Evolutionary gain and loss of a plant pattern-recognition receptor for HAMP recognition
Abstract
Pattern recognition receptors (PRR) recognize distinct pathogen and herbivore-associated molecular patterns (PAMPs and HAMPs) and mediate activation of immune responses, but the evolution of new PRR sensing functions is not well understood. We employed comparative genomics and functional analysis to define evolutionary events leading to the sensing of the peptide HAMP inceptin (In11) by the PRR Inceptin Receptor (INR). Existing and de novo genome assemblies revealed that the presence of a functional INR gene corresponded with In11 response across 55 million years (my) of legume evolution, and that In11 recognition is unique to the clade of Phaseoloid legumes. The INR loci of certain Phaseoloid and non-Phaseoloid species also contain diverse INR-like homologues, suggesting that the evolution of INR receptor function ∼28 mya occurred after an ancestral gene insertion ∼32 mya. Functional analysis of chimeric and ancestrally reconstructed receptors revealed that specific AA differences in the C1 leucine-rich repeat (LRR) domain and C2 intervening motif likely mediated gain of In11 recognition. In summary, we present a conceptual model for the evolution of a defined PRR function based on patterns of INR variation in legumes.
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Evaluation Summary:
This manuscript, of interest to those studying the evolution of immunity, investigates the evolutionary history of a recently described herbivore-associated molecular pattern (HAMP) receptor, INR, which perceives the caterpillar-derived peptide HAMP, In11. The authors compare INR homologs to identify evolutionarily conserved residues and use chimeric fusion proteins to investigate specificity. The findings presented are valuable and supported by convincing experiments and analysis.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #1 agreed to share their name with the authors.)
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Reviewer #1 (Public Review):
The paper by Snoeck et al. addresses the evolution of the recognition of inceptin, a peptide from insect saliva, by plant immune receptor INR, a member of LRR-type receptor-like protein family. As a first step, the authors surveyed how broad inceptin recognition is among legumes and found that it likely emerged in the common ancestor of Phaseolid legumes. By considering available genomic information and supplementing it with several de novo sequenced species, the authors were able to show that all extant inceptin receptor sequences form a single phylogenetic clade, supporting a single origin for INR evolution, an event that was followed by several independent losses. The authors also describe a closely related INR-like clade that lacks inceptin recognition. By considering chimeras between INR and INR-like …
Reviewer #1 (Public Review):
The paper by Snoeck et al. addresses the evolution of the recognition of inceptin, a peptide from insect saliva, by plant immune receptor INR, a member of LRR-type receptor-like protein family. As a first step, the authors surveyed how broad inceptin recognition is among legumes and found that it likely emerged in the common ancestor of Phaseolid legumes. By considering available genomic information and supplementing it with several de novo sequenced species, the authors were able to show that all extant inceptin receptor sequences form a single phylogenetic clade, supporting a single origin for INR evolution, an event that was followed by several independent losses. The authors also describe a closely related INR-like clade that lacks inceptin recognition. By considering chimeras between INR and INR-like receptors, the authors map specificity to C1 (leucine-rich repeat) and C2 (insertion domain) regions of the protein. By testing inferred ancestral INR sequences they limit the number of amino acid residues responsible for the original ability to recognize inceptin to just a few residues.
The approach is well reasoned, the two complementary functional assays - ROS time course and ethylene accumulation time point - are qualitatively concordant, and the controls - expression level in heterologous assay - appropriate. Phylogenetic conclusions are likewise well supported. The authors have also done well to make the data on newly sequenced organisms available through NCBI.
There are two aspects of the study that could be improved. One is following up on the genomic events leading to independent INR loss events. Were there deletions, transposon insertions, point mutations leading to early stop codons, etc.? The other missing part is a structural interpretation of mutations leading to inceptin recognition. While I agree with the authors that an experimental structure of INR/peptide/co-receptor would be ideal, an AlphaFold or RoseTTaFold model of the N3/N4/N14 series might highlight where the key changes occurred leading to inceptin recognition. It could also hint at the N3 function, for example, was N3 already a likely foreign peptide receptor?
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Reviewer #2 (Public Review):
The manuscript is of high quality. It is well written, concise, and easy to follow both in the text and figures. The methods give sufficient detail. Snoeck et al. conducted an in-depth analysis of the INR locus and In11 recognition across Legumes. They provide an extensive functional characterisation of INR orthologs and use ancestral sequence prediction to identify residues that appear to be important for INR function in In11 perception. In order to do this, they generate considerable genetic resources. To my knowledge, this represents the most extensive analysis of this sort on plant cell surface immune receptors.
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