Multiple functional self-association interfaces in plant TIR domains

The self-association of Toll/interleukin-1 receptor/resistance protein (TIR) domains has been implicated in signaling in plant and animal immunity receptors. Structure-based studies identified different TIR-domain dimerization interfaces required for signaling of the plant nucleotide-binding oligomerization domain-like receptors (NLRs) L6 from flax and disease resistance protein RPS4 from Arabidopsis. Here we show that the crystal structure of the TIR domain from the Arabidopsis NLR suppressor of npr1-1, constitutive 1 (SNC1) contains both an L6-like interface involving helices alpha D and alpha E (DE interface) and an RPS4-like interface involving helices alpha A and alpha E (AE interface). Mutations in either the AE- or DE-interface region disrupt cell-death signaling activity of SNC1, L6, and RPS4 TIR domains and full-length L6 and RPS4. Self-association of L6 and RPS4 TIR domains is affected by mutations in either region, whereas only AE-interface mutations affect SNC1 TIR-domain self-association. We further show two similar interfaces in the crystal structure of the TIR domain from the Arabidopsis NLR recognition of Peronospora parasitica 1 (RPP1). These data demonstrate that both the AE and DE self-association interfaces are simultaneously required for self-association and cell-death signaling in diverse plant NLRs.11139Ysciescopu

An Arabidopsis mutant with altered hypersensitive response to Xanthomonas campestris pv. campestris, hxc1, displays a complex pathophenotype

The hxc1 mutant was identified by screening an EMS (ethylmethane sulphonate) mutagenized population of Arabidopsis Col-0 plants for an altered hypersensitive response (HR), after spray inoculation with an HR-inducing isolate of Xanthomonas campestris pv. campestris (Xcc) (strain 147). The hxc1 mutant shows a susceptible phenotype several days after initiation of the interaction with the avirulent strain. This macroscopically observed phenotype was confirmed by measurement of in planta bacterial growth and by microscopical analysis. interestingly, the hxc1 mutation acts very specifically. Hxc1 displays a pathophenotype identical to that observed in the wild-type with several extensively characterized avirulent and virulent bacteria, except in response to Pseudomonas syringae pv. tomato strain DC3000/avrRpm1, for which a partial loss of resistance was observed. Finally, the mutation causes an attenuation of expression of several defence markers regulated through different signalling pathways. Together, these data underline the complexity of this novel defence mutant, and support the hypothesis of a mutation affecting a key component acting during the first steps of the plant defence response leading to resistance to Xcc147 and Pseudomonas syringae pv. tomato containing the avr gene, avrRpm1

A DHA14 drug efflux gene from Xanthomonas albilineans confers high-level albicidin antibiotic resistance in Escherichia coli

Aims: Identification of a gene for self-protection from the antibiotic-producing plant pathogen Xanthomonas albilineans, and functional testing by heterologous expression. Methods and Results: Albicidin antibiotics and phytotoxins are potent inhibitors of prokaryote DNA replication. A resistance gene (albF) isolated by shotgun cloning from the X. albilineans albicidin-biosynthesis region encodes a protein with typical features of DHA14 drug efflux pumps. Low-level expression of albF in Escherichia coli increased the MIC of albicidin 3000-fold, without affecting tsx-mediated albicidin uptake into the periplasm or resistance to other tested antibiotics. Bioinformatic analysis indicates more similarity to proteins involved in self-protection in polyketide-antibiotic-producing actinomycetes than to multi-drug resistance pumps in other Gram-negative bacteria. A complex promoter region may co-regulate albF with genes for hydrolases likely to be involved in albicidin activation or self-protection. Conclusions: AlbF is the first apparent single-component antibiotic-specific efflux pump from a Gram-negative antibiotic producer. It shows extraordinary efficiency as measured by resistance level conferred upon heterologous expression. Significance and Impact of the Study: Development of the clinical potential of albicidins as potent bactericidial antibiotics against diverse bacteria has been limited because of low yields in culture. Expression of albF with recently described albicidin-biosynthesis genes may enable large-scale production. Because albicidins are X. albilineans pathogenicity factors, interference with AlbF function is also an opportunity for control of the associated plant disease