Effector-triggered defence against apoplastic fungal pathogens

Abstract

R gene-mediated host resistance against apoplastic fungal pathogens is not adequately explained by the terms pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) or effector-triggered immunity (ETI). Therefore, it is proposed that this type of resistance is termed 'effector-triggered defence' (ETD). Unlike PTI and ETI, ETD is mediated by R genes encoding cell surface-localised receptor-like proteins (RLPs) that engage the receptor-like kinase SOBIR1. In contrast to this extracellular recognition, ETI is initiated by intracellular detection of pathogen effectors. ETI is usually associated with fast, hypersensitive host cell death, whereas ETD often triggers host cell death only after an elapsed period of endophytic pathogen growth. In this opinion, we focus on ETD responses against foliar fungal pathogens of crops.

Keywords: R gene-mediated resistance; apoplastic fungal pathogens; cell wall; extracellular matrix; receptor-like protein.

Figure 1

Phenotypes of effector-triggered defence (ETD), effector-triggered immunity (ETI), or effector-triggered susceptibility (ETS) associated with recognition of effectors from representative fungal or oomycete leaf pathogens (featured in Table 1 or Table 2, main text) by contrast with phenotypes associated with nonrecognition of these effectors. ETD (A–H) involves limited or no macroscopic symptom development when apoplastic fungal leaf pathogen effectors are recognised by the corresponding R genes in the individual hosts (A1, A2, C1, E1, G1). The operation of the R gene against apoplastic fungal leaf pathogens limits pathogen growth but does not eliminate the pathogen, which can often subsequently sporulate. ETD in the resistant oilseed rape cultivar ‘Imola’ limited asexual sporulation (acervuli) of Pyrenopeziza brassicae (light leaf spot) and dark flecking occurred on (A1) the lamina and (A2) especially along the leaf midrib, as observed 23 days post inoculation (dpi) . (A3) The operation of the R gene against P. brassicae limited subcuticular hyphal growth, as observed 13 dpi in scanning electron micrographs (SEM, scale bar = 100 μm) of leaf surfaces, but (A4) it did not prevent sexual sporulation because P. brassicae apothecia subsequently developed on senescent leaves (scale bar = 0.5 mm). (B3) By contrast, on a susceptible oilseed rape cultivar, extensive subcuticular hyphal growth was observed at 13 dpi (SEM, scale bar = 100 μm), (B1) followed by asexual sporulation (acervuli); (B2) apothecia subsequently developed on senescent leaves (scale bar = 0.5 mm). (C1) Recognition of the Rhynchosporium commune (leaf blotch) NIP1 effector by the corresponding Rrs1 receptor of the resistant barley cultivar Turk was not associated with macroscopically visible symptom development, whereas (D1) necrotic lesions developed by 21 dpi with a Δnip1 R. commune isolate . (C2) Limited colonisation and asexual sporulation were observed 21 dpi on the resistant barley cultivar Atlas 46 inoculated with the R. commune transformant T-R214-GFP (confocal imaging) in contrast to (D3) extensive sub-cuticular hyphal (H) growth of R. commune observed by 17 dpi on susceptible barley leaves (SEM, scale bars 10 μm) and (D2) extensive colonisation and sporulation on the susceptible cultivar Atlas by 21 dpi. (E1) ETD operated in a resistant tomato inoculated with Cladosporium fulvum (leaf mould) that did not develop any visible symptoms by 14 dpi. (F1) By contrast, the pathogen grew extensively on a susceptible tomato cultivar, with mould developing as light brown patches in which conidiophores erupted through the stomata to produce asexual spores. (E2) ETD against C. fulvum growing in the apoplast of a tomato was associated with cell-wall enforcement (black arrow) without visible cell death early after inoculation (3 dpi) but (F2) no cell-wall enforcement had taken place on susceptible tomato plants at 3 dpi with the virulent C. fulvum race (H: pathogen hyphae, white arrow) . (G1) ETD triggered by the Leptosphaeria maculans (phoma leaf spot) AvrLm6 effector when it was recognised by the Rlm6 receptor on the resistant oilseed rape cultivar DarmorMX did not involve symptom development by 11 dpi with ascospores (without wounding) . (G2) Small dark spots (black arrows) and green islands (white arrows) were observed on DarmorMX 18 dpi when the leaf started to senesce. (G3) There was a necrotic response on leaves of DarmorMX associated with dead plant cells (lack of red chlorophyll fluorescence); however, the pathogen was alive within these small necrotic areas (white arrows) after inoculation with conidia of GFP-expressing L. maculans, when viewed under a fluorescent microscope (inoculation with wounding) (scale bar 200 μm). When there was no effector recognition (H1, at 22 h post inoculation) (H2, 42 h post inoculation (SEM, scale bar 10 μm)), germ tubes produced from L. maculans ascospores penetrated stomata on oilseed rape leaves . (H3) There was extensive cell death and lesion formation (grey, >2 mm in diameter) on leaves of Darmor (without Rlm6) 11 dpi with ascospores of L. maculans carrying the effector gene AvrLm6. (H4) When there was no recognition of the AvrLm6 effector, the pathogen produced an extensive hyphal network with pycnidia, as demonstrated by using a GFP-expressing L. maculans isolate carrying the effector gene AvrLm6 (white arrows) (scale bar 200 μm) before growing along the leaf petiole to the stem, the organ in which sexual sporulation occurs. In contrast to ETD, ETI (I–J) resulted in a macroscopic hypersensitive response on resistant potato (genotype 7644-17, derived from Solanum avilesii genotype 478-2) when production of the Rpi-avl1 protein operated against the corresponding Phytophthora infestans (late blight) effector. When there was no recognition of pathogen effectors (J1), typical late blight lesions with necrosis and chlorosis developed after 13 dpi with P. infestans isolate IPO-C on the susceptible cultivar ‘Nicola’ in a field experiment with (J2) Phytophthora infestans sporulating in chlorotic areas on the lower surface of the leaf. In contrast to ETD, ETS (K) results in programmed cell death (PCD) and the pathogen proliferated by 5 days post infiltration with isoforms of the host-selective toxin ToxA from Phaeosphaeria nodorum (glume blotch) in the wheat line BG261 that carries the sensitivity gene Tsn1. (L) No obvious necrosis was induced in the recessive tsn1 line BR34 by 5 days post infiltration with the same ToxA isoform . Modified, with permission, from (A2, A3, A4, B1, B2, B3), (C2, D2), (E2, F2), (G1, G2, G3, H3, H4), (H1, H2), and (K,L). C1, D1 provided by Wolfgang Knogge (Leibniz Institute of Plant Biochemistry, Germany); D3 by Kevin King and Jean Devonshire (Rothamsted Research, UK); and I, J1, and J2 by Vivianne Vleeshouwers (Wageningen University, The Netherlands).

