Metabolism of Triose-phosphate in Wheat. Phosphorylation of cytosolic non-phosphorylating glyceraldehyde-3-phosphate dehydrogenase

A.A. IGLESIAS; C.V. PIATTONI; S.A. GUERRERO

Minneapolis

Congreso; Annual Meeting of the American Society of Plant Biologists of Plant Physiologists; 2011

Non phosphorylating glyceraldehyde-3-phosphate dehydrogenase (np-Ga3PDHase, EC1.2.1.9) is a glycolytic enzyme typical of the cytosol of plant cells, mainly involved inderiving triose-P to produce NADPH. The enzyme was found to be modified byphosphorylation in wheat heterotrophic tissues. Phosphorylated np-Ga3PDHase interactswith 14-3-3 regulatory proteins; afterward it turns less active and more sensitive toregulation by adenylates and pyrophosphate. Herein we identify that the Ser404 residue ofwheat np-Ga3PDHase is specifically phosphorylated by the SnRK (SNF1 related) proteinkinase family. Interestingly, only the kinase present in wheat heterotrophic tissues(endosperm), but not in leaves, was found active. The specific SnRK was partially purifiedand characterized as a ~200 kDa protein. The kinase also phosphorylated SAMS, AMARAand SP46 peptides and it was recognized by antibodies rose against a peptide conserved inSnRK1 from sorghum and maize developing seeds. The kinase required Mg2+ or Mn2+(but not Ca2+) for activity and it was allosterically inhibited by physiological concentrations of ribose-5P, and to a lesser extent by fructose-1,6-bisP and 3P-glycerate. Glucose-6P (a main effector of spinach leaf SnRK1) produced little effect. MALDI-TOF/TOF analysis evidenced that sucrose synthase copurified with SnRK1, also being a target of phosphorylation. Results support np-Ga3PDHase as a new target of SnRK1 action,identifying distinctive allosteric regulation of SnRK1s present in photosynthetic or nongreentissues. Phosphorylation of np-Ga3PDHase would be part of a mechanism regulatingcarbon partitioning, determining triose-P usage to produce NADPH or ATP in the cytosol ofheterotrophic plant cells.