An XA21-Associated Kinase (OsSERK2) Regulates Immunity Mediated by the XA21 and XA3 Immune Receptors

XueweiChen 0 1 ShiminZuo 0 1 BenjaminSchwessinger 0 1 MawshengChern 0 1 Patrick E.Canlas 0 1 DelingRuan 0 1 XiaogangZhou 0 1 JingWang 0 1 ArsalanDaudi 0 1 Christopher J.Petzold 0 1 Joshua L.Heazlewood 0 1 Pamela C.Ronald 0 1 0 a Department of Plant Pathology and the Genome Center, University of California , Davis, CA 95616 , USA b Rice Research Institute, Sichuan Agricultural University , Chengdu, Sichuan 611130, China c Joint Bioenergy Institute , Emeryville, CA 94710 , USA d Key Laboratory of Crop Genetics and Physiology of Jiangsu Province, Key Laboratory of Plant Functional Genomics of the Ministry of Education, College of Agriculture, Yangzhou University , Yangzhou 225009, China 1 Chen et al. OsSERK2 Regulates Receptor Kinase-Mediated Immunity in Rice The rice XA21 immune receptor kinase and the structurally related XA3 receptor confer immunity to Xanthomonas oryzae pv. oryzae (Xoo), the causal agent of bacterial leaf blight. Here we report the isolation of OsSERK2 (rice somatic embryogenesis receptor kinase 2)and demonstrate that OsSERK2 positively regulates immunity mediated by XA21 and XA3 as well as the rice immune receptor FLS2 (OsFLS2). Rice plants silenced for OsSerk2 display altered morphology and reduced sensitivity to the hormone brassinolide. OsSERK2 interacts with the intracellular domains of each immune receptor in the yeast two-hybrid system in a kinase activity-dependent manner. OsSERK2 undergoes bidirectional transphosphorylation with XA21 in vitro and forms a constitutive complex with XA21 in vivo. These results demonstrate an essential role for OsSERK2 in the function of three rice immune receptors and suggest that direct interaction with the rice immune receptors is critical for their function. Taken together, our findings suggest that the mechanism of OsSERK2-meditated regulation of rice XA21, XA3, and FLS2 differs from that of AtSERK3/BAK1-mediated regulation of Arabidopsis FLS2 and EFR. - In TRODu CTIOn The XA21 receptor kinase confers broad-spectrum resistance to Xanthomonas oryzae pv. oryzae ( Xoo) (Song et al., 1995). Animals and other plant species also carry membraneanchored receptors with striking structural similarities to XA21 (Ronald and Beutler, 2010). Many of these receptors play key roles in recognition of conserved microbial signatures (also called pathogen-associated molecular patterns (PAMPs)) and host defense (Song et al., 1995; Lemaitre et al., 1996; Medzhitov etal., 1997; Poltorak etal., 1998; Gomez-Gomez and Boller, 2000; Zipfel et al., 2006; Ronald and Beutler, 2010). XA21 and structurally similar immune receptors activate defense signaling via membrane-associated complexes that include non-RD (arginine-aspartic acid) kinases to induce a core set of defense responses (Ronald and Beutler, 2010; Dardick etal., 2012). The non-RD kinases are either associated with the receptor via adaptor proteins (animals) or integral to the receptor (plants) (Ronald and Beutler, 2010; Schwessinger and Ronald, 2012). In rice, the immune receptors XA21, XA3, Pid2, and FLS2 all belong to the non-RD subclass of kinases (Dardick etal., 2012; Schwessinger and Ronald, 2012). In contrast to non-RD kinases, which are associated with the immune response, most RD kinases appear to regulate non-immune responses or serve as co-regulators of receptor kinase-mediated immunity (Chinchilla etal., 2006; Heese et al., 2007; Roux et al., 2011; Schwessinger et al., 2011; Schwessinger and Ronald, 2012) with the notable exception of the RD-kinase CERK1 in Arabidopsis, which directly binds chitin (Liu etal., 2012b). In Arabidopsis, members of the somatic embryogenesis receptor kinase (SERK) regulate the function of multiple plasma-membrane-localized receptor kinases (RKs) including hormone receptors and immune RKs (Chinchilla et al., 2009; Li, 2010), and are members of the RD subclass of kinases (Schwessinger and Ronald, 2012). The best-studied member SERK3 is also referred to as BAK1 (brassinosteroidinsensitive 1 (BRI1) associated kinase 1), as it was initially identified as a key regulator of BRI1-mediated signaling (Li etal., 2002; Nam and Li, 2002). BRI1 is the main receptor of brassinosteroids (BR), an important class of plant hormones regulating growth and development (Li and Chory, 1997; Clouse, 2011). SERK3 and its closest paralog SERK4 are critical co-regulators of the immune response triggered by the ligand-activated Arabidopsis immune receptor kinase FLS2 (flagellin insensitive 2), EFR (EF-TU receptor), and PEPR1/2 (PEP receptors 1 and 2)(Chinchilla etal., 2006; Heese etal., 2007; Postel etal., 2009; Krol etal., 2010; Roux etal., 2011; Schwessinger et al., 2011). The pattern-recognition receptors (PRRs) FLS2 and EFR recognize the bacterial proteins (or derived epitopes) flagellin (flg22) or EF-TU (elf18), respectively (Zipfel et al., 2006; Chinchilla et al., 2007). In contrast, PEPR1/2 are paralogous receptors for the endogenously produced small danger-associated peptides, AtPeps (Yamaguchi etal., 2006, 2010). Arabidopsis FLS2 and EFR do not constitutively interact with SERK3 (or any other SERKfamily member) (Chinchilla et al., 2007; Heese et al., 2007; Schulze et al., 2010; Roux et al., 2011; Schwessinger et al., 2011). Only upon ligand binding, FLS2 and EFR undergo a nearly instantaneous complex formation with SERK3 and potentially with additional co-regulatory RKs (Chinchilla etal., 2009; Schulze etal., 2010; Roux etal., 2011). The FLS2/ EFRSERK3 complex formation is independent of the kinase activity of either interaction partner or any other associated kinase (Schulze etal., 2010; Schwessinger etal., 2011). Indeed, the co-crystal structure of the FLS2SERK3 ectodomains and flg22 suggests that flg22 acts as molecular glue by stabilizing the interaction between both receptors (Sun et al., 2013). This ligand-induced heterodimer formation is the molecular switch-on for transmembrane signaling of these Arabidopsis RKs (Albert etal., 2013). The tight association of the intracellular kinase domains is hypothesized to induce downstream signaling activation via specific structurally guided auto- and transphosphorylation events. SERK3 and FLS2/EFR undergo unidirectional phosphorylation in vitrothat is, SERK3 is able to transphosphorylate FLS2 or EFR but not vice versa (Schwessinger etal., 2011). In rice, XA21 confers robust resistance to Xoo (Song etal., 1995). XA21 biogenesis occurs in the endoplasmic reticulum (ER) (Park etal., 2010, 2013). After processing and transit to the plasma membrane, XA21 binds to XB24 (XA21 binding protein 24) (Chen et al., 2010b). XB24 physically associates with the XA21 juxtamembrane (JM) domain and catalyzes the autophosphorylation of serine and threonine residue(s) on XA21, keeping XA21 in an inactive state (Chen et al., 2010b). Upon pathogen recognition, XA21 kinase disassociates from XB24 and is activated (Chen et al., 2010b). This activation triggers a series of downstream events resulting in a robust resistance resp (...truncated)