Grasslands underpin critical ecosystem services, yet their biodiversity and functioning are increasingly threatened by nitrogen (N) deposition and influenced by biomass removal through mowing to provide fodder. Trait-based approaches have advanced our understanding of plant responses, but many commonly used traits are plastic and context-dependent, limiting their predictive power. Here, we propose that plant genome size (GS), a relatively stable genetic character between individuals of a species, offers predictive power through nucleotypic constraints that are hypothesized to influence plant nutrient use, growth dynamics, and competitive strategies. By utilizing a large dataset of aboveground net primary productivity (ANPP) from an N addition and mowing field experiment over 15 years, we showed that GS–N interactions caused a selective stimulation of large-GS species under N enrichment, but only when N addition treatment exceeded a saturation threshold of 15 g N m⁻² year⁻¹. Conversely, low levels of N addition preferentially enhanced ANPP of small-GS plants. These divergent responses between two genomic groups, pivoting at the N saturation threshold, could be ascribed to a shift from competition for N to competition for light, as N availability increases. Meanwhile, biomass removal by mowing delayed the impact of this threshold by selectively benefiting small-GS species and disadvantaging large-GS species. This response could be due to the increased amount of canopy light penetration following biomass removal favoring small-GS species even under higher N availabilities. Additionally, large-GS species showed greater sensitivity of ANPP to interannual fluctuations in temperature and precipitation compared to small-GS species. The climate sensitivity of large-GS species was further amplified with increasing N treatment, reflecting a pervasive large genome constraint among grassland plants. Our findings pinpoint the co-regulation of GS, nutrients, and climate on plant growth, providing evidence that GS is a predictive character for ANPP change trajectories and community dynamics under the combined influence of global changes and management scheme shifts.