Strong photosynthetic acclimation and enhanced water‐use efficiency in grassland functional groups persist over 21 years of CO 2 enrichment, independent of nitrogen supply

Uncertainty about long‐term leaf‐level responses to atmospheric CO 2 rise is a major knowledge gap that exists because of limited empirical data. Thus, it remains unclear how responses of leaf gas exchange to elevated CO 2 (eCO 2 ) vary among plant species and functional groups, or across different levels of nutrient supply, and whether they persist over time for long‐lived perennials. Here, we report the effects of eCO 2 on rates of net photosynthesis and stomatal conductance in 14 perennial grassland species from four functional groups over two decades in a Minnesota Free‐Air CO 2 Enrichment experiment, BioCON. Monocultures of species belonging to C 3 grasses, C 4 grasses, forbs, and legumes were exposed to two levels of CO 2 and nitrogen supply in factorial combinations over 21 years. eCO 2 increased photosynthesis by 12.9% on average in C 3 species, substantially less than model predictions of instantaneous responses based on physiological theory and results of other studies, even those spanning multiple years. Acclimation of photosynthesis to eCO 2 was observed beginning in the first year and did not strengthen through time. Yet, contrary to expectations, the response of photosynthesis to eCO 2 was not enhanced by increased nitrogen supply. Differences in responses among herbaceous plant functional groups were modest, with legumes responding the most and C 4 grasses the least as expected, but did not further diverge over time. Leaf‐level water‐use efficiency increased by 50% under eCO 2 primarily because of reduced stomatal conductance. Our results imply that enhanced nitrogen supply will not necessarily diminish photosynthetic acclimation to eCO 2 in nitrogen‐limited systems, and that significant and consistent declines in stomatal conductance and increases in water‐use efficiency under eCO 2 may allow plants to better withstand drought.