Effect of catalytic subunit phosphorylation on the properties of SnRK1 from Phaseolus vulgaris embryos.

Abstract

Legume seed development represents a high demand for energy and metabolic resources to support the massive synthesis of starch and proteins. However, embryo growth occurs in an environment with reduced O2 that forces the plant to adapt its metabolic activities to maximize efficient energy use. SNF1-related protein kinase1 (SnRK1) is a master metabolic regulator needed for cells adaptation to conditions that reduce energy availability, and its activity is needed for the successful development of seeds. In bean embryo extracts, SnRK1 can be separated by anion exchange chromatography into two pools: one where the catalytic subunit is phosphorylated (SnRK1-p) and another with reduced phosphorylation (SnRK1-np). The phosphorylation of the catalytic subunit produces a large increase in SnRK1 activity but has a minor effect in determining its sensitivity to metabolic inhibitors such as trehalose 6-P (T6P), ADP-glucose (ADPG), glucose 1-P (G1P) and glucose 6-P (G6P). In Arabidopsis thaliana, upstream activating kinases (SnAK) phosphorylate the SnRK1 catalytic subunit at T175/176, promoting and enhancing its activity. Recombinant Phaseolus vulgaris homologous to SnAK proteins (PvSnAK), can phosphorylate and activate the catalytic domains of the α-subunits of Arabidopsis, as well as the SnRK1-np pool purified from bean embryos. While the phosphorylation process is extremely efficient for catalytic domains, the phosphorylation of the SnRK1-np complex was less effective but produced a significant increase in activity. The presence of SnRK1-np could contribute to a quick response to unexpected adverse conditions. However, in addition to PvSnAK kinases, other factors might contribute to regulating the activation of SnRK1.