tae-miR399-UBC24 module regulates the ROS metabolism
response to freezing stress in Arabidopsis
Low temperature stress can significantly disturb ROS homeostasis in
plants, leading to lipid peroxidation, damage to membranes (Ning, Li,
Hicks, & Xiong, 2010) and increasing MDA content (J. J. Liu, Wei, &
Li, 2014). In this study, under low temperature stress, OEmiR399
Arabidopsis had lower MDA content, indicating that it suffered less
oxidative damage than WT. Further histochemical staining and
physiological experiment methods to detect ROS content, the results
showed that the H2O2 content and
O2.- production of OEmiR399
Arabidopsis plants were visibly lower than that of WT under freezing
stress (Fig. 11). Also, under freezing stress, the SOD, POD and CAT
enzyme activities of OEmiR399 Arabidopsis plants were significantly
higher than those of WT (Fig. 12, 13 and 14). These implies that
OEmiR399 Arabidopsis plants have a strong ROS scavenging ability.
Interestingly, the expression changes of the genes encoding SOD, POD and
CAT in OEmiR399 Arabidopsis plants were not consistent with the enzyme
activities. The expression of the most genes encoding SOD, POD and CAT
in OEmiR399 Arabidopsis plants was significantly up-regulated at 24°C
and 4°C, but not significantly changed or down-regulated at -10°C (Fig.
12, 13 and 14). Similary, the expression of genes encoding SOD and APX
in bluegrass are significantly down-regulated under drought stress,
while their enzyme activities are significantly up-regulated (Xu, Han,
& Huang, 2011). Based on previous studies, we speculate that the
changes in the enzyme activity of OEmiR399 Arabidopsis plants under
freezing stress may not be caused by mRNA levels, but are regulated at
the post-transcriptional level, which is due to the inhibition of enzyme
inactivation/activation or degradation/synthesis caused by freezing
stress. Homologous or heterologous overexpressing ICE1 in plants
such as tomato and tobacco can increase its antioxidant enzyme activity
to enhance the cold tolerance of plants (Feng et al., 2013; Luo et al.,
2020). Therefore, in this study, under low temperature stress, the high
ICE1 protein levels of OEmiR399 Arabidopsis plants may also be one of
the reasons for its stronger ROS scavenging ability.
Combining with our phenotypic data, we can conclude that
miR399-UBC24 module inhibit the degradation of ICE1 and promote
the CBF signalling pathway and ROS clearance to initiate plant response
to freezing stress. On the other hand, miR399-UBC24 module can
accelerate starch degradation to accumulate a large amount of soluble
sugar by promoting the Pi absorption and utilization efficiency, and
then enhance freezing tolerance in plants (Fig. 14). These findings
clarify the function of miR399-UBC24 module and suggest that
tae-miR399-TaUBC24 can potentially be used to improve freezing
tolerance in crops.