10The phytohormone ethylene is widely involved in many developmental processes and is 11 a crucial regulator of defense responses against biotic and abiotic stresses in plants. 12Ethylene-responsive element binding protein (EREBP), a member of the 13 APETALA2/ethylene response factor (AP2/ERF) superfamily, is a transcription factor that 14 regulates stress-responsive genes by recognizing a specific cis-acting element of target DNA. 15A previous study showed only the NMR structure of the AP2/ERF domain of AtERF100 in 16 complex with a GCC box DNA motif. In this report, we determined the crystal structure of 17 AtERF96 in complex with a GCC box at atomic resolution. We analyzed the binding residues 18 of the conserved AP2/ERF domain in the DNA recognition sequence. In addition to the 19 AP2/ERF domain, an N-terminal α-helix of AtERF96 participates in DNA interaction in the 20 flanking region. We also demonstrated the structure of AtERF96 EDLL motif, a unique 21 conserved motif in the group IX of AP2/ERF family, is critical for the transactivation of 22 defense-related genes. Our study establishes the structural basis of the AtERF96 transcription 23 factor in complex with the GCC box, as well as the DNA binding mechanisms of the 24 All rights reserved. No reuse allowed without permission.

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