Structural polymorphisms within a common powdery mildew effector scaffold as a driver of coevolution with cereal immune receptors

Abstract

In plants, host–pathogen coevolution often manifests in reciprocal, adaptive genetic changes through variations in host nucleotide-binding leucine-rich repeat immune receptors (NLRs) and virulence-promoting pathogen effectors. In grass powdery mildew (PM) fungi, an extreme expansion of a RNase-like effector family, termed RALPH, dominates the effector repertoire, with some members recognized as avirulence (AVR) effectors by cereal NLR receptors. We report the structures of the sequence-unrelated barley PM effectors AVR A6 , AVR A7 , and allelic AVR A10 /AVR A22 variants, which are detected by highly sequence-related barley NLRs MLA6, MLA7, MLA10, and MLA22 and of wheat PM AVR PM2 detected by the unrelated wheat NLR PM2. The AVR effectors adopt a common scaffold, which is shared with the RNase T1/F1 family. We found striking variations in the number, position, and length of individual structural elements between RALPH AVRs, which is associated with a differentiation of RALPH effector subfamilies. We show that all RALPH AVRs tested have lost nuclease and synthetase activities of the RNase T1/F1 family and lack significant binding to RNA, implying that their virulence activities are associated with neo-functionalization events. Structure-guided mutagenesis identified six AVR A6 residues that are sufficient to turn a sequence-diverged member of the same RALPH subfamily into an effector specifically detected by MLA6. Similar structure-guided information for AVR A10 and AVR A22 indicates that MLA receptors detect largely distinct effector surface patches. Thus, coupling of sequence and structural polymorphisms within the RALPH scaffold of PMs facilitated escape from NLR recognition and potential acquisition of diverse virulence functions.

The structural landscape and diversity ofPyricularia oryzaeMAX effectors revisited

“…These findings highlighted that AF2 predicts MAX effector structures in a highly reliable and precise manner even at relatively low pLDDT scores. Similar observations were made in other recent studies where experimentally resolved structures of fungal effectors were compared with AF2 predictions, as for example in the case of LARS [43], FOLD [44] and RALPH [45] effectors. Comparison with a published study testing ab initio approaches using Rosetta or the two web servers Robetta and QUARK to model MAX effectors with already known structures [46] confirmed that AF2 outcompetes other strategies for the prediction of MAX effector structures.…”

Powdery mildew effectors AVRA1and BEC1016 target the ER J-domain proteinHvERdj3B required for immunity in barley

“…AVRA1 and BEC1016 both interact with HvERdj3B and both enter the ER posttranslationally and signal peptide-independently. They have very diverse amino acid sequences, but appear derived from an ancient RNAse and may potentially be structurally related (Cao et al, 2023;Pedersen et al, 2012), which may explain their shared functions. Interestingly, the ERQC machinery is known to engage in the post-translational ER protein uptake through the Sec61 translocon complex (Hassdenteufel et al, 2019;Zimmermann et al, 2011).…”

“…For instance, CSEP0008 (gene ID BLGH_03023) is recognized by MLA1, and thus named AVRA1 (Lu et al, 2016). Other recognized CSEPs are CSEP0059 (AVRA7), CSEP0174 (AVRA9), CSEP0141 (AVRA10 and AVRA22), and CSEP0372 (AVRA13) (Bauer et al, 2021;Cao et al, 2023;Lu et al, 2016;Saur et al, 2019;). AVRA6 is represented by three near-identical copies in the DH14 genome, BLGH_00709 (CSEP0254), BLGH_00708 and BLGH_07091 (Bauer et al, 2021;Cao et al, 2023), which may be expressed in an isolate-specific manor (Velásquez-Zapata et al, 2023b).…”

Powdery mildew effectors AVRA1 and BEC1016 target the ER J‐domain protein HvERdj3B required for immunity in barley

“…For instance, CSEP0008 (gene ID BLGH_03023) is recognized by MLA1, and thus named AVR A1 (Lu et al., 2016 ). Other recognized CSEPs are CSEP0059 (AVR A7 ), CSEP0174 (AVR A9 ), CSEP0141 (AVR A10 and AVR A22 ), and CSEP0372 (AVR A13 ) (Bauer et al., 2021 ; Cao et al., 2023 ; Lu et al., 2016 ; Saur et al., 2019 ). AVR A6 is represented by three near‐identical copies in the DH14 genome, BLGH_00709 ( CSEP0254 ), BLGH_00708 and BLGH_07091 (Bauer et al., 2021 ; Cao et al., 2023 ), which may be expressed in an isolate‐specific manner (Velásquez‐Zapata, Smith, et al., 2023 ).…”