Insect saliva proteins switch on an unusual plant immune receptor

When an aphid or a brown planthopper pierces a plant stem and begins to feed, it injects more than just a probing mouthpart. Along with the watery saliva that lubricates its stylet comes a cocktail of secreted proteins, some of which act as effectors that manipulate the host’s biology. A new study published in Plant Cell Reports reveals how one family of these insect salivary proteins, the chemosensory proteins, or CSPs, is detected by a plant immune receptor with a set of structural quirks that sets it apart from nearly every other receptor of its kind. The work, led by Weiwei Rao and Tingting Ma with colleagues at Hubei University of Technology, the Chinese Academy of Sciences, Wuhan University and Shandong University, dissects the molecular machinery behind a plant-insect arms race at a level of detail rarely achieved for herbivore interactions.

The receptor in question, named RCSP, belongs to the TNL family of plant immune proteins, which carry a Toll-interleukin-1 receptor, or TIR, domain alongside a nucleotide-binding region and a leucine-rich repeat array. TNL receptors are famous in plant pathology for recognizing pathogen effectors and triggering a localized cell death response that walls off infection. What makes RCSP remarkable is its target: rather than a bacterial or oomycete effector, it responds to CSPs delivered by phloem-feeding insects such as aphids and the brown planthopper Nilaparvata lugens. Previous work from the same group had shown that recognition of a salivary effector by RCSP promotes effector-triggered immunity and systemic resistance in the model plant Nicotiana benthamiana, causing the insects to induce dwarfism in the plant as part of a defensive response. The new study asks how, at the level of atoms and residues, this recognition actually works.

The answer, it turns out, involves three departures from the canonical TIR domain playbook. First, the RCSP-TIR domain carries a deletion of nine amino acids in the loop connecting beta-strand B and alpha-helix B, a region known as the BB-loop. Second, it contains an insertion of eight amino acids in the loop between alpha-helices 5 and 6, the alpha5alpha6-loop. Third, the glutamate residue that normally serves as a key catalytic component in TIR enzymatic chemistry is replaced by glutamine, an E87Q substitution. Each of these changes would be expected to compromise the standard functions of a TIR domain, and indeed the researchers found that the BB-loop deletion disrupts the canonical NAD+-binding pocket and prevents the standard tetramerization that many TIR domains use to become active enzymes.

That loss of conventional architecture raises an obvious puzzle: if RCSP cannot form the usual tetramer or bind NAD+ in the usual way, how does it signal at all? The team turned to AlphaFold3 and DMFold structure prediction to find out. Their modeling revealed a striking compensatory mechanism: the alpha5alpha6-loop insertion creates a novel protein-protein interface that stabilizes an atypical tetrameric architecture. In other words, the receptor appears to have traded its ancestral assembly interface for a new one, built from an inserted loop, that restores the multimeric state needed for activity. This kind of structural re-engineering, in which one evolutionary change offsets another, offers a vivid example of how immune receptors can diversify while preserving function.

The enzymatic story is equally unconventional. TIR domains in plants and animals are known to act as NAD+ cleaving enzymes, and this activity underlies their ability to drive cell death signaling. Molecular docking combined with site-directed mutagenesis indicated that RCSP may utilize a noncanonical NAD+-binding pocket and depends on a pair of catalytic residues, aspartate 86 and glutamine 87, to mediate what the authors describe as weak cell death. The dual-residue dependence is notable because the canonical glutamate is absent; glutamine at position 87, the very residue that replaced it, appears to be part of the catalytic machinery rather than a silent placeholder. The weak cell death phenotype also fits the biology of insect defense, where a measured response may be more useful than the explosive hypersensitive reaction typical of microbial pathogen recognition.

