High‐throughput in planta expression screening identifies a class II ethylene‐responsive element binding factor‐like protein that regulates plant cell death and non‐host resistance

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Abstract
We performed high‐throughput screening using the potato virus X (PVX) system to overexpress Nicotiana benthamiana genes in planta and identify positive regulators of cell death. This screening identified NbCD1, a novel class II ethylene‐responsive element binding factor (ERF), as a potent inducer of the hypersensitive response (HR)‐like cell death. NbCD1 expression was induced by treatments with INF1 elicitor and a non‐host pathogen Pseudomonas cichorii. NbCD1 exhibited transcriptional repressor activity through its EAR motif, and this motif was necessary for NbCD1 to cause cell death. We identified 58 genes that displayed altered transcription following NbCD1 overexpression. NbCD1 overexpression downregulated the expression of HSR203, a negative regulator of hypersensitive death. Conditional expression of NbCD1 in Arabidopsis also caused cell death, indicating that NbCD1 downstream cascades are conserved in dicot plants. To further confirm the role of NbCD1 in defense, we used virus‐induced gene silencing to demonstrate that NbCD1 is required for non‐host resistance of N. benthamiana to the bacterial pathogen P. cichorii. Our data point to a model of transcriptional regulatory cascades. NbCD1 positively regulates cell death and contributes to non‐host resistance, possibly by downregulating the expression of other defense response genes.