Single‐Cell Transcriptomics Reveals FLS2‐Dependent Hypoxia Signaling and ERF13‐Mediated Transcription During flg22‐Triggered Immunity

ABSTRACT The flagellin peptide flg22 activates FLAGELLIN‐SENSING 2 (FLS2)‐mediated immunity in Arabidopsis, leading to growth inhibition and oxidative burst. While these responses are well‐studied, their cell‐type‐specific regulation remains poorly understood. Using single‐cell RNA sequencing, genetics, and phenotyping, we systematically mapped flg22‐induced responses. flg22 suppressed growth and elevated reactive oxygen species (ROS) in wild‐type, but not in fls2 mutants. Epidermal (EP_3) and mesophyll (MPC_2) cells showed FLS2‐dependent transcriptional reprogramming. Pseudotime analysis revealed developmental trajectories toward immune‐activated states. flg22 also induced a hypoxia‐like response; hypoxic signaling mutants ( ate1 , prt6 , zpr2 ) showed reduced flg22 sensitivity, indicating crosstalk between immune and hypoxia pathways. ERF13 was identified as a central regulator: erf13 mutants impaired flg22‐triggered ROS and growth inhibition but enhanced effector‐triggered immunity (ETI), while overexpressors showed stronger pattern‐triggered immunity (PTI). flg22 altered ploidy and cell‐cycle gene expression in WT, which was stabilized in ate1 and erf13 mutants. Cell‐cycle mutants sim smr and e2fabc enhanced flg22 responses, whereas cpr5 was less sensitive. Thus, immune, hypoxia, and ROS signals converge via ERF13 to balance immunity and growth, providing a single‐cell view of spatial immune organization and stress adaptation.