Abstract text: Root development relies on the coordination of longitudinal cell division, expansion, and maturation. This process is modulated by signaling networks involving peptide ligand-receptor interactions and reactive oxygen species (ROS) homeostasis, and is tightly linked to dynamic cell-wall remodeling. However, these regulatory modules remain largely unconnected, and the systematic mechanisms orchestrating root development remain elusive. Here, we identify an intrinsic regulatory hub in rice that orchestrates root development by linking ROS transmembrane signaling to pectin structural organization. THESEUS3, a receptor-like kinase, interacts with and phosphorylates RbohB to stimulate superoxide anion (O₂⁻) production, thereby negatively regulating pectin demethylesterification in the root meristem. Rice RAPID ALKALINIZATION FACTOR4 (OsRALF4) acts as a ligand that maintains O₂⁻ homeostasis by repressing THESEUS3-RbohB signaling in the meristem, thereby modulating pectin demethylesterification. Demethylesterified pectin further interacts with OsRALF4 to mediate pectin bundling into a condensed structure, which marks the initiation of cell expansion. Collectively, our work establishes a unified mechanistic framework for root development regulation.