## 4 Discussion  
Drought stress reduced the leaf RWC in both the NT and transformed plants, where the transgenic wheat lines performed differently than the NT plants. The transgenic lines maintained their RWC more efficiently, as it was higher after the eighth day of drought than that of the NT plants (Table 1). These data suggest that the overexpression of the _ScTPS1-TPS2_ gene aided in retaining their RWC when the soil water diminished. In the NT plants, the transpiration rate decreased during drought (Figure 2c), indicating that stomatal closure occurred. This result explains the decrease in the photosynthetic rate (Figure 2a). However, the _Ci_ in the NT plants did not change during drought (Figure 2b), suggesting that the reduction in the photosynthetic rate was more affected by non-stomatal factors than by a decreased gas interchange in the leaf. One of these factors could be Rubisco activity, which declined for the NT plants under drought stress (Figure 3a). Although the NT plants showed a recovery in their photosynthetic rate on day eight of drought (Figure 2a), a definitive decline in photosynthesis could be expected if the drought treatment were to be extended further [31]. In the transgenic line 35S, the photosynthetic rate and _Ci_ declined on the sixth day of drought (Figure 2a), but the transpiration (Figure 2c) and Rubisco activity (Figure 3a) data denote that the decline in the photosynthetic rate was not due to a reduced CO2 uptake. We observed that the constitutive overexpression of the TPS-TPP bifunctional enzyme in the 35S line caused a decreased activity of FBPase compared to the NT plants and the rd-1B and rd-1C lines (Figure 3b). This suggests that triose phosphates synthesized in the Calvin cycle in the 35S line were not directed towards RuBP regeneration, which could explain the decreased photosynthetic rate observed in this line (Figure 3a). The gas exchange parameters (A, _Ci_, and E) were less affected by drought in plants transformed with the inducible rd29A promoter, as both lines kept a higher photosynthetic rate on the eighth day of drought (Figure 2). The rd-1B plants and rd-1C plants maintained their CO2 uptake during drought, which aided in sustaining the photosynthetic rate (Figure 2). Moreover, the rd-1B and rd-1C lines maintained a higher Rubisco activity under drought conditions than the NT plants (Figure 3a). A higher FBPase activity was also observed in these transgenic lines on the sixth day of drought (Figure 3b), suggesting that RuBP regeneration was sustained and contributed to keeping a high photosynthetic rate during drought (Figure 2a). The photosynthesis data in this work agree with other studies, in which the overexpression of a bifunctional bacterial TPS-TPP in rice avoids stomata closure when subjected to drought and salinity stress, promoting a higher photosynthetic rate [32]. Foliar trehalose application avoids stomata closure, increases photosynthesis during abiotic stress, and protects the chloroplast structure [33; 34]. Although trehalose metabolism seems involved in controlling the stomata aperture and the activation of various stress-response pathways, the precise mechanism for protecting photosynthesis during abiotic stress is still unclear [35]. Transcriptomic and proteomic approaches have found that trehalose application upregulates Rubisco and chloroplast FBPase in wheat, even under non-stressed conditions [36; 37]. In this work, the three lines transformed with the _ScTPS1-TPS2_ gene had higher Rubisco activity during drought stress than non-transformed wheat plants (Figure 4a), implying that increased trehalose synthesis in wheat promotes better photosynthesis because it helps maintain an efficient CO2 fixation under drought conditions. The activity of SPS declined in the transgenic wheat lines on day eight of drought (Figure 3c). This may have been caused by the increased demand for glucose and UDP-glucose for trehalose synthesis under stress, induced by the transformation with the _ScTPS1-TPS2_ gene.