Alajoleen, R. M.; Chau, T. N.; Shuman, J.; Bargmann, B.; Li, S.

Understanding how plant roots coordinate immune responses at the cellular level is key to unraveling host-pathogen interactions. Using single-nucleus RNA sequencing (snRNA-seq) of Arabidopsis thaliana roots 24 hours after Phytophthora. capsici (P. capsici) inoculation, we captured the transcriptional landscape of early infection at single-cell resolution. Four libraries (two infected and two mock-treated) were generated with approximately 26,000 high-quality nuclei with consistent sequencing depth and viability. A reference-based pipeline distinguished host and pathogen transcripts, enabling species-resolved mapping and host-focused single-nucleus transcriptomic analysis. Integration and clustering identified 12 transcriptionally distinct root cell types, encompassing major tissues such as the meristem, cortex, endodermis, and vasculature. Cluster-specific marker analysis confirmed cell-type identities, while differential expression and Gene Ontology enrichment revealed a global transcriptional shift from metabolic and translational processes in mock samples to defense, stress, and pathogen-response pathways upon infection. Hormone-related enrichment indicated broad salicylic acid activation across root tissues, spatially confined ethylene signaling in vascular-associated clusters, and localized jasmonic acid responses in cortex and phloem. Together, these results provide a high-resolution view of Arabidopsis root immunity, highlighting a coordinated yet tissue-specific defense architecture in which salicylic acid underpins systemic protection, ethylene modulates vascular defense, and jasmonic acid contributes targeted reinforcement during early P. capsici infection. Keywords: single-nucleus RNA sequencing, root immunity, cell type specific defense. hormone signaling, salicylic acid, jasmonic acid, ethylene signaling