Natural variation and CRISPR/Cas9 gene editing demonstrate the potential for a group VII ethylene response factor HvERF62 in regulating barley (Hordeum vulgare L.) waterlogging tolerance

Abstract Waterlogging stress is a serious problem all over the world, which inhibits plant growth and development, and even leads to plant death in severe circumstances. Ethylene-responsive VII transcription factor (ERFVII) is a key factor regulating plant waterlogging tolerance. In this study, a major QTL conferring chlorophyll content under waterlogging stress was detected on chromosome 4H through GWAS using a natural population. Combined with RNA-Seq analysis, the group VII ethylene response factor HvERF62 was identified as the candidate gene. CRISPR/Cas9-guided knockout of HvERF62 mutants showed sensitivity to waterlogging, with lower chlorophyll content, less adventitious roots and lower root activity. Physiological and transcriptomic analyses showed that HvERF62 plays an important role in aerenchyma formation, ROS homeostasis and carbohydrate accumulation under waterlogging stress, and regulates waterlogging tolerance through starch and sucrose metabolism, MAPK signaling pathway, plant hormone signal transduction and glycolysis/gluconeogenesis signaling. Haplotype analysis showed that the amino acid coding of the waterlogging-intolerant haplotype Hap3 was terminated prematurely. This study provides a new genetic resource and a relevant marker for identification of waterlogging-sensitive materials and deepens our understanding of the molecular mechanisms of waterlogging response in barley.