An RLP23/Cf‐9 ^TM‐IC Chimeric Receptor Enhances nlp24‐Triggered Immunity and Resistance to Phytophthora nicotianae in Nicotiana be

ABSTRACT

Receptor‐like protein AtRLP23 from Arabidopsis thaliana perceives the conserved microbial peptide nlp24 derived from necrosis‐ and ethylene‐inducing peptide 1‐like proteins (NLPs), but immune outputs are often weak in heterologous systems. Here, we engineered RLP23/Cf‐9 chimeric receptors by fusing the extracellular domain of AtRLP23 to either the Cf‐9 intracellular region (IC) or the Cf‐9 transmembrane–intracellular region (TM–IC), and assessed their activity in Nicotiana benthamiana . In transient expression assays, AtRLP23 alone did not trigger hypersensitive cell death after infiltration with 10 μM nlp24, whereas both chimeras conferred nlp24‐induced cell death, with RLP23/Cf‐9 ^TM‐IC producing markedly stronger macroscopic symptoms than RLP23/Cf‐9 ^IC . Stable transgenic N. benthamiana expressing an untagged RLP23/Cf‐9 ^TM‐IC chimera showed enhanced nlp24 responsiveness, including stronger seedling growth inhibition and cell death after repeated 10 μM nlp24 treatment. In these plants, virus‐induced gene silencing showed that chimera‐mediated cell death requires SOBIR1 and the helper NLR NRC3, and transient NRC3 expression further sensitized the response, enabling cell death after a single 0.1 μM nlp24 treatment. In addition, the chimera significantly enhanced resistance to the oomycete Phytophthora nicotianae . Together, these results identify RLP23/Cf‐9 ^TM‐IC as an effective chimeric receptor and reveal transient NRC3 expression as a practical step to further potentiate nlp24‐triggered output in N. benthamiana .