Figure 2

Three types of host response to filamentous leaf pathogens, based on examples from Table 1 or Table 2 (main text). This diagram illustrates specific interactions between single pathogen effectors and corresponding host gene products. In reality, pathogens secrete numerous effectors that directly or indirectly interact with corresponding host gene products. (A) Resistance (R) gene-mediated effector-triggered defence (ETD) results in incompatible interactions with hemibiotrophic apoplastic fungal leaf pathogens. Extracellular recognition of effectors from fungal pathogens growing underneath the host cuticle (C) (Rhychosporium commune, Pyrenopeziza brassicae, and Venturia inaequalis) or between mesophyll cells (Cladosporium fulvum, Leptosphaeria maculans, and Zymoseptoria tritici) by receptor-like proteins (RLPs) can result in resistance without macroscopically visible host cell death (C. fulvum and R. commune) (▨). Host cell death typically occurs in only a few cells several days (C. fulvum and L. maculans) or weeks (R. commune and P. brassicae) after infection. The pathogen does not die () but can resume growth after host senescence begins or after the immune response is otherwise compromised. (B) In compatible interactions, in the absence of an RLP, the host stays alive (□) and the virulence function of the effector can promote extensive fungal proliferation (). (C) In the absence of the effector, the pathogen may proliferate less (). (D)R gene-mediated effector-triggered immunity (ETI) results in incompatible interactions with obligate biotrophic fungal (Blumeria graminis and Puccinia striiformis), oomycete (Bremia lactucae) pathogens, or some hemibiotrophic oomycete (Phythophthora infestans) or fungal (Magnaporthe grisea) pathogens. Upon formation of an appressorium (A) to breach the cell wall (CW) and penetrate an epidermal cell, specific fungal or oomycete effectors are secreted and delivered into the host cytoplasm, where recognition by corresponding nucleotide-binding site (NBS) leucine-rich repeat (LRR) receptors (NLRs) occurs. This recognition event triggers a rapid hypersensitive response (typically <1 day after infection) that boosts host defence and usually results in host (■) and pathogen cell death (). (E) Compatible interactions lead to the formation of haustoria (H) or a biotrophic interfacial complex through plasma membrane (PM) invaginations. In this case, the host cells stay alive (□). The effector stimulates pathogen proliferation (). (F) In the absence of the effector that compromises basal plant defence responses, pathogen growth () is slower. (G) Effector-triggered susceptibility (ETS) results in compatible interactions with necrotrophic fungal pathogens that secrete host-selective toxins (HSTs). Before entry through the leaf epidermis by means of penetration structures (P) such as hyphopodia (Phaeosphaeria nodorum) or appressoria (Cochliobolus victoriae), HSTs are released to target specific host proteins that are sensitive to the toxin (some are R gene products) and trigger host cell death (■) (typically within a day). Arrows indicate the final cellular destination of effectors of HSTs. Effectors are not injected into but taken up by the host cell. This leads to fungal proliferation (). Membrane invaginations do not occur. Entry into the leaf is also possible through stomata without development of penetration structures (P. nodorum). (H) In incompatible interactions and absence of host cell death (□), the fungal pathogen attempts to penetrate but cannot invade leaves. The pathogen can grow and survive on the plant surface for several days before it dies when nutrients are exhausted (). (I) Presence or absence of HST or its target has no impact on superficial growth. Colour codes for molecules and domains, which are not drawn to scale: effector or HST ; LRR domains ; NBS ; coiled-coil or Toll/interleukin-1 receptor domains ; transmembrane domain . NLRs are colour-coded the same for ETI and ETS because the same receptor may confer resistance against a biotrophic pathogen and susceptibility to a necrotrophic pathogen.

Figure I

Pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) is conserved and this host defence response against apoplastic pathogens (AP) and haustoria-forming (and other cell-penetrating) pathogens (HFP) does not generally differ. Although R gene-mediated resistance operates against both AP and HFP, these defence responses differ in that effector-triggered immunity (ETI) and effector-triggered defence (ETD) operate against HFP and AP, respectively. Specific differences are explained in Table I.