Aquaporins are involved in CO₂ transport from the leaf intercellular air space to the chloroplast, which contributes to CO₂ assimilation. However, the mechanism of CO₂ transport by rice (Oryza sativa L.) aquaporins is unknown. Here, we investigated the function of the aquaporin OsPIP1;2 in CO₂ diffusion-associated photosynthesis and phloem sucrose transport. Moreover, the grain yield of rice lines overexpressing OsPIP1;2 was determined. OsPIP1;2 was localized to the plasma membrane and the relative expression of OsPIP1;2 was approximately 5-fold higher in leaves in the presence of an elevated CO₂ concentration. Overexpression of OsPIP1;2 increased mesophyll conductance by approximately 150% compared with wild-type (WT) rice. The OsPIP1;2-overexpressing lines had higher biomass than the WT, possibly due to increased phloem sucrose transport. In addition, the grain yield of OsPIP1;2-overexpressing lines was approximately 25% higher than that of the WT in three-season field experiments, due to the increased numbers of effective tillers and spikelets per panicle. Our results suggest that OsPIP1;2 modulates rice growth and grain yield by facilitating leaf CO₂ diffusion, which increases both the net CO₂ assimilation rate and sucrose transport.