A greater understanding of natural variation in photosynthesis will inform strategies for crop improvement by revealing overlooked opportunities. We use Arabidopsis thaliana ecotypes as a model system to assess (i) how photosynthesis and photorespiration covary and (ii) how mesophyll conductance influences water use efficiency (WUE). Phenotypic variation in photorespiratory CO2 efflux was correlated with assimilation rates and two metrics of photosynthetic capacity (i.e. VCmax and Jmax); however, genetic correlations were not detected between photosynthesis and photorespiration. We found standing genetic variation-as broad-sense heritability-for most photosynthetic traits, including photorespiration. Genetic correlation between photosynthetic electron transport and carboxylation capacities indicates that these traits are genetically constrained. Winter ecotypes had greater mesophyll conductance, maximum carboxylation capacity, maximum electron transport capacity, and leaf structural robustness when compared with spring ecotypes. Stomatal conductance varied little in winter ecotypes, leading to a positive correlation between integrated WUE and mesophyll conductance. Thus, variation in mesophyll conductance can modulate WUE among A. thaliana ecotypes without a significant loss in assimilation. Genetic correlations between traits supplying energy and carbon to the Calvin-Benson cycle are consistent with biochemical models, suggesting that selection on either of these traits would improve all of them. Similarly, the lack of a genetic correlation between photosynthesis and photorespiration suggests that the positive scaling of these two traits can be broken.