Alterations in climate factors such as rising CO2 concentration ([CO2]), warming and reduced precipitation may have significant impacts on plant physiology and growth. This research investigated the interactive effects of elevated [CO2], warming and soil water deficit on biomass production, leaf-level physiological responses and whole-plant water use efficiency (WUEP) in cotton (Gossypium hirsutum L.). Cotton was grown in the glasshouse under two [CO2] treatments (CA, 400µLL-1; CE, 640µLL-1) and two temperature treatments (TA, 28°C:17°C day:night; TE, 32°C:21°C day:night). Plants were subjected to two progressive water deficit cycles, with a 5-day recovery period between the water deficit periods. CE increased vegetative biomass and photosynthetic rates, and decreased stomatal conductance in TA; however, these responses to CE were not evident under TE. CE increased whole-plant water loss under TA, but increased WUEp, whereas increased whole-plant water loss in TE decreased WUEp regardless of atmospheric [CO2]. CE may provide some positive growth and physiological benefits to cotton at TA if sufficient water is available but CE will not mitigate the negative effects of rising temperature on cotton growth and physiology in future environments.