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Summary Affecting biodiversity, plants with larger genome sizes (GS) may be restricted in nutrient‐poor conditions. This pattern has been attributed to their greater cellular nitrogen (N) and phosphorus (P) investments and hypothesized nutrient–investment tradeoffs between cell synthesis and physiological attributes associated with growth. However, the influence of GS on cell size and functioning may also contribute to GS‐dependent growth responses to nutrients.To test whether and how GS is associated with cellular nutrient, stomata, and/or physiological attributes, we examined > 500 forbs and grasses from seven grassland sites conducting a long‐term N and P fertilization experiment.Larger GS plants had increased cellular nutrient contents and larger, but fewer stomata than smaller GS plants. Larger GS grasses (but not forbs) also had lower photosynthetic rates and water‐use efficiencies. However, nutrients had no direct effect on GS‐dependent physiological attributes and GS‐dependent physiological changes likely arise from how GS influences cells. At the driest sites, large GS grasses displayed high water‐use efficiency mostly because transpiration was reduced relative to photosynthesis in these conditions.We suggest that climatic conditions and GS‐associated cell traits that modify physiological responses, rather than resource–investment tradeoffs, largely explain GS‐dependent growth responses to nutrients (especially for grasses).