O-GlcNAc signaling and OGT biochemistry
Although the monosaccharide addition of GlcNAc to serine and threonine residues of nuclear and cytosolic proteins was discovered in the early 80’s, we know little about its function.
O-GlcNAc background
O-GlcNAc is a single N-acetylglucosamine coupled to serine and threonine residues of nuclear and cytoplasmic proteins. Analogous to phosphorylation, O-GlcNAc signaling is dynamic, rapidly added and removed from proteins in a site-specific manner in response to cellular perturbations and extracellular cues. Because both modifications occur on the same residues it is hypothesized that there is a functional crosstalk between OGlcNAc and phosphorylation, where one may affect deposition or removal the other. Unlike phosphorylation, however, which is catalyzed by over 500 kinases and roughly 300 phosphatases, the mammalian genome only encodes a single O-GlcNAc transferase (OGT) and a single hydrolase (OGA). While many kinases recognize specific amino acid sequences in their substrates, the determinants guiding OGT are unclear and likely manifold. This intracellular glycosylation is implicated in nearly every cellular process from gene expression and signal transduction to cell division and differentiation. Despite the ubiquitous nature of this post-translational modification in health and disease, the specific functions of OGT and the basic principles of O-GlcNAc signaling remain largely elusive.
O-GlcNAc signaling
Unambiguously identifying O-GlcNAcylated proteins, especially the exact residue being modified, has been a notorious challenge. This is largely due to A) O-GlcNAc being chemically similar to other GlcNAc-containing glycans, making specific enrichment reagents challenging, and B) the labile nature of the glycosidic bond hampering confident modified site identification. We develop and apply novel enrichment and mass spectrometric approaches to confidently identify and quantify O-GlcNAc-modified peptides from cells undergoing cell state transitions, such as immune cell activation or stem cell differentiation.
OGT biochemistry
The sole O-GlcNAc transferase (OGT) is essential for cell survival, and is the least mutated glycosyltransferase in the human genome. Moreover, OGT interacts with nearly every cellular process. These features make studying OGT via genetic mechanisms extremely challenging. Thus, we develop proteomic technologies to better understand how OGT can be involved in so many cellular processes, and what its function is in each one.
