The Role of SNF1-Related Protein Kinase 1 (SnRK1) in Regulating Intermediary Metabolism in Arabidopsis thaliana
The Sucrose Non-fermenting-1 (SNF1)-Related Protein Kinase 1 (SnRK1) is a highly conserved heterotrimeric protein kinase which plays an important role in the control of energy balance in plants. It possesses a catalytic subunit (α) and two regulatory subunits (β and γ). This research project focuses on understanding the role of one of its regulatory subunits, AKINβ1, on metabolism in Arabidopsis. The effects of altered expression of AKINβ1 on carbohydrate metabolism in plants was investigated in an Arabidopsis T-DNA insertion mutant. The concentrations of key intermediates in the tricarboxylic acid cycle of the mutant leaves were markedly reduced throughout the diurnal cycle, and this was coupled with a decrease in measurable respiration rate. The subcellular localization of AKINα1 (AKIN11), AKINβ1, and AKINγ1 subunits of SnRK1 was investigated in tobacco and Arabidopsis leaves, using fluorescence-tagged proteins. The regulatory function of N-myristoylation on the subcellular localization of AKINβ1 was also investigated, indicating that AKINβ1 binds to the Golgi stack, and that the N-terminal 74-amino acids of AKINβ1 possesses a putative nuclear localization signal. Recombinant fusion proteins (AKINα1, AKINβ1 and AKINγ1) of SnRK1 were expressed in E. coli and used as bait to study their interaction with other proteins in plant leaf cells. LHCB1.1/LHCB1.2 or/and LHCB1.3 were identified as putative interacting protein(s) of AKINα1. Finally, the effect of altered expression of AKINβ1 on transcriptional regulation was studied. Compared to WT, 2485 genes and 188 genes were expressed differentially in the akinβ1 mutant leaves in response to light and darkness respectively. Very large changes in expression were observed in several genes including glycerol-3-phosphate acyltransferase 5 (-243 fold), NADP-dependent malic enzyme 1 (-70 fold), and nitrate transporter 1.8 (-52 fold), indicating that the AKINβ subunit plays a significant role in modulating carbohydrate, lipid, and nitrogen metabolism. A model is hypothesized to explain the effects of AKINβ1 on metabolism in Arabidopsis. The results in this study provide new insight into the role of SnRK1, especially AKINβ1, in regulating plant metabolism.