Spotlights on the post-translational regulation of enzymes involved in glucitol metabolism. Phosphorylation of aldose-6-phosphate reductase

IGLESIAS, ALBERTO A.; FIGUEROA, CARLOS M.; HARTMAN, MATÍAS D.

Honolulu

Congreso; Plant Biology 2017; 2017

American Society of Plant Biologists

In addition to sucrose and starch, sugar alcohols(like glucitol) are primary photosynthates in a significant number of plantspecies. Glucitol, also known as sorbitol, is produced in mature leaves fromglucose-6-phosphate by the combined action of aldose-6-phosphate reductase(Ald6PRase) and a specific phosphatase. Thereafter, glucitol is translocated tosink tissues where it is oxidized to fructose by glucitol dehydrogenase.Research performed during the last two decades established that glucitolmetabolism is regulated at the post-transcriptional level. More recently, wedemonstrated that redox regulation of peach Ald6PRase and glucitoldehydrogenase orchestrates glucitol metabolism in both source and sink tissues,respectively. In this work, we show that Ald6PRase from peach leaves wasrecovered using Fe3+-IMAC, suggesting that the enzyme could be phosphorylatedin vivo. Recombinant peach Ald6PRase was phosphorylated in vitroby a partially purified protein kinase from peach leaves. Based on immunodetectionand phosphorylation of synthetic peptides we hypothesize this protein kinasebelongs to the SnRK1 family. The partially purified protein kinase wasinhibited by fructose-1,6-bisphosphate, glucose-6-phosphate,glucitol-6-phosphate, pyrophosphate and phosphoenolpyruvate. Treatmentof protein extracts from peach leaves with alkaline phosphatase increased theactivity of Al6PRase and prevented its binding to the IDA-Fe3+resin, indicating that phosphorylation would inhibit enzyme activity. Databasesearch allowed us to identify a homolog of peach Ald6PRase in Arabidopsis,phosphorylated at a highly conserved Thr residue in many plant aldo-ketoreductases. Interestingly, the recombinant peach Ald6PRase T257D phosphomimeticmutant had negligible enzymatic activity. Overall, our results suggest thatphosphorylation of Ald6PRase would be a mechanism regulating glucitol synthesisin leaves.