SummaryA paradigm-changing discovery in biology came about when it was found that nuclear and cytosolic proteins could be dynamically glycosylated with a single O-linked b-N-acetylglucosamine (O-GlcNAc) moiety. O-GlcNAcylation is akin to phosphorylation: it occurs on serine and/or threonine side chains of proteins, and cycles rapidly upon cellular activation. O-GlcNAc and phosphate show a complex interplay: they can either competitively occupy a single site or proximal sites, or noncompetitively occupy different sites on a substrate. Phosphorylation regulates O-GlcNAc-cycling enzymes and, conversely, O-GlcNAcylation controls phosphate-cycling enzymes. Such crosstalk is evident in all compartments of the cell, a finding that is congruent with the fundamental role of O-GlcNAc in regulating nutrient-and stress-induced signal transduction. O-GlcNAc transferase is recruited to the plasma membrane in response to insulin and is targeted to substrates by forming transient holoenzyme complexes that have different specificities. Cytosolic O-GlcNAcylation is important for the proper transduction of signaling cascades such as the NFkB pathway, whereas nuclear O-GlcNAc is crucial for regulating the activity of numerous transcription factors. This Commentary focuses on recent findings supporting an emerging concept that continuous crosstalk between phosphorylation and O-GlcNAcylation is essential for the control of vital cellular processes and for understanding the mechanisms that underlie certain neuropathologies.

At least 20 core ribosome proteins are modified by O-GlcNAc. O-GlcNAcase is localized to the nucleolus and O-GlcNAc transferase is excluded from the nucleolus. Both enzymes associate with active polysomes. Overexpression of OGT disrupts ribosomal subunit homeostasis. Data suggest that O-GlcNAc regulates translation and ribosome biogenesis.

Contractile dysfunction and increased deposition of O-linked β-N-acetyl-d-glucosamine (O-GlcNAc) in cardiac proteins are a hallmark of the diabetic heart. However, whether and how this posttranslational alteration contributes to lower cardiac function remains unclear. Using a refined β-elimination/Michael addition with tandem mass tags (TMT)–labeling proteomic technique, we show that CpOGA, a bacterial analog of O-GlcNAcase (OGA) that cleaves O-GlcNAc in vivo, removes site-specific O-GlcNAcylation from myofilaments, restoring Ca2+ sensitivity in streptozotocin (STZ) diabetic cardiac muscles. We report that in control rat hearts, O-GlcNAc and O-GlcNAc transferase (OGT) are mainly localized at the Z-line, whereas OGA is at the A-band. Conversely, in diabetic hearts O-GlcNAc levels are increased and OGT and OGA delocalized. Consistent changes were found in human diabetic hearts. STZ diabetic hearts display increased physical interactions of OGA with α-actin, tropomyosin, and myosin light chain 1, along with reduced OGT and increased OGA activities. Our study is the first to reveal that specific removal of O-GlcNAcylation restores myofilament response to Ca2+ in diabetic hearts and that altered O-GlcNAcylation is due to the subcellular redistribution of OGT and OGA rather than to changes in their overall activities. Thus, preventing sarcomeric OGT and OGA displacement represents a new possible strategy for treating diabetic cardiomyopathy.