Genome-wide analysis of RAD23 gene family and a functional characterization of AcRAD23D1 in drought resistance in Actinidia

• Six AcRAD23 genes were identified in kiwifruit. • All AcRAD23 genes showed differential expression patterns under various stress conditions. • AcRAD23D1 may play a positive role in regulating kiwifruit’s response to drought stress. UBL-UBA protein functions as a shuttle factor in the 26S ubiquitin degradation pathway, playing a critical role in plant growth and development, and responding to various biotic and abiotic stresses. Although RAD23, a type of UBL-UBA protein, has been extensively studied in several plants, there is currently no comprehensive analysis available for kiwifruit ( Actinidia chinensis ). In this study, we identified six AcRAD23 genes in kiwifruit and further analyzed their phylogenetic relationships, gene structure, conserved motif composition and cis -acting element in the promoter. Subcellular localization experiments revealed that all AcRAD23 were localized in the nucleus and the cell membranes. Quantitative real-time PCR (qRT-PCR) analysis demonstrated differential expression patterns of these AcRAD23 genes across different tissues and under various stress conditions (drought, waterlogging, salt stress, etc.), with AcRAD23D1 showing the highest responsiveness to abiotic stress. Additionally, we investigated the biological function of AcRAD23D1 using VIGS-mediated gene silencing methods under drought stress conditions. Suppression of AcRAD23D1 expression resulted in reduced relative water content (RWC) but increased malondialdehyde (MDA) content and relative electrolyte leakage (REL) levels in D1-VIGS lines compared to control lines. Furthermore, D1-VIGS lines exhibited a higher accumulation of reactive oxygen species (ROS) along with decreased superoxide dismutase (SOD) and peroxidase (POD) enzyme activities. These findings suggest that AcRAD23D1 may play a positive role in regulating kiwifruit’s response to drought stress. Our results provide new insights into the potential involvement of AcRAD23 under abiotic stress conditions while offering a theoretical foundation for understanding the molecular mechanisms underlying kiwifruit’s adaptation to stresses.