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The endoplasmic reticulum (ER) is the entry point to the secretory pathway and, as such, is critical for adaptive responses to biotic stress, when the demand for de novo synthesis of immunity-related proteins and signalling components increases significantly. Successful phytopathogens have evolved an arsenal of small effector proteins which collectively reconfigure multiple host components and signalling pathways to promote virulence; a small, but important, subset of which are targeted to the endomembrane system including the ER. We identified and validated a conserved C-terminal tail-anchor motif in a set of pathogen effectors known to localize to the ER from the oomycetes Hyaloperonospora arabidopsidis and Plasmopara halstedii (downy mildew of Arabidopsis and sunflower, respectively) and used this protein topology to develop a bioinformatic pipeline to identify putative ER-localized effectors within the effectorome of the related oomycete, Phytophthora infestans, the causal agent of potato late blight. Many of the identified P. infestans tail-anchor effectors converged on ER-localized NAC transcription factors, indicating that this family is a critical host target for multiple pathogens.

MotivationA common class of behaviour encountered in the biological sciences involves branching and recombination. During branching, a statistical process bifurcates resulting in two or more potentially correlated processes that may under-go further branching; the contrary is true during recombination, where two or more statistical processes converge into one. A key objective is to identify the time of this bifurcation (branch time) from time series measurements e.g., comparing a control time series with a perturbed time series. Whilst statistical treatments for the two branch (control versus treatment) case exists, the ability to infer more complex branching structure from time series data remains open. Gaussian processes (GPs) represents an ideal framework for such analysis, allowing for nonlinear regression that includes a rigorous treatment of uncertainty. Currently, however, GP models only exist for two-branch systems. Here we highlight how arbitrarily complex branching processes can be built using the correct composition of covariance functions within a GP framework, thus outlining a general framework for the treatment of branching and recombination in the form of branch-recombinant Gaussian processes (B-RGPs). We first demonstrate the performance of B-RGPs compared to a variety of existing regression approaches, and demonstrate robustness to model misspecification. B-RGPs are then used to investigate the branching patterns ofArabidopsis thalianagene expression following inoculation with the hemibotrophic bacteria,Pseudomonas syringae DC3000, and a disarmed mutant strain, hrpA. By grouping genes according to the number of branches, we could naturally separate out genes involved in basal immune response from those subverted by the virulent strain, and show enrichment for targets of pathogen protein effectors. Finally, we identify two early branching genes WRKY11 and WRKY17, and showed that groups of genes that branched at similar times to WRKY11/17 were enriched for W-box binding motifs, and overrepresented for genes differentially expressed in WRKY11/17 knockouts, suggesting that branch time could be used for identifying direct and indirect binding targets of key transcription factors.Software is available from:https://github.com/cap76/BranchingGPs.

The Asp-His-His-Cys-Cys-rich domain-containing Protein S-Acyl Transferases (PATs) are multipass transmembrane proteins that catalyze S-acylation (commonly known as S-palmitoylation), the reversible posttranslational lipid modification of proteins. Palmitoylation enhances the hydrophobicity of proteins, contributes to their membrane association, and plays roles in protein trafficking and signaling. In Arabidopsis (Arabidopsis thaliana), there are at least 24 PATs; previous studies on two PATs established important roles in growth, development, and stress responses. In this study, we identified a, to our knowledge, novel PAT, AtPAT14, in Arabidopsis. Complementation studies in yeast (Saccharomyces cerevisiae) and Arabidopsis demonstrate that AtPAT14 possesses PAT enzyme activity. Disruption of AtPAT14 by T-DNA insertion resulted in an accelerated senescence phenotype. This coincided with increased transcript levels of some senescence-specific and pathogen-resistant marker genes. We show that early senescence of pat14 does not involve the signaling molecules jasmonic acid and abscisic acid, or autophagy, but associates with salicylic acid homeostasis and signaling. This strongly suggests that AtPAT14 plays a pivotal role in regulating senescence via salicylic acid pathways. Senescence is a complex process required for normal plant growth and development and requires the coordination of many genes and signaling pathways. However, precocious senescence results in loss of biomass and seed production. The negative regulation of leaf senescence by AtPAT14 in Arabidopsis highlights, to our knowledge for the first time, a specific role for palmitoylation in leaf senescence.

