13, a conserved immunogenic peptide pattern from plant pathogenic Phytophthora species. PERU, a leucine-rich repeat receptor kinase, is a bona fide PRR that binds Pep-13 and enhances immunity to Phytophthora infestans infection. Diversification in ligand binding specificities of PERU can be traced to sympatric wild tuber-bearing Solanum populations in the Central Andes. Our study reveals the evolution of cell surface immune receptor alleles in wild potato populations that recognize ligand variants not recognized by others.

Recognition of Pep-13/25 MAMPs of Phytophthora localizes to an RLK locus in Solanum microdontum.

Pattern-triggered immunity (PTI) in plants is mediated by cell surface-localized pattern recognition receptors (PRRs) upon perception of microbe-associated molecular pattern (MAMPs). MAMPs are conserved molecules across microbe species, or even kingdoms, and PRRs can confer broad-spectrum disease resistance. Pep-13/25 are well-characterized MAMPs in Phytophthora species, which are renowned devastating oomycete pathogens of potato and other plants, and for which genetic resistance is highly wanted. Pep-13/25 are derived from a 42 kDa transglutaminase GP42, but their cognate PRR has remained unknown. Here, we genetically mapped a novel surface immune receptor that recognizes Pep-25. By using effectoromics screening, we characterized the recognition spectrum of Pep-13/25 in diverse Solanaceae species. Response to Pep-13/25 was predominantly found in potato and related wild tuber-bearing Solanum species. Bulk-segregant RNA sequencing (BSR-Seq) and genetic mapping the response to Pep-25 led to a 0.081 cM region on the top of chromosome 3 in the wild potato species Solanum microdontum subsp. gigantophyllum. Some BAC clones in this region were isolated and sequenced, and we found the Pep-25 receptor locates in a complex receptor-like kinase (RLK) locus. This study is an important step toward the identification of the Pep-13/25 receptor, which can potentially lead to broad application in potato and various other hosts of Phytophthora species.

Identification of Solanum Immune Receptors by Bulked Segregant RNA-Seq and High-Throughput Recombinant Screening.

The identification, understanding, and deployment of immune receptors are crucial to achieve high-level and durable resistance for crops against pathogens. In potato, many R genes have been identified using map-based cloning strategies. However, this is a challenging and laborious task that involves the development of a high number of molecular markers for the initial mapping, and the screening of thousands of plants for fine mapping. Bulked segregant RNA-Seq (BSR-Seq) has proven to be an efficient technique for the mapping of resistance genes. The RNA from two bulks of plants with contrasting phenotypes is sequenced and analyzed to identify single-nucleotide polymorphism (SNPs) markers linked to the target gene. Subsequently, the SNP markers that are identified can be used to delimit the mapping interval. Additionally, we designed an in vitro recombinant screening strategy that is advantageous for analyzing a large number of plants, in terms of time, space, and cost. Tips and detailed protocols, including BSR-Seq, bioinformatic analysis, and recombinant screening, are provided in this chapter.