To investigate the response of a dominant freshwater diatom to changes in nutrient stoichiometry, Cyclotella meneghiniana isolated from the Chaiwen River, an important tributary of the lower Yellow River, and Dongping Lake, a shallow eutrophic freshwater lake in northern China, was used as the study species. Indoor culture experiments were conducted under four nitrogen-to-phosphorus (N:P) ratios, namely 4:1, 8:1, 16:1, and 64:1. By combining nutrient monitoring with integrated transcriptomic and proteomic analyses, we investigated the physiological and molecular responses of Cyclotella meneghiniana to different N:P conditions. The results showed that lower N:P ratios were associated with higher biomass accumulation and faster nitrogen and phosphorus consumption, whereas the 64:1 treatment showed lower final biomass and reduced nutrient utilization efficiency. Integrated transcriptomic and proteomic analyses further showed that the high-N:P treatment exhibited a more distinct transcriptional profile, whereas proteomic responses were comparatively more dispersed, suggesting partial transcript–protein decoupling. GO and KEGG enrichment analyses suggested that different N:P ratios were mainly associated with changes in broadly conserved processes related to protein synthesis, energy metabolism, and cellular homeostasis, including ribosome-related functions, protein processing, oxidative phosphorylation, and glyoxylate metabolism. This study provides new insight into the physiological and molecular adaptation of a dominant freshwater diatom to altered nitrogen and phosphorus stoichiometry, and offers a basis for understanding how nutrient imbalance may influence algal responses in eutrophic waters.