Cyanate enhances methane-derived carbon assimilation in methanotrophs

Nitrogen availability strongly constrains soil methane oxidation, a major biological sink for atmospheric methane, yet the role of reactive nitrogen intermediates in methanotroph metabolism remains unresolved. Here, cyanate is identified as a previously overlooked nitrogen source that supports aerobic methanotrophy and promotes methane-derived carbon retention across multiple methanotrophic strains. Mechanistic analyses in Methylomonas sp. LW13 showed that cyanate utilization depends on coordinated transport, detoxification, and enzymatic hydrolysis to ammonium. 15 N tracing further demonstrated that cyanate-derived nitrogen was incorporated predominantly into biomass, with no detectable diversion to nitrification or denitrification products, and remained bioavailable under mixed-nitrogen conditions. Relative to ammonium, nitrate, and urea, cyanate increased biomass C/N ratios and cellular carbon content while reducing CO 2 release, consistent with enhanced retention of methane-derived carbon. Transcriptomic profiling revealed concerted upregulation of methane oxidation and central carbon metabolism during cyanate-dependent growth. Genomic and functional gene evidence further indicated that the capacity for cyanate utilization is widespread among aerobic methanotrophs. Together, these findings uncover cyanate as an underappreciated component of the soil nitrogen pool that links nitrogen acquisition to methane-derived carbon assimilation, thereby expanding current understanding of nitrogen control over terrestrial methane sinks.