A lineage-specific Exo70 is required for receptor kinase–mediated immunity in barley

In the evolution of land plants, the plant immune system has experienced expansion in immune receptor and signaling pathways. Lineage-specific expansions have been observed in diverse gene families that are potentially involved in immunity but lack causal association. Here, we show that Rps8-mediated resistance in barley to the pathogen Puccinia striiformis f. sp. tritici (wheat stripe rust) is conferred by a genetic module: Pur1 and Exo70FX12, which are together necessary and sufficient. Pur1 encodes a leucine-rich repeat receptor kinase and is the ortholog of rice Xa21, and Exo70FX12 belongs to the Poales-specific Exo70FX clade. The Exo70FX clade emerged after the divergence of the Bromeliaceae and Poaceae and comprises from 2 to 75 members in sequenced grasses. These results demonstrate the requirement of a lineage-specific Exo70FX12 in Pur1-mediated immunity and suggest that the Exo70FX clade may have evolved a specialized role in receptor kinase signaling

Using CRISPR/Cas9 genome editing in tomato to create a gibberellin-responsive dominant dwarf DELLA allele

The tomato PROCERA gene encodes a DELLA protein, and loss-of-function mutations derepress growth. We used CRISPR/Cas9 and a single guide RNAs (sgRNA) to target mutations to the PROCERA DELLA domain, and recovered several loss-of-function mutations and a dominant dwarf mutation that carries a deletion of one amino acid in the DELLA domain. This is the first report of a dominant dwarf PROCERA allele. This allele retains partial responsiveness to exogenously applied gibberellin. Heterozygotes show an intermediate phenotype at the seedling stage, but adult heterozygotes are as dwarfed as homozygotes

Extracellular proteolytic cascade in tomato activates immune protease Rcr3

Proteolytic cascades regulate immunity and development in animals, but these cascades in plants have not yet been reported. Here we report that the extracellular immune protease Rcr3 of tomato is activated by P69B and other subtilases (SBTs), revealing a proteolytic cascade regulating extracellular immunity in solanaceous plants. Rcr3 is a secreted papain-like Cys protease (PLCP) of tomato that acts both in basal resistance against late blight disease (Phytophthora infestans) and in gene-for-gene resistance against the fungal pathogen Cladosporium fulvum (syn. Passalora fulva) Despite the prevalent model that Rcr3-like proteases can activate themselves at low pH, we found that catalytically inactive proRcr3 mutant precursors are still processed into mature mRcr3 isoforms. ProRcr3 is processed by secreted P69B and other Asp-selective SBTs in solanaceous plants, providing robust immunity through SBT redundancy. The apoplastic effector EPI1 of P. infestans can block Rcr3 activation by inhibiting SBTs, suggesting that this effector promotes virulence indirectly by preventing the activation of Rcr3(-like) immune proteases. Rcr3 activation in Nicotiana benthamiana requires a SBT from a different subfamily, indicating that extracellular proteolytic cascades have evolved convergently in solanaceous plants or are very ancient in the plant kingdom. The frequent incidence of Asp residues in the cleavage region of Rcr3-like proteases in solanaceous plants indicates that activation of immune proteases by SBTs is a general mechanism, illuminating a proteolytic cascade that provides robust apoplastic immunity

The barley immune receptor Mla recognizes multiple pathogens and contributes to host range dynamics.

Funder: Gatsby Charitable Foundation; doi: https://doi.org/10.13039/501100000324Crop losses caused by plant pathogens are a primary threat to stable food production. Stripe rust (Puccinia striiformis) is a fungal pathogen of cereal crops that causes significant, persistent yield loss. Stripe rust exhibits host species specificity, with lineages that have adapted to infect wheat and barley. While wheat stripe rust and barley stripe rust are commonly restricted to their corresponding hosts, the genes underlying this host specificity remain unknown. Here, we show that three resistance genes, Rps6, Rps7, and Rps8, contribute to immunity in barley to wheat stripe rust. Rps7 cosegregates with barley powdery mildew resistance at the Mla locus. Using transgenic complementation of different Mla alleles, we confirm allele-specific recognition of wheat stripe rust by Mla. Our results show that major resistance genes contribute to the host species specificity of wheat stripe rust on barley and that a shared genetic architecture underlies resistance to the adapted pathogen barley powdery mildew and non-adapted pathogen wheat stripe rust

