Cross-talk between Two Essential Nutrient-sensitive Enzymes O-GlcNAc TRANSFERASE ( OGT) AND AMP-ACTIVATED PROTEIN KINASE ( AMPK)

Background: OGT and AMPK collectively target hundreds of intracellular signaling processes, but no study has addressed whether they regulate each other. Results: AMPK activity mediates the substrate selectivity of OGT, and O-GlcNAcylation modulates the activity of AMPK. Conclusion: There is significant cross-talk between the O-GlcNAc and AMPK systems. Significance: OGT and AMPK may synergistically regulate numerous nutrient-sensitive processes essential for life. Nutrient-sensitive pathways regulate both O-GlcNAc transferase (OGT) and AMP-activated protein kinase (AMPK), cooperatively connecting metabolic homeostasis to regulation of numerous intracellular processes essential for life. Similar to phosphorylation, catalyzed by kinases such as AMPK, O-GlcNAcylation is a highly dynamic Ser/Thr-specific post-translational modification of nuclear, cytoplasmic, and mitochondrial proteins catalyzed exclusively by OGT. OGT and AMPK target a multitude of intracellular proteins, with the net effect to protect cells from the damaging effects of metabolic stress. Despite hundreds of studies demonstrating significant overlap in upstream and downstream signaling processes, no study has investigated if OGT and AMPK can directly regulate each other. We show acute activation of AMPK alters the substrate selectivity of OGT in several cell lines and nuclear localization of OGT in C2C12 skeletal muscle myotubes. Nuclear localization of OGT affects O-GlcNAcylation of numerous nuclear proteins and acetylation of Lys-9 on histone 3 in myotubes. AMPK phosphorylates Thr-444 on OGT in vitro; phosphorylation of Thr-444 is tightly associated with AMPK activity and nuclear localization of OGT in myotubes, and phospho-mimetic T444E-OGT exhibits altered substrate selectivity. Conversely, the - and -subunits of AMPK are O-GlcNAcylated, O-GlcNAcylation of the 1-subunit increases with AMPK activity, and acute inhibition of O-GlcNAc cycling disrupts activation of AMPK. We have demonstrated significant cross-talk between the O-GlcNAc and AMPK systems, suggesting OGT and AMPK may cooperatively regulate nutrient-sensitive intracellular processes that mediate cellular metabolism, growth, proliferation, and/or tissue function.