The diazotrophic cyanobacteria Trichodesmium spp. contribute approximately half of the known marine dinitrogen (N2) fixation. Rapidly changing environmental factors such as the rising atmospheric partial pressure of carbon dioxide (pCO2) and shallower mixed layers (higher light intensities) are likely to affect N2-fixation rates in the future ocean. Several studies have documented that N2 fixation in laboratory cultures of T. erythraeum increased when pCO2 was doubled from present-day atmospheric concentrations (∼380 ppm) to projected future levels (∼750 ppm). We examined the interactive effects of light and pCO2 on two strains of T. erythraeum Ehrenb. (GBRTRLI101 and IMS101) in laboratory semicontinuous cultures. Elevated pCO2 stimulated gross N2-fixation rates in cultures growing at 38 μmol quanta · m−2 · s−1 (GBRTRLI101 and IMS101) and 100 μmol quanta · m−2 · s−1 (IMS101), but this effect was reduced in both strains growing at 220 μmol quanta · m−2 · s−1. Conversely, CO2-fixation rates increased significantly (P < 0.05) in response to high pCO2 under mid- and high irradiances only. These data imply that the stimulatory effect of elevated pCO2 on CO2 fixation and N2 fixation by T. erythraeum is correlated with light. The ratio of gross:net N2 fixation was also correlated with light and trichome length in IMS101. Our study suggests that elevated pCO2 may have a strong positive effect on Trichodesmium gross N2 fixation in intermediate and bottom layers of the euphotic zone, but perhaps not in light-saturated surface layers. Climate change models must consider the interactive effects of multiple environmental variables on phytoplankton and the biogeochemical cycles they mediate.