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To understand the mechanisms of agricultural ecosystem structure and function in response to soil water deficit under future elevated atmospheric CO2 concentration, we examined the effects of soil water deficit and elevated CO2 concentration ([CO2]) on the stomatal traits, leaf photosynthesis, water use efficiency as well as Rubisco activity and gene expression of winter wheat with environmental growth chambers, whereby the [CO2] was controlled accurately with CO2 sensors. Our results showed that water deficit significantly decreased the plant biomass and net photosynthesis rates (Pn) of winter wheat by 33% and 29%, whereas elevated [CO2] partially mitigated the negative effects of water deficit on plant growth and physiological processes of winter wheat. Meanwhile, water deficit also reduced the stomatal width and regularity of stomatal distribution pattern on winter wheat leaves, but higher [CO2] could optimize the leaf gas exchange efficiency with more regular distribution pattern of stomata. Moreover, elevated [CO2] not only enhanced the Pn of winter wheat plants under water deficit, but also substantially reduced the transpiration rates (Tr) by 25%, and thus elevated [CO2] increased the water use efficiency by 61% when winter wheat plants subjected to water deficit. In addition, elevated [CO2] boosted the initial activity and activation state of Rubisco as well as soluble protein content by 66%, 38%, and 15%, and meanwhile significantly enhanced the gene expression levels of RbcL3 and RbcS2 by 453% and 417%, respectively. These results suggested that elevated [CO2] may optimize leaf gas exchange through modifying stomatal traits as well as the activity and gene express of Rubisco, and thus increased plant biomass, Pn, and water use efficiency to efficiently alleviate the physiological stress of water deficit on growth and development processes of winter wheat. Our findings may not only provide data for further understanding the impacts of water deficit on grain yield and water use efficiency of winter wheat under elevated [CO2], but also have important significance for adaptation management of agricultural ecosystems under global change. [ABSTRACT FROM AUTHOR]