Emerging evidence points out that the responses of soil organic carbon (SOC) to nitrogen (N) addition differ along the soil profile, highlighting the importance of synthesizing results from different soil layers. Here, using a global meta-analysis, we found that N addition significantly enhanced topsoil (0–30 cm) SOC by 3.7% (±1.4%) in forests and grasslands. In contrast, SOC in the subsoil (30–100 cm) initially increased with N addition but decreased over time. The model selection analysis revealed that experimental duration and vegetation type are among the most important predictors across a wide range of climatic, environmental, and edaphic variables. The contrasting responses of SOC to N addition indicate the importance of considering deep soil layers, particularly for long-term continuous N deposition. Finally, the lack of depth-dependent SOC responses to N addition in experimental and modeling frameworks has likely resulted in the overestimation of changes in SOC storage under enhanced N deposition.