The Cyst Nematode SPRYSEC Protein RBP-1 Elicits Gpa2- and RanGAP2-Dependent Plant Cell Death

Melanie Ann Sacco,Kamila Koropacka,Marianne J Jaubert,Geert Smant,Eric Grenier,Alexandra Blanchard,Peter Moffett,Aska Goverse,Charles Opperman

doi:10.1371/journal.ppat.1000564

Abstract

Plant NB-LRR proteins confer robust protection against microbes and metazoan parasites by recognizing pathogen-derived avirulence (Avr) proteins that are delivered to the host cytoplasm. Microbial Avr proteins usually function as virulence factors in compatible interactions; however, little is known about the types of metazoan proteins recognized by NB-LRR proteins and their relationship with virulence. In this report, we demonstrate that the secreted protein RBP-1 from the potato cyst nematode Globodera pallida elicits defense responses, including cell death typical of a hypersensitive response (HR), through the NB-LRR protein Gpa2. Gp-Rbp-1 variants from G. pallida populations both virulent and avirulent to Gpa2 demonstrated a high degree of polymorphism, with positive selection detected at numerous sites. All Gp-RBP-1 protein variants from an avirulent population were recognized by Gpa2, whereas virulent populations possessed Gp-RBP-1 protein variants both recognized and non-recognized by Gpa2. Recognition of Gp-RBP-1 by Gpa2 correlated to a single amino acid polymorphism at position 187 in the Gp-RBP-1 SPRY domain. Gp-RBP-1 expressed from Potato virus X elicited Gpa2-mediated defenses that required Ran GTPase-activating protein 2 (RanGAP2), a protein known to interact with the Gpa2 N terminus. Tethering RanGAP2 and Gp-RBP-1 variants via fusion proteins resulted in an enhancement of Gpa2-mediated responses. However, activation of Gpa2 was still dependent on the recognition specificity conferred by amino acid 187 and the Gpa2 LRR domain. These results suggest a two-tiered process wherein RanGAP2 mediates an initial interaction with pathogen-delivered Gp-RBP-1 proteins but where the Gpa2 LRR determines which of these interactions will be productive.

Highlights

In plants, immune receptors encoded by disease resistance (R) genes confer resistance to a broad spectrum of biotrophic organisms including bacteria, fungi, oomycete, viruses, nematodes and arthropods [1]
Identification of a G. pallida AvrGpa2 candidate The NB-leucine-rich repeat (LRR) protein Gpa2 has previously been shown to interact with the Ran GTPase activating protein Ran GTPase-activating protein 2 (RanGAP2), which in turn is predicted to interact with Ran GTPase as part of its normal cellular function in nucleocytoplasmic trafficking and mitosis [15,23]
A Gp-Rbp-1 cDNA derived from G. pallida pathotype (Pa-) 2/3 population Chavornay was cloned into the binary vector pBIN61 under control of the cauliflower mosaic virus 35S promoter as a C-terminal HA-tagged EGFP fusion (Gp-RBP-1:EGFP:HA), but lacking its secretion signal peptide

Summary

Introduction

Immune receptors encoded by disease resistance (R) genes confer resistance to a broad spectrum of biotrophic organisms including bacteria, fungi, oomycete, viruses, nematodes and arthropods [1]. Plant NB-LRR proteins show striking similarities in domain organization and predicted structure to NOD-LRR proteins, which are involved in innate immune responses in animals [2,3]. Unlike NOD-LRRs, which tend to recognize pathogen-associated molecular patterns (PAMPs) associated with broad classes of pathogens, NB-LRR proteins recognize proteins which are specific to a particular pathogen or pathogen isolate(s). These proteins are known as avirulence (Avr) proteins as they render the pathogen unable to infect a host encoding a corresponding R gene and the interaction between host and pathogen genotypes is referred to as gene-for-gene resistance. Recognition of Avr proteins by NBLRR proteins results in the activation of defense responses that limit infection, and may lead to a characteristic form of cell death referred to as the hypersensitive response (HR)

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