Evolution trajectory of auxin response factor across Triticeae species underpin candidate genes for the regulation of salt/alkali stress responsive networks.

key messageSystematically characterized the ARF family in Triticeae revealed its evolutionary expansion patterns and validated that TaARF4.1 acts as a repressor enhancing salt/alkali tolerance potentially by regulating cell wall metabolism and ROS homeostasis. Auxin response factors (ARFs) are core transcription factor families mediating auxin signaling, which not only regulate plant growth and development but also bridge the trade-off between growth and stress adaptation. However, their roles in responding to abiotic stresses-particularly salt/alkali stress, a major constraint to global wheat production-remain poorly understood in wheat (Triticum aestivum). To address this, we integrated evolutionary genomics, multi-tissue expression profiling, and functional validation across 10 Poaceae species (including wheat, rice, maize, and other Triticeae crops) to systematically identify stress-regulatory wheat ARFs (TaARFs). We identified 349 ARF members and reconstructed the evolutionary trajectory across 10 species, clarified orthologous relationships between TaARFs and rice ARFs (OsARFs), and revealed that TaARF expansion is driven by two mechanisms: ancient whole-genome duplication (WGD) conserving core ARF functions and recent gene duplication burst (RBGD) generating Triticeae-specific duplicates (e.g., TaARF4.1/4.2). GO enrichment and stress-induced expression analyses highlighted Group IV repressor ARFs (TaARF4.1/4.2/9) as candidate regulators of abiotic stress responses. Transcriptome integration across salt/alkali-treated wheat leaves and roots identified stably stress-responsive TaARFs, while functional assays confirmed TaARF4.1, as a repressor ARF with the whole-cell localization, was associated with key genes involved in stress signaling, cell wall metabolism, and reactive oxygen species (ROS) homeostasis. This study demonstrates that underutilized genome evolution data aids gene mining in complex crop genomes, providing novel genetic resources for wheat salt/alkali tolerance breeding and insights into auxin-mediated stress adaptation mechanisms.