1Cell-autonomous immunity is widespread in plant-fungus interactions and terminates fungal pathogenesis either at the cell surface or after pathogen entry. Although post-invasive resistance responses typically coincide with a self-contained cell death of plant cells undergoing attack by parasites, these cells survive pre-invasive defence. Mutational analysis in Arabidopsis identified PEN1 syntaxin as one component of two pre-invasive resistance pathways against ascomycete powdery mildew fungi 1-3 . Here we show that plasma-membrane-resident PEN1 promiscuously forms SDS-resistant soluble N-ethylmaleimide sensitive factor attachment protein receptor (SNARE) complexes together with the SNAP33 adaptor and a subset of vesicle-associated membrane proteins (VAMPs). PEN1-dependent disease resistance acts in vivo mainly through two functionally redundant VAMP72 subfamily members, VAMP721 and VAMP722. Unexpectedly, the same two VAMP proteins also operate redundantly in a default secretory pathway, suggesting dual functions in separate biological processes owing to evolutionary co-option of the default pathway for plant immunity. The disease resistance function of the secretory PEN1-SNAP33-VAMP721/722 complex and the pathogen-induced subcellular dynamics of its components are mechanistically reminiscent of immunological synapse formation in vertebrates, enabling execution of immune responses through focal secretion.Arabidopsis is immune to non-adapted powdery mildew fungi such as Blumeria graminis and Erysiphe pisi, which in nature colonize grass and pea species, respectively. This non-host resistance requires both pre-and post-invasive immune responses, which are under separate genetic control 2 . The former response engages PEN1 syntaxin, peroxisomal PEN2 b-glycosyl hydrolase and the plasmamembrane-resident PEN3 ABC transporter 1-3 . PEN2 and PEN3 act in the same pathway and are implicated in the cytoplasmic synthesis and transport of small antimicrobial compounds across the plasma membrane at attempted fungal entry sites, respectively 2,3 . PEN1 syntaxin acts in a second pathway and could, by analogy to known syntaxin functions in yeast and animals, either participate in vesicle fusion processes 4 or modulate ion-channel activity through interactions with plasma-membrane-resident ion channels 5 . Genetic studies defy mechanistic interpretations but suggest direct or indirect PEN1 repressor activity in defence responses that are dependent on salicylic acid, as well as an overlapping function with the closely related syntaxin of plant 122 (SYP122) 6 . Compared with largely resistant PEN1 wild type and severely defence-compromised pen1-1 null mutants, plants containing the pen1-3 allele allow intermediate B. graminis entry rates, indicating residual PEN1-3 resistance activity ( Supplementary Fig. 1a). In the deduced PEN1-3 protein, a glycine residue is substituted by a glutamate in the SNARE domain 1 (Supplementary Fig. 1b). Because this mutation affects a hydrophobic residue that is thought to stabilize interactions with ...

Plant innate immunity against invasive biotrophic pathogens depends on the intracellular defense regulator ENHANCED DISEASE SUSCEPTIBILITY1 (EDS1). We show here that Arabidopsis thaliana EDS1 interacts in vivo with another protein, SENESCENCE-ASSOCIATED GENE101 (SAG101), discovered through a proteomic approach to identify new EDS1 pathway components. Together with PHYTOALEXIN-DEFICIENT4 (PAD4), a known EDS1 interactor, SAG101 contributes intrinsic and indispensable signaling activity to EDS1-dependent resistance. The combined activities of SAG101 and PAD4 are necessary for programmed cell death triggered by the Toll-Interleukin-1 Receptor type of nucleotide binding/leucine-rich repeat immune receptor in response to avirulent pathogen isolates and in restricting the growth of normally virulent pathogens. We further demonstrate by a combination of cell fractionation, coimmunoprecipitation, and fluorescence resonance energy transfer experiments the existence of an EDS1-SAG101 complex inside the nucleus that is molecularly and spatially distinct from EDS1-PAD4 associations in the nucleus and cytoplasm. By contrast, EDS1 homomeric interactions were detected in the cytoplasm but not inside the nucleus. These data, combined with evidence for coregulation between individual EDS1 complexes, suggest that dynamic interactions of EDS1 and its signaling partners in multiple cell compartments are important for plant defense signal relay.