The principal function of isoprene biosynthesis in plants remains unclear, but emission rates are positively correlated with temperature and light, supporting a role for isoprene in maintaining photosynthesis under transient heat and light stress from sunflecks. Isoprene production is also inversely correlated with CO(2) concentrations, implying that rising CO(2) may reduce the functional importance of isoprene. To understand the importance of isoprene in maintaining photosynthesis during sunflecks, we used RNAi technology to suppress isoprene production in poplar seedlings and compared the responses of these transgenic plants to wild-type and empty-vector control plants. We grew isoprene-emitting and non-emitting trees at low (190 ppm) and high (590 ppm) CO(2) concentrations and compared their photosynthetic responses to short, transient periods of high light and temperature, as well as their photosynthetic thermal response at constant light. While there was little difference between emitting and non-emitting plants in their photosynthetic responses to simulated sunflecks at high CO(2), isoprene-emitting trees grown at low CO(2) had significantly greater photosynthetic sunfleck tolerance than non-emitting plants. Net photosynthesis at 42°C was 50% lower in non-emitters than in isoprene-emitting trees at low CO(2), but only 22% lower at high CO(2). Dark respiration rates were significantly higher in non-emitting poplar from low CO(2), but there was no difference between isoprene-emitting and non-emitting lines at high CO(2). We propose that isoprene biosynthesis may have evolved at low CO(2) concentrations, where its physiological effect is greatest, and that rising CO(2) will reduce the functional benefit of isoprene in the near future.