An RLP23/Cf-9TM-IC Chimeric Receptor Enhances nlp24-Triggered Immunity and Resistance to Phytophthora nicotianae in Nicotiana benthamiana.

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-9TM-IC producing markedly stronger macroscopic symptoms than RLP23/Cf-9IC. Stable transgenic N. benthamiana expressing an untagged RLP23/Cf-9TM-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-9TM-IC as an effective chimeric receptor and reveal transient NRC3 expression as a practical step to further potentiate nlp24-triggered output in N. benthamiana.