Plant resistance to necrotrophic fungi is regulated by a complex set of signaling pathways that includes those mediated by the hormones salicylic acid (SA), ethylene (ET), jasmonic acid (JA), and abscisic acid (ABA). The role of ABA in plant resistance remains controversial, as positive and negative regulatory functions have been described depending on the plant-pathogen interaction analyzed. Here, we show that ABA signaling negatively regulates Arabidopsis (Arabidopsis thaliana) resistance to the necrotrophic fungus Plectosphaerella cucumerina. Arabidopsis plants impaired in ABA biosynthesis, such as the aba1-6 mutant, or in ABA signaling, like the quadruple pyr/pyl mutant (pyr1pyl1pyl2pyl4), were more resistant to P. cucumerina than wild-type plants. In contrast, the hab1-1abi1-2abi2-2 mutant impaired in three phosphatases that negatively regulate ABA signaling displayed an enhanced susceptibility phenotype to this fungus. Comparative transcriptomic analyses of aba1-6 and wild-type plants revealed that the ABA pathway negatively regulates defense genes, many of which are controlled by the SA, JA, or ET pathway. In line with these data, we found that aba1-6 resistance to P. cucumerina was partially compromised when the SA, JA, or ET pathway was disrupted in this mutant. Additionally, in the aba1-6 plants, some genes encoding cell wall-related proteins were misregulated. Fourier transform infrared spectroscopy and biochemical analyses of cell walls from aba1-6 and wild-type plants revealed significant differences in their Fourier transform infrared spectratypes and uronic acid and cellulose contents. All these data suggest that ABA signaling has a complex function in Arabidopsis basal resistance, negatively regulating SA/JA/ET- mediated resistance to necrotrophic fungi. Fil: Sánchez Vallet, Andrea. Universidad Politécnica de Madrid; España Fil: López, Gemma. Universidad Politécnica de Madrid; España Fil: Ramos, Brisa. Universidad Politécnica de Madrid; España Fil: Delgado Cerezo, Magdalena. Universidad Politécnica de Madrid; España Fil: Riviere, Marie Pierre. Universidad Politécnica de Madrid; España Fil: Llorente, Francisco. Universidad Politécnica de Madrid; España Fil: Fernández, Paula Virginia. Universidad de Buenos Aires. Facultad de Agronomía. Departamento de Biología Aplicada y Alimentos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Miedes, Eva. Universidad Politécnica de Madrid; España Fil: Estevez, Jose Manuel. Universidad de Buenos Aires. Facultad de Agronomía. Departamento de Biología Aplicada y Alimentos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Grant, Murray. University of Exeter; Reino Unido Fil: Molina, Antonio. Universidad Politécnica de Madrid; España

Banana Xanthomonas wilt is a newly emerging disease that is currently threatening the livelihoods of millions of farmers in East Africa. The causative agent is Xanthomonas campestris pathovar musacearum (Xcm), but previous work suggests that this pathogen is much more closely related to species Xanthomonas vasicola than to X. campestris. We have generated draft genome sequences for a banana-pathogenic strain of Xcm isolated in Uganda and for a very closely related strain of X. vasicola pathovar vasculorum, originally isolated from sugarcane, that is nonpathogenic on banana. The draft sequences revealed overlapping but distinct repertoires of candidate virulence effectors in the two strains. Both strains encode homologues of the Pseudomonas syringae effectors HopW, HopAF1 and RipT from Ralstonia solanacearum. The banana-pathogenic and non-banana-pathogenic strains also differed with respect to lipopolysaccharide synthesis and type-IV pili, and in at least several thousand single-nucleotide polymorphisms in the core conserved genome. We found evidence of horizontal transfer between X. vasicola and very distantly related bacteria, including members of other divisions of the Proteobacteria. The availability of these draft genomes will be an invaluable tool for further studies aimed at understanding and combating this important disease.

Alterations in the proteome of Arabidopsis (Arabidopsis thaliana) leaves during responses to challenge by Pseudomonas syringae pv tomato DC3000 were analyzed using two-dimensional gel electrophoresis. Protein changes characteristic of the establishment of disease, basal resistance, and resistance-gene-mediated resistance were examined by comparing responses to DC3000, a hrp mutant, and DC3000 expressing avrRpm1, respectively. The abundance of each protein identified was compared with that of selected transcripts obtained from comparable GeneChip experiments. We report changes in three subcellular fractions: total soluble protein, chloroplast enriched, and mitochondria enriched over four time points (1.5–6 h after inoculation). In total, 73 differential spots representing 52 unique proteins were successfully identified. Many of the changes in protein spot density occurred before significant transcriptional reprogramming was evident between treatments. The high proportion of proteins represented by more than one spot indicated that many of the changes to the proteome can be attributed to posttranscriptional modifications. Proteins found to show significant change after bacterial challenge are representative of two main functional groups: defense-related antioxidants and metabolic enzymes. Significant changes to photosystem II and to components of the mitochondrial permeability transition were also identified. Rapid communication between organelles and regulation of primary metabolism through redox-mediated signaling are supported by our data.

The availability of potassium to the plant is highly variable, due to complex soil dynamics, which are strongly influenced by root-soil interactions. A low plant potassium status triggers expression of high affinity K+ transporters, up-regulates some K+ channels, and activates signalling cascades, some of which are similar to those involved in wounding and other stress responses. The molecules that signal low K+ status in plants include reactive oxygen species and phytohormones, such as auxin, ethylene and jasmonic acid. Apart from up-regulation of transport proteins and adjustment of metabolic processes, potassium deprivation triggers developmental responses in roots. All these acclimation strategies enable plants to survive and compete for nutrients in a dynamic environment with a variable availability of potassium.

The RPM1 protein confers resistance to Pseudomonas syringae pv tomato DC3000 expressing either of the Type III effector proteins AvrRpm1 or AvrB. Here, we describe the isolation and functional characterization of RPM1 Interacting Protein 13 (RIN13), a resistance protein interactor shown to positively enhance resistance function. Ectopic expression of RIN13 (RIN13s) enhanced bacterial restriction mechanisms but paradoxically abolished the normally rapid hypersensitive response (HR) controlled by RPM1. In contrast with wild-type plants, leaves expressing RIN13s did not undergo electrolyte leakage or accumulate H2O2 after bacterial delivery of AvrRpm1. Overexpression of RIN13 also altered the transcription profile observed during a normal HR. By contrast, RIN13 knockout plants had the same ion leakage signatures and HR timing of wild-type plants in response to DC3000(avrRpm1) but failed to suppress bacterial growth. The modified phenotypes seen in the RIN13s/as plants were specific to recognition of AvrRpm1 or AvrB, and wild-type responses were observed after challenge with other incompatible pathogens or the virulent DC3000 isolate. Our results suggest that cell death is not necessary to confer resistance, and engineering enhanced resistance without activation of programmed cell death is a real possibility.