The nucleotide-binding domain and leucine-rich repeat (NLR) family of plant receptors detects pathogen-derived molecules, designated effectors, inside host cells and mediates innate immune responses to pathogenic invaders. Genetic evidence revealed species-specific coevolution of many NLRs with effectors from host-adapted pathogens, suggesting that the specificity of these NLRs is restricted to the host or closely related plant species. However, we report that an NLR immune receptor (MLA1) from monocotyledonous barley is fully functional in partially immunocompromised dicotyledonous Arabidopsis thaliana against the barley powdery mildew fungus, Blumeria graminis f. sp. hordei. This implies ∼200 million years of evolutionary conservation of the underlying immune mechanism. A time-course RNA-seq analysis in transgenic Arabidopsis lines detected sustained expression of a large MLA1-dependent gene cluster. This cluster is greatly enriched in genes known to respond to the fungal cell wallderived microbe-associated molecular pattern chitin. The MLA1-dependent sustained transcript accumulation could define a conserved function of the nuclear pool of MLA1 detected in barley and Arabidopsis. We also found that MLA1-triggered immunity was fully retained in mutant plants that are simultaneously depleted of ethylene, jasmonic acid, and salicylic acid signaling. This points to the existence of an evolutionarily conserved and phytohormone-independent MLA1-mediated resistance mechanism. This also suggests a conserved mechanism for internalization of B. graminis f. sp. hordei effectors into host cells of flowering plants. Furthermore, the deduced connectivity of the NLR to multiple branches of immune signaling pathways likely confers increased robustness against pathogen effector-mediated interception of host immune signaling and could have contributed to the evolutionary preservation of the immune mechanism.innate immunity | defense phytohormone T he nucleotide-binding domain and leucine-rich repeat (NLR)-containing proteins in plants and animals mediate pathogensensing. Unlike animal NLRs that detect mostly conserved microbial molecules, plant-encoded NLRs typically recognize the presence of isolate-specific pathogen effectors (1). Most plant NLRs are modular, containing coiled-coil (CC) or TOLL/IL-1 receptor domains at the N terminus, a central nucleotide-binding domain, and C-terminal leucine-rich repeats. Plant NLRs are often polymorphic between individual plants of a host population, and the sum of genes encoding NLRs in a host population defines the repertoire available for the detection of structure or action of polymorphic pathogen effectors (1).Interfamily transfer of NLR function was shown by coexpression of an NLR, its cognate effector and the effector target (e.g., Arabidopsis RPS5, Pseudomonas syringae AvrPphB, and Arabidopsis PBS1; reviewed in ref.2). However, these data are based on transient gene expression and use host cell death as proxy for NLR activity even though cell death has been uncoupled from NLRmedia...