Barley MLA3 recognizes the host-specificity effector Pwl2 from Magnaporthe oryzae

Plant nucleotide-binding leucine-rich repeat (NLRs) immune receptors directly or indirectly recognize pathogen-secreted effector molecules to initiate plant defense. Recognition of multiple pathogens by a single NLR is rare and usually occurs via monitoring for changes to host proteins; few characterized NLRs have been shown to recognize multiple effectors. The barley (Hordeum vulgare) NLR gene Mildew locus a (Mla) has undergone functional diversification, and the proteins encoded by different Mla alleles recognize host-adapted isolates of barley powdery mildew (Blumeria graminis f. sp. hordei [Bgh]). Here, we show that Mla3 also confers resistance to the rice blast fungus Magnaporthe oryzae in a dosage-dependent manner. Using a forward genetic screen, we discovered that the recognized effector from M. oryzae is Pathogenicity toward Weeping Lovegrass 2 (Pwl2), a host range determinant factor that prevents M. oryzae from infecting weeping lovegrass (Eragrostis curvula). Mla3 has therefore convergently evolved the capacity to recognize effectors from diverse pathogens

Solanum americanum genome-assisted discovery of immune receptors that detect potato late blight pathogen effectors

Potato (Solanum tuberosum) and tomato (Solanum lycopersicon) crops suffer severe losses to late blight caused by the oomycete pathogen Phytophthora infestans. Solanum americanum, a relative of potato and tomato, is globally distributed and most accessions are highly blight resistant. We generated high-quality reference genomes of four S. americanum accessions, resequenced 52 accessions, and defined a pan-NLRome of S. americanum immune receptor genes. We further screened for variation in recognition of 315P. infestans RXLR effectors in 52 S. americanum accessions. Using these genomic and phenotypic data, we cloned three NLR-encoding genes, Rpi-amr4, R02860 and R04373, that recognize cognate P. infestans RXLR effectors PITG_22825 (AVRamr4), PITG_02860 and PITG_04373. These genomic resources and methodologies will support efforts to engineer potatoes with durable late blight resistance and can be applied to diseases of other crops

Domain swapping and gene shuffling identify sequences required for induction of an Avr-dependent hypersensitive response by the tomato Cf-4 and Cf-9 proteins

The tomato Cf-4 and Cf-9 genes confer resistance to infection by the biotrophic leaf mold pathogen Cladosporium. Their protein products induce a hypersensitive response (HR) upon recognition of the fungus-encoded Avr4 and Avr9 peptides. Cf-4 and Cf-9 share >91% sequence identity and are distinguished by sequences in their N-terminal domains A and B, their N-terminal leucine-rich repeats (LRRs) in domain C1, and their LRR copy number (25 and 27 LRRs, respectively). Analysis of Cf-4/Cf-9 chimeras, using several different bioassays, has identified sequences in Cf-4 and Cf-9 that are required for the Avr-dependent HR in tobacco and tomato. A 10–amino acid deletion within Cf-4 domain B relative to Cf-9 was required for full Avr4-dependent induction of an HR in most chimeras analyzed. Additional sequences required for Cf-4 function are located in LRRs 11 and 12, a region that contains only eight of the 67 amino acids that distinguish it from Cf-9. One chimera, with 25 LRRs that retained LRR 11 of Cf-4, induced an attenuated Avr4-dependent HR. The substitution of Cf-9 N-terminal LRRs 1 to 9 with the corresponding sequences from Cf-4 resulted in attenuation of the Avr9-induced HR, as did substitution of amino acid A433 in LRR 15. The amino acids L457 and K511 in Cf-9 LRRs 16 and 18 are essential for induction of the Avr9-dependent HR. Therefore, important sequence determinants of Cf-9 function are located in LRRs 10 to 18. This region contains 15 of the 67 amino acids that distinguish it from Cf-4, in addition to two extra LRRs. Our results demonstrate that sequence variation within the central LRRs of domain C1 and variation in LRR copy number in Cf-4 and Cf-9 play a major role in determining recognition specificity in these proteins