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4D spatial transcriptomics reveals nodule identity emerges through stacked parallel developmental programs

2026/06/24 by Min-Yao Jhu, Jo Heffer, Chongjing Xia +8 · 1 voice
Agricultural and Biological Sciences · Energy · #Legume Nitrogen Fixing Symbiosis #Metalloenzymes and iron-sulfur proteins #Plant Molecular Biology Research

paper · pdf · doi:10.64898/2026.06.23.733685

Abstract

SUMMARY Legume root nodules enable symbiotic nitrogen fixation through the development of specialized cells that accommodate nitrogen-fixing bacteria intracellularly and support bacterial nitrogenase activity. Here, we present a 4D (3D space and time) spatial transcriptomic atlas of Medicago truncatula nodules and lateral roots, revealing specialized symbiotic cell types that develop alongside a conserved lateral-root-derived program that underpins vascularization. Spatial profiling of both plant and bacterial transcripts resolves distinct transcriptional states and previously unrecognized cell states. Spatial analysis of developmental regulator mutants uncovers a cascading series of cell-type-specific programs during nodule maturation. LSH1/LSH2 are central regulators of these programs, and lsh1/lsh2 mutants exhibit collapse of hormonal gradients and nodule identity. Strikingly, loss of nodule identity collapses to a primary-root identity rather than lateral-root fate. This work reveals how tissue complexity emerges through stacked developmental programs sustained in distinct cellular compartments, allowing the emergence of cell types specialized for harboring nitrogen-fixing bacteria. HIGHLIGHTS >136,000 cells define a 4D spatial atlas of nodules and lateral roots 3D spatial and dual-species analysis resolves dynamic host and rhizobial cell states A shared meristem generates 3 spatially coordinated symbiotic, non-symbiotic, and vascular cell programs LSH1/LSH2 are critical for regulatory programs underlying nodule development and identity Nodule identity loss collapses toward a primary root-associated state, not a lateral root fate

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