Nitrogen (N) deficiency has been recorded in the top surface of Tibetan Plateau. However, the variation of soil N availability across the elevational gradient in alpine forests remains poorly understood. Here, the elevational patterns and determinants of soil N composition, key N transformation processes and functional microbes across three typical mountains on the southeastern Tibetan Plateau were characterized by multiple techniques. Our results showed that soil total N and ammonium were markedly enriched in high elevation zones where a stable N release via mineralization and extremely low net nitrification were observed. Further, the increasing biological N fixation rates along the elevation driven by abiotic (i.e., high organic carbon) and biotic (i.e., key diazotrophic taxa like Bradyrhizobium, Herbaspirillum and Klebsiella) factors greatly benefited N accumulation at high elevations. Our study offers new insights into the N dynamics in alpine forests on the Tibetan Plateau under scenarios of future climate change.