Protein-protein interactions in the RPS4/RRS1 immune receptor complex

Abstract

Plant NLR (Nucleotide-binding domain and Leucine-rich Repeat) immune receptor proteins are encoded by Resistance (R) genes and confer specific resistance to pathogen races that carry the corresponding recognized effectors. Some NLR proteins function in pairs, forming receptor complexes for the perception of specific effectors. We show here that the Arabidopsis RPS4 and RRS1 NLR proteins are both required to make an authentic immune complex. Over-expression of RPS4 in tobacco or in Arabidopsis results in constitutive defense activation; this phenotype is suppressed in the presence of RRS1. RRS1 protein co-immunoprecipitates (co-IPs) with itself in the presence or absence of RPS4, but in contrast, RPS4 does not associate with itself in the absence of RRS1. In the presence of RRS1, RPS4 associates with defense signaling regulator EDS1 solely in the nucleus, in contrast to the extra-nuclear location found in the absence of RRS1. The AvrRps4 effector does not disrupt RPS4-EDS1 association in the presence of RRS1. In the absence of RRS1, AvrRps4 interacts with EDS1, forming nucleocytoplasmic aggregates, the formation of which is disturbed by the co-expression of PAD4 but not by SAG101. These data indicate that the study of an immune receptor protein complex in the absence of all components can result in misleading inferences, and reveals an NLR complex that dynamically interacts with the immune regulators EDS1/PAD4 or EDS1/SAG101, and with effectors, during the process by which effector recognition is converted to defense activation.

Constant vigilance: plant functions guarded by resistance proteins

“…However, the observation that RPS4 overexpression, when the amount of RPS4 protein exceeds that of RRS1, induces auto‐immunity and HR (Huh et al ., ) is more consistent with the dissociation model. In addition, co‐expression of RRS1 can abolish the RPS4 overexpression‐induced phenotype (Huh et al ., ), indicating that RRS1 is required to lock RPS4 in an inactive state under normal conditions and that higher RPS4 protein amounts compared with RRS1 could activate this system (Figure c). RPS4 is induced by AvrRps4 (Zhang and Gassmann, ), and protein complexes containing RPS4 are also modulated by additional proteins such as SRFR1 and SGT1 (Kwon et al ., ; Kim et al ., ; Li et al ., ), which are not usually included in transient expression studies.…”

“…This model may be an oversimplification, as it was shown that AvrRps4 does not disrupt the RPS4–EDS1 interaction in the presence of RRS1, indicating that RPS4 activation may be mediated by AvrRps4‐induced RPS4‐RRS1 conformational changes (Figure b; Huh et al ., ). However, the observation that RPS4 overexpression, when the amount of RPS4 protein exceeds that of RRS1, induces auto‐immunity and HR (Huh et al ., ) is more consistent with the dissociation model. In addition, co‐expression of RRS1 can abolish the RPS4 overexpression‐induced phenotype (Huh et al ., ), indicating that RRS1 is required to lock RPS4 in an inactive state under normal conditions and that higher RPS4 protein amounts compared with RRS1 could activate this system (Figure c).…”

“…Because the interaction between EDS1 and the two unrelated pathogen effectors AvrRps4 and HopA1 also occurred in vitro, it was concluded that EDS1 is a direct target of these effectors (Bhattacharjee et al, 2011). Although previously disputed by work from a separate group (Sohn et al, 2012), recently published data confirm the interaction between AvrRps4 and EDS1 by co-immunoprecipitation and bimolecular fluorescence complementation analyses (Huh et al, 2017).…”

“…Bhattacharjee et al showed that AvrRps4 or HopA1 disrupt RPS4-EDS1 or RPS6-EDS1 interactions, and proposed that the disassociation of RPS4 and RPS6 from EDS1 leads to activation (Figure 7a) (Bhattacharjee et al, 2011). This model may be an oversimplification, as it was shown that AvrRps4 does not disrupt the RPS4-EDS1 interaction in the presence of RRS1, indicating that RPS4 activation may be mediated by AvrRps4-induced RPS4-RRS1 conformational changes (Figure 7b;Huh et al, 2017). However, the observation that WRKY transcription factors play key roles in plant immunity, and the Ralstonia solanacearum effector PopP2 acetylates a subset of WRKY TFs to induce ETS.…”

Multiple strategies for pathogen perception by plant immune receptors

“…Indeed, genetically linked pairs of NLR genes were found to function cooperatively in pathogen resistance (Cesari et al, 2014a). In cases where the functional mechanism of paired NLRs was studied, one NLR behaved as an effector receptor involving an ID, while the second NLR induced signaling (Cesari et al, 2014b;Williams et al, 2014;Huh et al, 2017). In other instances, NLRs form signaling networks beyond genetically linked pairs, thereby broadening the concept of NLR cooperation .…”