A Single Binding Site Mediates Resistance- and Disease-Associated Activities of the Effector Protein NIP1 from the Barley Pathogen Rhynchosporium secalis

Abstract

The effector protein NIP1 from the barley (Hordeum vulgare) pathogen Rhynchosporium secalis specifically induces the synthesis of defense-related proteins in cultivars of barley expressing the complementary resistance gene, Rrs1. In addition, it stimulates the activity of the barley plasma membrane H 1 -ATPase in a genotype-unspecific manner and it induces necrotic lesions in leaf tissues of barley and other cereal plant species. NIP1 variants type I and II, which display quantitative differences in their activities as elicitor and H 1 -ATPase stimulator, and the inactive mutant variants type III* and type IV*, were produced in Escherichia coli. Binding studies using 125 I-NIP1 type I revealed a single class of binding sites with identical binding characteristics in microsomes from near-isogenic resistant (Rrs1) and susceptible (rrs1) barley. Binding was specific, reversible, and saturable, and saturation ligand-binding experiments yielded a K d of 5.6 nM. A binding site was also found in rye (Secale cereale) and the nonhost species wheat (Triticum aestivum), oat (Avena sativa), and maize (Zea mays), but not in Arabidopsis (Arabidopsis thaliana). For NIP1 types I and II, equilibrium competition-binding experiments revealed a correlation between the difference in their affinities to the binding site and the differences in their elicitor activity and H 1 -ATPase stimulation, indicating a single target molecule to mediate both activities. In contrast, the inactive proteins type III* and type IV* are both characterized by high affinities similar to type I, suggesting that binding of NIP1 to this target is not sufficient for its activities.

Physiological and proteomic approaches to evaluate the role of sterol binding in elicitin-induced resistance

“…In both cases a similar hyperbolical binding curve was found with similar binding parameters to those of elicitins (Kooman-Gersmann et al , 1998; van’t Slot et al , 2007). In addition, as for AVR9 and NIP1, an elicitor high-affinity binding site has been identified in different plant cultivars or species, but the effective response was observed only in the resistant genotype.…”

Proteinaceous necrotrophic effectors in fungal virulence

“…Nip1 and 3 has been shown to stimulate barley plasma H + ATPase, which may be the likely cause of host tissue necrosis (Wevelsiep et al 1991(Wevelsiep et al , 1993. Nip1 has recently been shown to bind to a single unidentified receptor that triggers the plant's defence response (van't Slot et al 2007). In addition, Nip1 also functions as an avirulence effector on barley varieties that possess the uncloned Rrs1 gene (Rohe et al 1995).…”

The emergence of the multi‐species NIP1 effector in Rhynchosporium was accompanied by high rates of gene duplications and losses

“…NIP1 was first identified in R. commune and shown to stimulate a plasma membrane-localized H + -ATPase (Wevelsiep et al, 1991(Wevelsiep et al, , 1993Rohe et al, 1995;van't Slot et al, 2007). NIP1 promotes the formation of necrotic lesions on barley leaves but can also trigger strong host defence reactions if the host expresses the cognate receptor encoded by the resistance gene Rrs1.…”