Effects of growth light intensity on the temperature dependence of CO2 assimilation rate were studied in tobacco (Nicotiana tabacum) because growth light intensity alters nitrogen allocation between photosynthetic components. Leaf nitrogen, ribulose 1·5-bisphosphate carboxylase/oxygenase (Rubisco) and cytochrome f (cyt f) contents increased with increasing growth light intensity, but the cyt f/Rubisco ratio was unaltered. Mesophyll conductance to CO2 diffusion (gm) measured with carbon isotope discrimination increased with growth light intensity but not with measuring light intensity. The responses of CO2 assimilation rate to chloroplast CO2 concentration (Cc) at different light intensities and temperatures were used to estimate the maximum carboxylation rate of Rubisco (Vcmax) and the chloroplast electron transport rate (J). Maximum electron transport rates were linearly related to cyt f content at any given temperature (e.g. 115 and 179 µmol electrons mol−1 cyt f s−1 at 25 and 40 °C, respectively). The chloroplast CO2 concentration (Ctrans) at which the transition from RuBP carboxylation to RuBP regeneration limitation occurred increased with leaf temperature and was independent of growth light intensity, consistent with the constant ratio of cyt f/Rubisco. In tobacco, CO2 assimilation rate at 380 µmol mol−1 CO2 concentration and high light was limited by RuBP carboxylation above 32 °C and by RuBP regeneration below 32 °C.