ZmWAK3-mediated cell wall remodeling and stomatal dynamics modulate drought tolerance in maize seedlings

ZmWAK3 negatively regulates drought tolerance in maize seedlings by modulating cell wall remodeling and stomatal dynamics, and is directly regulated by the ZmWRKY44 transcription factor. Drought, a significant abiotic stress affecting maize production in key growing regions, necessitates a deeper understanding of its regulatory mechanisms to develop drought-resistant varieties and ensure yield stability. WAKs are pivotal receptor kinases in cell wall signaling, mediating extracellular-to-intracellular communication and participating in diverse processes, including cell expansion, stress adaptation, and pathogen defense. However, the role of WAKs in the drought response remains poorly elucidated. Functional analysis of ZmWAK3 through overexpression and mutant lines has revealed its negative regulatory role in maize drought tolerance. It was found that ZmWAK3 reduces pectin content by increasing polygalacturonase activity, thereby promoting cell wall relaxation. Furthermore, ZmWAK3 was observed to regulate stomatal aperture. Additionally, we demonstrated that the transcription factor ZmWRKY44 directly activates ZmWAK3 expression by binding to a W-box cis-element within its promoter. Crucially, we identified a drought-associated InDel locus within ZmWAK3 and developed the functional marker ZmWAK3-177, which effectively distinguishes drought-tolerant alleles and serves as a practical tool for marker-assisted selection in breeding programs. Collectively, these results reveal a novel mechanism of ZmWAK3 in drought stress response and provide actionable genetic resources for improving maize drought resilience.