Drought is a major abiotic stress that significantly limits maize productivity. However, previous transcriptomic studies with limited time-point sampling have hindered the construction of robust co-expression networks, making it challenging to identify reliable hub genes involved in drought tolerance. To overcome this limitation, we generated a high-temporal-resolution transcriptome dataset spanning 108 time points from maize seedlings subjected to two consecutive rounds of drought and re-watering treatments. A total of 8477 drought-responsive genes (DRGs) were identified by comparing drought-stressed and well-watered controls. Using weighted gene co-expression network analysis (WGCNA), we constructed 17 co-expression modules, of which 8 were strongly associated with drought stress responses and collectively contained 353 hub genes. Among them, we validated the drought resistance functions of ZmCPK35, a known drought-responsive gene, and Zinc-finger Inflorescence Meristem 23 (ZmZIM23), a newly identified drought-regulatory gene, within the M10 module. Functional analysis revealed that ZmZIM23 enhances drought tolerance by improving water-use efficiency, reducing transpiration rates, and promoting biomass accumulation. Furthermore, yeast one-hybrid (Y1H) and dual-luciferase (LUC) assays demonstrated that ZmWRKY40, another M10 module member, transcriptionally regulates both ZmZIM23 and ZmCPK35. By integrating high-resolution transcriptomic data with co-expression network analyses, this study unveils key drought-responsive regulatory networks in maize and identifies novel candidate genes for improving drought tolerance. These findings provide valuable insights into the genetic foundation of drought adaptation and offer potential targets for the development of drought-resistant maize cultivars.

The KERNEL NUMBER PER ROW6 (KNR6)-mediated phosphorylation of an adenosine diphosphate ribosylation factor (Arf) GTPase-activating protein (AGAP) forms a key regulatory module for the numbers of spikelets and kernels in the ear inflorescences of maize (Zea mays L.). However, the action mechanism of the KNR6–AGAP module remains poorly understood. Here, we characterized the AGAP-recruited complex and its roles in maize cellular physiology and agronomically important traits. AGAP and its two interacting Arf GTPase1 (ARF1) members preferentially localized to the Golgi apparatus. The loss-of-function AGAP mutant produced by CRISPR/Cas9 resulted in defective Golgi apparatus with thin and compact cisternae, together with delayed internalization and repressed vesicle agglomeration, leading to defective inflorescences and roots, and dwarfed plants with small leaves. The weak agap mutant was phenotypically similar to knr6, showing short ears with fewer kernels. AGAP interacted with KNR6, and a double mutant produced shorter inflorescence meristems and mature ears than the single agap and knr6 mutants. We hypothesized that the coordinated KNR6–AGAP–ARF1 complex modulates vegetative and reproductive traits by participating in vesicle trafficking in maize. Our findings provide a novel mechanistic insight into the regulation of inflorescence development, and ear length and kernel number, in maize.

Maize stalk rot, primarily caused by Fusarium spp., is a significant disease in many maize-growing areas worldwide, often leading to considerable yield losses under favorable conditions for infection. In this study, a genome-wide association study (GWAS) for Fusarium stalk rot resistance was carried out using 230,506 single nucleotide polymorphism (SNP) markers on phenotypic data from 178 maize inbred lines. Four SNPs were significantly associated with stalk rot resistance, with three located within previously reported QTL regions and one novel locus identified on chromosome 9 (9_149289012). Candidate genes within a 600-kb window around each SNP were retrieved from the MaizeGDB database, revealing 174 candidate genes in the B73_RefGen_v5 reference genome, including 8 R genes and 8 differentially expressed genes (DEGs). Notably, three potential candidate genes were identified around the novel SNP 9_149289012 on chromosome 9. Additionally, transcriptome analysis identified 1,597 genes related to disease resistance, associated with GO terms involved in resistance responses, nutrient metabolism, and toxin metabolism. Overall, this study provides valuable insights into the genetic basis of Fusarium stalk rot resistance in maize, highlighting both known and novel loci along with candidate genes for further functional characterization.