BAK1 is required for the attenuation of ethylene-inducing xylanase (Eix)-induced defense responses by the decoy receptor LeEix1

Abstract

SUMMARYElicitor recognition plays a key role in the reaction of plants to pathogens and the induction of plant defense responses. Furthermore, plant-microbe interactions involve numerous regulatory systems essential for plant defense against pathogens. Ethylene-inducing xylanase (Eix) is a potent elicitor of plant defense responses in specific cultivars of tobacco (Nicotiana tabacum) and tomato (Solanum lycopersicum). The Eix receptors (LeEix1 and LeEix2) belong to a superclade of leucine-rich repeat receptor-like proteins (RLP) with a signal for receptor-mediated endocytosis, which was shown to be essential for proper induction of defense responses. Both receptors are able to bind Eix, while only LeEix2 mediates defense responses. Here we demonstrate that LeEix1 heterodimerizes with LeEix2 upon application of the Eix elicitor. We show that LeEix1 attenuates Eix-induced internalization and signaling of the LeEix2 receptor. Furthermore, we demonstrate, using yeast two-hybrid and in planta bimolecular fluorescence complementation assays, that the brassinosteroid co-receptor, BAK1, binds LeEix1 but not LeEix2. In BAK1-silenced plants, LeEix1 was no longer able to attenuate plant responses to Eix, indicating that BAK1 is required for this attenuation. We suggest that LeEix1 functions as a decoy receptor for LeEix2, a function which requires BAK1.

Cautionary Notes on the Use of C-Terminal BAK1 Fusion Proteins for Functional Studies

“…affect (in a nonmutually exclusive manner) phosphorylation events within the PRR-BAK1 complex, association and/or dissociation with downstream components, and phosphorylation events with direct substrates. It is becoming increasingly clear that, in addition to their role in BR responses, BAK1 and other SERKs in Arabidopsis and other plant species are master regulators of plant innate immunity (Heese et al, 2007;Shan et al, 2008;Fradin et al, 2009Fradin et al, , 2011Santos et al, 2009;Bar et al, 2010;Krol et al, 2010;Chaparro-Garcia et al, 2011;Mantelin et al, 2011;Roux et al, 2011;Yang et al, 2011) as well as of cell death control Kemmerling et al, 2007;Gao et al, 2009;Wang et al, 2011), pollen development (Albrecht et al, 2005;Colcombet et al, 2005), and organ abscission (Lewis et al, 2010). One must not assume that the model based on analyses between BRI1 and BAK1 is universal, and, in light of our results, it is essential that researchers analyze in detail and quantify the impact of modifying the SERK proteins in transgenic approaches when studying SERKdependent signaling pathways.…”

Clathrin-dependent endocytosis is required for immunity mediated by pattern recognition receptor kinases

“…Induced trafficking into late endosomes was also observed for the receptor-like proteins Cf-4 and LeEix2, which recognize fungal Avr4 and xylanase, respectively (12,13). While this depended on BAK1/SERK3 for Avr4-induced Cf-4 internalization (12), BAK1 was not required for xylanasetriggered endocytosis of LeEix2 (13). Thus, although internalization of PRRs (FLS2, LeEix2, and Cf-4) is found in plants, whether receptor-mediated endocytosis is a more general feature of activated PRRs and involves shared trafficking routes remains unclear.…”

“…This highlights a common pathway for the transport of activated plant receptors, supporting the hypothesis that all receptors are targeted to the same final destination, possibly the vacuole. This extends to the receptor-like protein PRRs Cf-4 and LeEix2, which reportedly localize to late endosomes (12,13), as well as to the BRI1 hormone receptor. It provides evidence that this pathway is common across different types of PRR families and receptor kinases, possibly to accumulate cargoes before delivery to the vacuole.…”

“…This induces trafficking of FLS2 into the late endosomal pathway and is associated with receptor degradation (10,11). Induced trafficking into late endosomes was also observed for the receptor-like proteins Cf-4 and LeEix2, which recognize fungal Avr4 and xylanase, respectively (12,13). While this depended on BAK1/SERK3 for Avr4-induced Cf-4 internalization (12), BAK1 was not required for xylanasetriggered endocytosis of LeEix2 (13).…”

Fungal Endopolygalacturonases Are Recognized as Microbe-Associated Molecular Patterns by the Arabidopsis Receptor-Like Protein RESPONSIVENESS TO BOTRYTIS POLYGALACTURONASES1

“…Although it was originally identified as a protein that interacts with the brassinosteroid receptor BRI1 Nam and Li, 2002), BAK1 also forms ligandinduced complexes with FLS2 and EFR and contributes to disease resistance against the pathogens Pseudomonas syringae, Hyaloperonospora arabidopsidis (Hpa), and Phytophthora infestans (Chinchilla et al, 2007;Heese et al, 2007;Chaparro-Garcia et al, 2011;Roux et al, 2011). Tomato BAK1 interacts in a ligand-independent manner with LeEIX1 but not with LeEIX2, and the BAK1-LeEIX1 interaction is required for the ability of LeEIX1 to attenuate the signaling of LeEIX2 (Bar et al, 2010). BAK1 has also been shown to interact with another LRR-RLK, BAK1-INTERACTING RECEPTOR-LIKE KINASE1 (BIR1).…”