Chloride (Cl−) has been recently described as a beneficial macronutrient, playing specific roles in promoting plant growth and water use efficiency (WUE). However, it is still unclear how Cl− could be beneficial, especially in comparison with nitrate (NO3−), an essential macronutrient that shares with Cl− similar physical and osmotic properties, as well as common transport mechanisms. In tobacco plants, macronutrient levels of Cl− specifically reduce stomatal conductance (gs) without a concomitant reduction of the net photosynthesis rate (AN). Since stomata-mediated water loss through transpiration is inherent in the need of C3 plants to capture CO2, simultaneous increase of photosynthesis and WUE is of great relevance to achieve a sustainable increase of C3 crop productivity. Our results show that Cl−-mediated stimulation of larger leaf cells leads to a reduction of stomatal density, which in turn reduces gs and water consumption. On the other hand, Cl− improves mesophyll diffusion conductance to CO2 (gm) and photosynthetic performance due to a higher surface area of chloroplasts exposed to the intercellular airspace of mesophyll cells, possibly as a consequence of the stimulation of chloroplast biogenesis. A key finding of this study is the simultaneous improvement of AN and WUE due to macronutrient Cl− nutrition. This work identifies relevant and specific functions in which Cl− participates as a beneficial macronutrient for higher plants, uncovering a sustainable approach to improve crop yield.