Nitrogen fixation, the microbial reduction of dinitrogen to ammonia, is increasingly recognized to occur in the Arctic Ocean. However, knowledge about the composition, biogeography, abundance, and ecology of nitrogen-fixing organisms (diazotrophs) is poor. This ultimately hinders the prediction of ecosystem productivity fueled by nitrogen fixation in this rapidly changing and predominantly nitrogen-limited ocean. We assessed the composition and abundance of total and nifH-expressing diazotrophs in subsurface water (8 m; amplicon sequencing and quantification of the marker gene nifH) over similar to 3400 km from the mouth of the brackish Baltic Sea to the sea ice edge in the Arctic Ocean. Upon entering nutrient-rich waters in the Atlantic gateway to the Arctic, we discovered an abrupt transition from autotrophic to heterotrophic diazotrophy (nifH expression). Our findings therefore suggest that diazotrophy is functionally distinct in the Arctic Ocean compared to adjacent temperate-boreal waters-a difference likely driven by inorganic nutrients, salinity, and temperature. We identify three key non-cyanobacterial diazotroph groups in the Arctic Ocean with Arctic-specific (Rhodocyclales and Oceanospirillales) or more widespread (unknown Gammaproteobacterium) distribution patterns and report their nifH gene transcription levels (up to 10³ nifH transcripts L^-1). In contrast, nifH expression in the warmer and more nutrient-poor Norwegian Sea with coastal-influenced water was dominated by sublineages of Candidatus Atelocyanobacterium thalassa (UCYN-A1, UCYN-A2, UCYN-A4; up to 10⁴ nifH transcripts L^-1). With ongoing atlantification of the Arctic pushing oceanic provinces and biogeographical ranges poleward, we predict a future displacement of the transition from autotrophic to heterotrophic diazotrophy with likely significant changes in nitrogen fixation.