摘要Although root nodules are essential for bio-logical nitrogen fixation in legumes,the cell types and molecular regulatory mechanisms contributing to nodule development and ni-trogen fixation in determinate nodule legumes,such as soybean(Glycine max),remain in-completely understood.Here,we generated a single-nucleus resolution transcriptomic atlas of soybean roots and nodules at 14 days post inoculation(dpi)and annotated 17 major cell types,including six that are specific to nodules.We identified the specific cell types responsible for each step in the ureides synthesis pathway,which enables spatial compartmentalization of biochemical reactions during soybean nitrogen fixation.By utilizing RNA velocity analysis,we reconstructed the differentiation dynamics of soybean nodules,which differs from those of indeterminate nodules in Medi-cago truncatula.Moreover,we identified several putative regulators of soybean nod-ulation and two of these genes,GmbHLH93 and GmSCL1,were as-yet uncharacterized in soy-bean.Overexpression of each gene in soybean hairy root systems validated their respective roles in nodulation.Notably,enrichment for cytokinin-related genes in soybean nodules led to identification of the cytokinin receptor,GmCRE1,as a prominent component of the nodulation pathway.GmCRE1 knockout in soybean resulted in a striking nodule pheno-type with decreased nitrogen fixation zone and depletion of leghemoglobins,accompanied by downregulation of nodule-specific gene ex-pression,as well as almost complete abroga-tion of biological nitrogen fixation.In summary,this study provides a comprehensive per-spective of the cellular landscape during soybean nodulation,shedding light on the un-derlying metabolic and developmental mecha-nisms of soybean nodule formation.