Key message <p>The drought stress-responsive transcription factor ZmbZIP38 interacts with ZmDnaJ to regulate maize drought tolerance through modulating ABA biosynthesis and signaling, stomatal closure and root growth under stress.</p> Abstract <p>Basic leucine zipper (bZIP) transcription factors (TFs) crucially regulate plant drought stress response. However, how bZIP TFs regulate maize drought tolerance remains elusive. Previously, we demonstrated that <i>ZmDnaJ</i> enhances maize drought tolerance by promoting ABA biosynthesis and stomatal closure. In this study, we have fished out ZmbZIP38 (by yeast one-hybrid analysis) as key interacting partner of ZmDnaJ, and elucidate its function in <i>ZmDnaJ</i>-mediated drought tolerance in maize. <i>ZmDnaJ</i> promoter analysis results showed that ZmbZIP38 directly targets ZmDnaJ by binding to ABRE motifs in the corresponding gene promoters. Overexpression of <i>ZmbZIP38</i> significantly enhanced maize survival rate under drought stress, which was accompanied by expanded leaf area and higher stomatal closure. <i>ZmbZIP38</i>-overexpressing maize lines also showed enhanced ROS scavenging, reduced H₂O₂ and MDA accumulation, and up-regulated expression of antioxidant enzymes-associated genes. Moreover, <i>ZmbZIP38</i>-OE maize lines exhibited elevated ABA levels under drought stress, correlating with the up-regulated expression of ABA biosynthetic genes. Conversely, <i>zmbzip38</i> knockdown mutants displayed reduced drought tolerance, evidenced by increased cell damage and decreased leaf area. Collectively, our findings demonstrate that <i>ZmbZIP38</i> regulates maize seedling drought tolerance by modulating ABA biosynthesis and signaling, ROS scavenging and root growth, highlighting its potential role in abiotic stress response, and particularly enhancing maize drought tolerance.</p>