GhWRKY70 interacted with GhMPK6 positively mediates drought tolerance in cotton

WRKY transcription factors (TFs) are key regulators of plant development and abiotic stress responses and characterized by the presence of one or two WRKY domains (conserved motifs WRKYGQK) at the N-terminal and zinc-like motifs at the C-terminal; however, their functional roles in cotton drought resistance remain largely unexplored. Here, we identified GhWRKY70, a group III WRKY TF from Upland cotton (Gossypium hirsutum), along with its homolog GsWRKY70 from the diploid wild cotton Gossypium stocksii. GhWRKY70 localizes predominantly to the nucleus and exhibits its highest expression in leaves. Its transcript levels were markedly upregulated under drought stress. Silencing GhWRKY70 compromised drought tolerance, leading to impaired stomatal closure, accelerated water loss, and reduced chlorophyll content in seedling and mature plants. These plants also accumulated more reactive oxygen species (ROS) and malondialdehyde (MDA), while showing diminished antioxidant enzyme activities. Conversely, transgenic Arabidopsis lines overexpressing GhWRKY70 promoted stomatal closure, reduce water loss, increase antioxidant enzyme activity to scavenge ROS and MDA, and thereby enhanced the drought resistance of GhWRKY70-OE Arabidopsis lines. Further analyses indicated that GhWRKY70 modulated the expression of ABA-dependent signaling genes GhNCED3, GhRD29A and GhP5CS, and promoted ABA-induced stomatal closure. Through BiFC, LCI, and Y2H assays, we demonstrated that GhWRKY70 interacted with GhMPK6. Importantly, knockdown of GhMPK6 resulted in heightened drought sensitivity, resembling the GhWRKY70-silenced phenotype, suggesting their coordinated action within the MAPK signaling cascade during drought response. Collectively, our study establishes GhWRKY70 as a positive regulator of drought tolerance in cotton, acting through the regulation of stomatal movement, enhancement of ROS scavenging capacity, and involvement in both ABA signaling and MAPK pathways.