Modulation of antioxidant capacities according to features of the high temperature sequences
Heat stress is known to trigger oxidative bursts that lead to a wide spectrum of responses including enzymatic and non-enzymatic components such as polyphenols, hormones and sugars, which have been shown to scavenge ROS (Nishizawa, Yabuta & Shigeoka 2008; Soengas, Rodríguez, Velasco & Cartea 2018; Serrano, Ling, Bahieldin & Mahfouz 2019; Soares, Carvalho, Azevedo & Fidalgo 2019). In our study, measurements of the antioxidant capacity indicated complex results that differed according to the S supply. In the non-limiting S conditions the greatest increase was not observed under the CES or late heat peaks, but it occurred following the EMS event. These observations correlated with seed polyphenol concentrations, which displayed similar rankings. However, other studies have reported the enhancement of subcellular antioxidant activities using several antioxidant enzymes as proxies (superoxide dismutase, glutathione reductase, and peroxidase in mitochondria and chloroplasts of wheat leaves) under multiple heat priming sequences prior to later high temperatures (Wang et al.2014), which might not be observed with the antioxidant capacity tests used herein. In addition, the concentrations of raffinose and stachyose, which have been reported to protect against oxidative damage (Nishizawaet al. 2008), were not increased under EMS under either of the S conditions, and so it was not possible to ascertain their role in the antioxidant defense pathways. However, this observation should be interpreted carefully regarding the timing and optimal biosynthesis of sugars within the maturing seeds, as previously discussed (Baud, Boutin, Miquel, Lepiniec & Rochat 2002; Leprince, Pellizzaro, Berriri & Buitink 2017). Indeed, our results might suggest that the temperature sequences (whether mild or intense) were above the temperature threshold of the stepwise transfer reactions involving raffinose synthase and stachyose synthase, leading to impairment of sugar biosynthesis (Gangl, Behmüller & Tenhaken 2015). Nevertheless, because the EMS had little to no negative impacts on the seed quality criteria (Tables 2 and 3) and it triggered the highest levels of antioxidant capacity and polyphenols and low levels of RFOs, the induced antioxidant response to the mild stress might rely on an effective stress response mediated by specific components of the non-enzymatic antioxidant machinery (mainly dealing with polyphenols and not sugars).