Semi-closed greenhouses have been developed in which window ventilation is minimized due to active cooling, enabling enhanced CO2 concentrations at high irradiance. Cooled and dehumidified air is blown into the greenhouse from below or above the canopy. Cooling below the canopy may induce vertical temperature gradients along the length of the plants. Our first aim was to analyze the effect of the positioning of the inlet of cooled and dehumidified air on the magnitudes of vertical temperature and VPD gradients in the semi-closed greenhouses. The second aim was to investigate the effects of vertical temperature gradients on assimilate production, partitioning, and fruit growth. Tomato crops were grown year-round in four semiclosed greenhouses with cooled and dehumidified air blown into the greenhouses from below or above the crop. Cooling below the canopy induced vertical temperature and VPD gradients. The temperature at the top of the canopy was over 5°C higher than at the bottom, when outside solar radiation was high (solar radiation >250 J cm-2 h-1). Total dry matter production was not affected by the location of the cooling (4.64 and 4.80 kg m-2 with cooling from above and from below, respectively). Percentage dry matter partitioning to the fruits was 74% in both treatments. Average over the whole growing season the fresh fruit weight of the harvested fruits was not affected by the location of cooling (118 vs 112 g fruit-1). However, during summer period the average fresh fruit weight of the harvested fruits in the greenhouse with cooling from below was higher than in the greenhouse with cooling from above (124 vs 115 g fruit-1).