Exogenous spermine pretreatment confers tolerance to combined high-temperature and drought stress in vitro in trifoliate orange seedlings via modulation of antioxidative capacity and expression of stress-related genes

Spermine (Spm) is thought to play an important role in drought or high-temperature (HT) tolerance. However, it is not clear whether Spm confers similar resistance in the presence of both drought and HT, which often occur simultaneously. In the present study, the trifoliate orange (Poncirus trifoliata (L.) Raf.) seedlings were pretreated with 1 mmol L-1 Spm to evaluate their tolerance to combined drought and HT (45 ºC) stress. Spm-pretreated seedlings showed less leaf wilting, less water loss and less electrolyte leakage than control leaves not treated with Spm within 180 min of treatment. Histochemical staining with diaminobenzidine and nitro blue tetrazolium showed that Spm-pretreated seedlings accumulated less hydrogen peroxide and superoxide than those of control plants 60, 120 and 180 min after treatment when exposed to both drought and HT (45 ºC). However, superoxide dismutase, peroxidase and catalase were significantly more active in Spm-pretreated seedlings than in control seedlings. In addition, Spm-pretreated seedlings showed significantly higher expression of heat shock proteins, abscisic acid (ABA)-responsive element binding factor and 9-cis-epoxycarotenoid dioxygenase 3 than controls either before (0 min) or after (60, 120 and 180 min) combined drought and HT treatment. All of these data suggest that exogenous Spm pretreatment confers tolerance to simultaneously occurring drought and HT stresses. Spm may influence this by activating antioxidant enzymes, increasing the effectiveness of scavenging of reactive oxygen species. It may also increase the expression levels of stress-related genes that protect trifoliate orange seedlings from stress damage.