Integrated Single‐Cell and Spatial Transcriptomics Reveal Cell‐Type‐Specific Immune Regulatory Networks in Maize Responding to Southern Corn Rust

Southern corn rust (SCR), caused by Puccinia polysora Underw. (P. polysora), poses a significant threat to maize production, yet the cell-type-specific defense mechanisms remain insufficiently characterized. To address this, we integrated single-nucleus RNA sequencing (snRNA-seq) and spatial transcriptomic sequencing (stRNA-seq) to elucidate the cell-type-specific transcriptional dynamics in maize leaves during early infection with P. polysora. Analyses at 24 and 48 h post infection (hpi) revealed eight major cell types and highlighted key defense responses, which are primarily initiated in the mesophyll and epidermal cells 24 hpi. Notably, the cell-type-specific activation of RLPs/RLKs and jasmonic acid was observed. Functional defense modules were activated in specific cell types at 24 hpi, with pseudotime and cell-cell communication analyses further uncovering immune-related cellular dynamics. Importantly, multi-omics analysis identified core DEGs across critical cell types and time points. Functional validation through virus-induced gene silencing (VIGS) demonstrated that silencing the ZmXET1 gene significantly reduced disease severity and pathogen biomass, while silencing the positive regulator ZmRBG, increased susceptibility. This study provides a high-resolution spatiotemporal atlas of maize defense against P. polysora, identifying ZmXET1 as a key susceptibility factor and ZmRBG as a resistance component, thereby offering valuable targets for disease resistance breeding.