Pseudomonas syringae pv. actinidiae Type III Effectors Localized at Multiple Cellular Compartments Activate or Suppress Innate Immune Responses in Nicotiana benthamiana

Bacterial phytopathogen type III secreted (T3S) effectors have been strongly implicated in altering the interaction of pathogens with host plants. Therefore, it is useful to characterize the whole effector repertoire of a pathogen to understand the interplay of effectors in plants. Pseudomonas syringae pv. actinidiae is a causal agent of kiwifruit canker disease. In this study, we generated an Agrobacterium-mediated transient expression library of YFP-tagged T3S effectors from two strains of Psa, Psa-NZ V13 and Psa-NZ LV5, in order to gain insight into their mode of action in Nicotiana tabacum and N. benthamiana. Determining the subcellular localization of effectors gives an indication of the possible host targets of effectors. A confocal microscopy assay detecting YF-tagged Psa effectors revealed that the nucleus, cytoplasm and cell periphery are major targets of Psa effectors. Agrobacterium-mediated transient expression of multiple Psa effectors induced HR-like cell death (HCD) in Nicotiana spp., suggesting that multiple Psa effectors may be recognized by Nicotiana spp.. Virus-induced gene silencing (VIGS) of several known plant immune regulators, EDS1, NDR1, or SGT1 specified the requirement of SGT1 in HCD induced by several Psa effectors in N. benthamiana. In addition, the suppression activity of Psa effectors on HCD-inducing proteins and PTI was assessed. Psa effectors showed differential suppression activities on each HCD inducer or PTI. Taken together, our Psa effector repertoire analysis highlights the great diversity of T3S effector functions in planta.112Ysciescopu

Effector-assisted breeding for bacterial wilt resistance in horticultural crops

Ralstonia solanacearum (Rso) is a causal agent of bacterial wilt disease in a wide range of horticultural crops. Rso strains are heterogeneous in nature and are therefore difficult in terms of both classification and developmentof disease resistance. Rso pathogen-associated molecular patterns (PAMPs) and effector proteins are secreted into plant cells, where they respectively activate and suppress plant immunity, thereby affecting Rso virulence. We review the current knowledge of Rso disease resistance and efforts to generate Rso-resistant crop plants. Further, we propose the introduction into plants of known pattern recognition receptors (PRRs) that recognize Rso PAMPs in order to confer resistance to a large number of strains. Additionally, the conserved ‘core’ effectors from Rso phylotypes could be used to identify and deploy nucleotide-binding leucine-rich repeat (NLR) resistance genes in a desired crop cultivar. We suggest that a phylotype-specific effector-assisted breeding program be instituted to rapidly identify disease resistance genes in available plant germplasm collections. Furthermore, stacking multiple NLRs that recognize Rso effectors would provide durable disease resistance by minimizing the chance for Rso to evade the implemented resistance. Finally, we propose that this strategy would most efficiently be achieved through development of transgenic crop lines.1120Nsciescopuskc

Differential Suppression of Nicotiana benthamiana Innate Immune Responses by Transiently Expressed Pseudomonas syringae Type III Effectors

The plant pathogen Pseudomonas syringae injects about 30 different virulence proteins, so-called effectors, via a type III secretion system into plant cells to promote disease. Although some of these effectors are known to suppress either pattern-triggered immunity (PTI) or effector-triggered immunity (ETI), the mode of action of most of them remains unknown. Here, we used transient expression in Nicotiana benthamiana, to test the abilities of type III effectors of Pseudomonas syringae pv. tomato (Pto) DC3000 and Pseudomonas syringae pv. tabaci (Pta) 11528 to interfere with plant immunity. We monitored the sequential and rapid bursts of cytoplasmic Ca2+ and reactive oxygen species (ROS), the subsequent induction of defense gene expression, and promotion of cell death. We found that several effector proteins caused cell death, but independently of the known plant immune regulator NbSGT1, a gene essential for ETI. Furthermore, many effectors delayed or blocked the cell death-promoting activity of other effectors, thereby potentially contributing to pathogenesis. Secondly, a large number of effectors were able to suppress PAMP-induced defense responses. In the majority of cases, this resulted in suppression of all studied PAMP responses, suggesting that these effectors target common elements of PTI. However, effectors also targeted different steps within defense pathways and could be divided into three major groups based on their suppressive activities. Finally, the abilities of effectors of both Pto DC3000 and Pta 11528 to suppress plant immunity was conserved in most but not all cases. Overall, our data present a comprehensive picture of the mode of action of these effectors and indicate that most of them suppress plant defenses in various ways.This work was funded by the Gatsby Charitable Foundation and the Spanish Ministry of Science and Innovation Grant BIO2014- 55884-JIN to SG-I. We acknowledge support of the publication fee by the CSIC Open Access Publication Support Initiative through its Unit of Information Resources for Research (URICI)

Host adaptation and microbial competition drive Ralstonia solanacearum phylotype I evolution in the Republic of Korea

Bacterial wilt caused by the Ralstonia solanacearum species complex (RSSC) threatens the cultivation of important crops worldwide. We sequenced 30 RSSC phylotype I (R. pseudosolanacearum) strains isolated from pepper (Capsicum annuum) and tomato (Solanum lycopersicum) across the Republic of Korea. These isolates span the diversity of phylotype I, have extensive effector repertoires and are subject to frequent recombination. Recombination hotspots among South Korean phylotype I isolates include multiple predicted contact-dependent inhibition loci, suggesting that microbial competition plays a significant role in Ralstonia evolution. Rapid diversification of secreted effectors presents challenges for the development of disease-resistant plant varieties. We identified potential targets for disease resistance breeding by testing for allele-specific host recognition of T3Es present among South Korean phyloype I isolates. The integration of pathogen population genomics and molecular plant pathology contributes to the development of location-specific disease control and development of plant cultivars with durable resistance to relevant threats.11Ysciescopu

Structural Basis for Assembly and Function of a Heterodimeric Plant Immune Receptor

Cytoplasmic plant immune receptors recognize specific pathogen effector proteins and initiate effector-triggered immunity. In Arabidopsis, the immune receptors RPS4 and RRS1 are both required to activate defense to three different pathogens. We show that RPS4 and RRS1 physically associate. Crystal structures of the N-terminal Toll–interleukin-1 receptor/resistance (TIR) domains of RPS4 and RRS1, individually and as a heterodimeric complex (respectively at 2.05, 1.75, and 2.65 angstrom resolution), reveal a conserved TIR/TIR interaction interface. We show that TIR domain heterodimerization is required to form a functional RRS1/RPS4 effector recognition complex. The RPS4 TIR domain activates effector-independent defense, which is inhibited by the RRS1 TIR domain through the heterodimerization interface. Thus, RPS4 and RRS1 function as a receptor complex in which the two components play distinct roles in recognition and signaling