Starch-related α-glucan/water dikinase is involved in the cold-induced development of freezing tolerance in arabidopsis

Cold- induced soluble sugar accumulation enhances the degree of freezing tolerance in various cold- hardy plants including Arabidopsis ( Arabidopsis thaliana), where soluble sugars accumulate in only a few hours at 2 degrees C. Hence, along with photosynthesis, starch degradation might play a significant role in cold- induced sugar accumulation and enhanced freezing tolerance. Starch- related alpha- glucan/ water dikinase ( EC 2.7.9.4), encoded by Arabidopsis STARCH EXCESS 1 ( SEX1), is hypothesized to regulate starch degradation in plastids by phosphorylating starch, thereby ensuring better accessibility by starch- degrading enzymes. Here, we show that Arabidopsis sex1 mutants, when incubated at 2 degrees C for 1 d, were unable to accumulate maltooligosaccharides or normal glucose and fructose levels. In addition, they displayed impaired freezing tolerance. After 7 d at 2 degrees C, sex1 mutants did not show any of the above abnormal phenotypes but displayed slightly higher leaf starch contents. The impaired freezing tolerance of sex1 mutants was restored by overexpression of wild- type SEX1 cDNA using the cauliflower mosaic virus 35S promoter. The results demonstrate a genetic link between the SEX1 locus and plant freezing tolerance, and show that starch degradation is important for enhanced freezing tolerance during an early phase of cold acclimation. However, induction of starch degradation was not accompanied by significant changes in a- glucan/ water dikinase activity in leaf extracts and preceded cold- induced augmentation of SEX1 transcripts. Therefore, we conclude that augmentation of SEX1 transcripts might be a homeostatic response to low temperature, and that starch degradation during an early phase of cold acclimation could be regulated by a component( s) of a starch degradation pathway( s) downstream of SEX1.