## 3 Discussion
Consistent with the high mass fraction of AsA found in young leaves (Figure 10B), gene expression of the D-mannose/L-galactose route indicated a remarkable activation, followed by a moderate enhancement of the L-glucose and D-galacturonate pathways in leaves at the vegetative stage (Figure 5A; Supplementary Table S5). Mutant analyses showed that D-mannose/L-galactose is dominant among the AsA biosynthetic pathways in photosynthetic tissues [55]. In addition, the intense increase in mRNA levels of organellar AsA-GSH cycle genes in the same experiments supports the biosynthetic requirement of AsA [10]. Indeed, the essential role of AsA in the vegetative phase has been observed in several plant species, and exogenous AsA application substantially improved biochemical and physiological parameters [56; 57]. Interestingly, the aging of the leaves changed the expression profile of the AsA biosynthesis gene, with a prevailing downregulation of D-mannose/L-galactose genes and upregulation of specific genes from the L-glucose (_GullO_1f_ and _lg_), D-galacturonate (_GalUIR_2, 4 and 5), and Myo-inositol (_MIOX_1a_ and _1b_) pathways (Figure 5A; Supplementary Table S5). These findings support the decreased AsA mass fractions observed across the developmental stages of leaves (Figure 10). Cotyledon senescence maintained this tendency but with the upregulation of specific genes such as _GMP_1a_, _2a_, _2b_, and all _GGP_ of the D-mannose/L-galactose pathways (Figure 5B; Supplementary Table S5). The upregulation of Myo-inositol genes in aged tissues presented a remarkable difference compared to leaves at the vegetative stage (Figure 3). A decrease in Myo-inositol biosynthesis in apple leaves was associated with extensive programmed cell death [58]. Hence, the activation of the Myo-inositol AsA biosynthesis pathway in aging _G. max_ tissues could contribute to controlling Myo-inositol and may have a minor contribution to increasing AsA synthesis during senescence. In stress conditions, a similar tendency was observed during senescence, stimulating/inducing the gene expression of the L-glucose, D-galacturonate, and Myo-inositol pathways. This was remarkable, with substantial increases in mRNA levels or a strong induction of specific genes. However, stress responses varied based on the type of stress, duration of exposure, and plant tissue types. Depending on stress, specific D-mannose/L-galactose genes were also upregulated or occurred with significant downregulation but always maintained basal mRNA levels (Figures 7-9; Supplementary Tables S6 and S7). This general expression pattern supports the conclusion that the D-mannose/L-galactose pathway could function as the major and constitutive AsA biosynthesis pathway. In contrast, the other three AsA pathways could be complementary and indispensable pathways in response to development and environmental fluctuations. Given the vital importance of AsA for plant metabolism, it is essential to take into account that stresses may reduce the AsA mass fraction in plant tissues, affecting the efficiency of the D-mannose/L-galactose pathway and/or the increased ROS scavenging activity using AsA [59]. In this regard, the downregulation of key genes from the D-mannose/L-galactose pathway was consistent with the reduced AsA mass fractions in leaves and roots under PEG and NaCl conditions (Figures 10 and 11). In addition, the upregulation of AsA-GSH cycle genes under the same experiments from the current study supports the idea of increased AsA consumption for ROS scavenging activities [10]. In former studies, our group observed an extensive upregulation of specific genes encoding APX from different cell compartments in response to the same abiotic and biotic stresses in _G. max_ that were investigated here [10]. In this scenario, it is possible to assume that alternative AsA biosynthesis pathways could be activated to attenuate AsA decrease or maintain or increase AsA levels in plant cells.