Estuarine and coastal systems are global hotspots of marine nitrous oxide (N₂O) emissions, where microbial nitrification and denitrification are the primary processes regulating N₂O dynamics. However, how interactions among different N2O-associated microorganisms influence ecosystem-scale N₂O emissions remains poorly understood. This study combined in situ N₂O concentrations, ¹⁵N-based potential rates, metagenomics, metatranscriptomics, and genome-scale metabolic model analysis to explore N₂O production and reduction processes in estuarine and coastal systems. Potential N₂O production and reduction rates, together with in situ concentrations, the relative abundance, and the transcriptional activity of associated genes, were significantly higher at low salinity and declined toward coastal regions. Based on the gene content of 974 recovered N₂O-associated genomes, microorganisms were classified into three functional groups: net N₂O producers, net N₂O consumers, and self-sustaining N₂O players. The abundance, composition, and activity of these functional groups shifted along estuarine-coastal gradients. A larger exchange gap between NO and N₂O was found at low salinity, which contributed to elevated N₂O emissions. Taken together, our results showed that community-level division of labour and the resulting imbalance of NO/N₂O exchange potential provide a conceptual framework linking N₂O-related functional groups to N₂O accumulation in estuarine-coastal ecosystems.

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