The surface mixed layer (ML) in the Mediterranean Sea is a well stratified domain characterized by low macro-nutrient and low chlorophyll content, during almost 6 months of the year. Nutrient dynamics in the ML depend on allochthonous inputs, through atmospheric deposition and on biological recycling. Here we characterize the biogeochemical cycling of N in the ML by combining simultaneous in situ measurements of atmospheric deposition, nutrients, hydrological conditions, primary production, heterotrophic prokaryotic production, N2 fixation and leucine aminopeptidase activity. The measurements were conducted along a 4300 km transect across the central and western open Mediterranean Sea in spring 2017. Dry deposition was measured on a continuous basis while two wet deposition events were sampled, one in the Ionian Sea and one in the Algerian basin. Along the transect, N budgets were computed to compare sources and sinks of N in the mixed layer. On average, phytoplankton N demand was 2.9 fold higher (range 1.5–8.1) than heterotrophic prokaryotic N demand. In situ leucine aminopeptidase activity contributed from 14 to 66 % of heterotrophic prokaryotic N demand, and N2 fixation rate represented 1 to 4.5 % of the phytoplankton N demand. Dry atmospheric deposition of inorganic nitrogen, estimated from dry deposition of (nitrate + ammonium) in aerosols, was higher than N2 fixation rates in the ML (on average 4.8 fold). The dry atmospheric input of inorganic N represented a highly variable proportion of biological N demand in the ML, 10–82 % for heterotrophic prokaryotes and 1–30 % for phytoplankton. Stations visited for several days allowed following the evolution of biogeochemical properties in the ML and within the nutrient depleted layers. At the site in the Algerian Basin and on a basis of high frequency sampling of CTD casts before and after a wet dust deposition event, different scenarios were considered to explain a delayed appearance of peaks in dissolved inorganic phosphate in comparison to nitrate within the ML. After the rain, nitrate was higher in the ML than in the nutrient depleted layer below. Estimates of nutrient transfer from the ML to the nutrient depleted layer could explain 1/3 of the nitrate fate out of the ML. Luxury consumption of P by heterotrophic prokaryotes, further transferred in the microbial food web, and remineralized by grazers, is one explanation for the delayed phosphate peak of DIP. The second explanation is a transfer from ML to the nutrient depleted layer below through adsorption/desorption processes on particles. Phytoplankton did not benefit directly from atmospheric inputs in the ML, probably due to a high competition with heterotrophic prokaryotes, also limited by N and P availability at the time of this study. Primary producers, in competition for nutrients with heterotrophic prokaryotes, decreased their production after the rain, recovering their initial state of activity after 2 days lag in the vicinity of the deep chlorophyll maximum layer.