Transcriptome and Co-Expression Network Analysis Reveals Key Modules and Hub Genes Associated with Husk Number in Maize

Background: Husk number is a key agronomic trait in maize that affects grain dehydration rate and yield, yet its regulatory mechanisms remain largely unknown. This study aimed to identify gene modules and hub genes associated with husk number by integrating transcriptome profiling, hierarchical clustering, and weighted gene co-expression network analysis (WGCNA). Methods: RNA-seq was performed on ear tissues from two maize inbred lines with extreme husk numbers (LJ311, ~20 husks; PHG35, ~5 husks) at four developmental stages (V7–V10). A total of 12,605 putative husk-associated genes were identified. Differential expression analysis, hierarchical clustering, and WGCNA were combined to prioritize candidate genes. Results: Hierarchical clustering revealed a late-stage-specific gene cluster (Cluster 3, 44 genes) strongly up-regulated at V8–V10 in the high-husk line. WGCNA identified three modules significantly correlated with husk number: MEblue and MEbrown (positively) and MEturquoise (negatively). Cluster 3 was enriched in the tricarboxylic acid (TCA) cycle, DNA replication, and glycerophospholipid metabolism. Intersecting hub genes (|kME| > 0.9) with high-confidence differentially expressed genes (DEGs) (|log2FoldChange| > 2) yielded two prioritized candidate gene sets: up-regulated genes enriched in cell cycle, brassinosteroid (BR) signaling, and phospholipid metabolism; down-regulated genes enriched in mitogen-activated protein kinase (MAPK) signaling, glycolysis, and cell wall degradation. Conclusions: These results suggest that husk number may be regulated by a potential BR–MAPK–cell cycle regulatory axis, providing a regulatory network and candidate genes for future validation and molecular breeding.