To place these findings in context, it helps to recall what is known about TIR domain signaling more broadly. Seminal work published in Science in 2019 demonstrated that animal and plant TIR domains cleave NAD+ in cell death pathways, and subsequent structural studies of proteins such as SARM1 in animals illuminated how NADase activity is regulated. In plants, activated TNL receptors assemble into large resistosome complexes, as visualized for ZAR1 and ROQ1, and signal through helper NLR proteins that function as calcium-permeable channels. The EDS1 family of proteins, together with SAG101 and NRG1, forms a coevolved module that mediates cell death signaling downstream of TIR-domain receptors. RCSP presumably plugs into this downstream framework in Nicotiana benthamiana, but its upstream activation mechanism, as the new study shows, is built on a distinctly nonstandard scaffold.

The evolutionary analysis adds another layer of interest. By surveying RCSP-TIR homologs across plant genomes, the researchers classified them into four distinct clades, with evidence highlighting Solanaceae-specific adaptations. This pattern suggests that the structural innovations seen in RCSP are not one-off accidents but part of a broader, family-wide diversification within the nightshade lineage, the group that includes tobacco, tomato, potato and pepper. Such lineage-specific expansion of atypical TIR domains could reflect an ongoing evolutionary dialogue with phloem-feeding insects, whose CSP effectors are conserved across aphids and planthoppers. If plants in this family have repeatedly reworked their TIR domains to detect these proteins, it would imply that CSP recognition has been a persistent selective pressure.

CSPs themselves are an intriguing class of effectors. Originally named for their putative roles in insect chemoreception, they are small, conserved proteins found in the saliva of phloem-feeding insects. Proteomic surveys of aphid and planthopper saliva have repeatedly identified them among secreted proteins, and functional studies have shown that some CSPs can modulate plant defenses. One aphid CSP, SmCSP4, has been reported to stimulate salicylic acid-mediated defense responses in wheat by interacting with a transcription factor. The fact that the same protein family can function as both a virulence factor and a trigger of immunity, depending on the host genotype, underscores the double-edged nature of effector biology: what is a weapon in a susceptible plant becomes a molecular signature in a plant carrying the right receptor.

The broader significance of this work lies in expanding the known repertoire of what plant immune receptors can recognize and how they can be built. Most research on NLR receptors has focused on microbial pathogens, and the structural biology of resistosomes has largely been worked out in that context. Herbivorous insects cause enormous agricultural losses, and the global burden of pests on major food crops is well documented, yet the molecular mechanisms by which plants detect insect attack have remained comparatively opaque. RCSP provides one of the clearest examples to date of a conventional NLR scaffold being repurposed for insect effector recognition, and it does so through structural changes, a deleted loop, an inserted loop and a swapped catalytic residue, that would have been difficult to predict from sequence alone.

For crop improvement, the implications are tantalizing. If the structural determinants of CSP recognition can be pinned down precisely, as this study begins to do with its docking and mutagenesis data, then engineering RCSP-like receptors or their recognition specificities into crop species could offer a route to durable resistance against aphids and planthoppers. The finding that weak cell death, mediated by an atypical catalytic mechanism, suffices for signaling also hints that immune outputs can be tuned, which matters because overly aggressive cell death can damage plant tissue. Much remains to be resolved, including the exact geometry of the noncanonical NAD+-binding pocket and how CSP binding allosterically activates the receptor, but the study establishes a compelling framework. A plant receptor has been caught in the act of reinventing itself, and the reinvention is aimed squarely at the insects that would eat the plant.

Subject of Research: Structural and catalytic mechanisms by which insect chemosensory protein effectors activate the atypical plant TNL immune receptor RCSP

Article Title: The insect effector CSPs activate an atypical plant TNL immune receptor

Article References: Rao, W., Li, T., Yang, J., Zhu, L., Zhao, W., & Ma, T. (2026). The insect effector CSPs activate an atypical plant TNL immune receptor. Plant Cell Reports, 45(10), Article 291. https://doi.org/10.1007/s00299-026-03972-w

Image Credits: AI Generated

DOI: 10.1007/s00299-026-03972-w

Keywords: plant immunity, chemosensory proteins, TNL receptor, TIR domain, RCSP, insect effectors, brown planthopper, aphids, NAD+ cleavage, Nicotiana benthamiana, effector-triggered immunity, plant-insect interaction