The allochthonous supply of dissolved organic nitrogen (DON) from gyre margins into the interior of the ocean's oligotrophic subtropical gyres potentially provides an important source of new N to gyre surface waters, thus sustaining export production. This process requires that a fraction of the transported DON be available to euphotic zone photoautotroph communities via mineralization. In this study, we investigated the biological and physical controls on the distribution and fate of DON within global ocean surface waters. Inputs of nitrate to the euphotic zone at upwelling zones fuel net accumulation of a DON pool that appears to resist rapid microbial remineralization, allowing subsequent advective transport into the subtropical gyres. Zonal gradients in DON concentrations across these gyres imply a DON sink in the surface layer. Assessment of the physical dynamics of gyre circulation and winter mixing revealed a pathway for DON removal from the mixed layer via vertical transport to the deep euphotic zone, which establishes the observed zonal gradients. Incubation experiments from the Florida Straits indicated surface-accumulated DON was largely resistant (over a few months) to utilization by the extant surface bacterioplankton community. In contrast, this same material was remineralized three times more rapidly when exposed to upper mesopelagic bacterioplankton. These results suggest the primary fate of surface DON to be removal via vertical mixing and subsequent mineralization below the mixed layer, implying a limited role for direct DON support of gyre export production from the surface layer. DON may contribute to export production at the eastern edges of the subtropical gyres, but only after its mineralization within the deep euphotic zone.