Targeting and Regulation of O-GlcNAc Transferase at M Phase

O-GlcNAc是由糖基乙酰氨基葡萄糖在丝氨酸/苏氨酸氨基酸上对蛋白质进行共价修饰。O-GlcNAc修饰不能延伸到更复杂的寡糖结构，只存在于核蛋白和细胞质蛋白上。这种修饰是高度动态的，对细胞外环境如激素、压力和营养物质的信号作出反应。之前，我证明了0-GlcNAc对真核细胞中细胞周期的正常进展至关重要；函数的增益

O-GlcNAc is the covalent modification of proteins at serine/threonine amino acids by the sugar moiety Nacetylglucosamine. The O-GlcNAc modification is not elongated to more complex oligosaccharide structures and is found only on nuclear and cytoplasmic proteins. The modification is highly dynamic and responds to signals of the extracellular environment such hormones, stress, and nutrients. Previously, I demonstrated that 0-GlcNAc is critical for the proper progression of the cell cycle in eukaryotic cells; gain of function of either 0-GlcNAc transferase (OGT), the enzyme which adds the modification, or 0-GlcNAcase (OGA), the enzyme which removes the modification, causes mitotic exit delays. Furthermore, OGT and OGA can be found in a signaling complex with mitotic kinases such as Aurora Kinase B. The goal of this proposal is to investigate the interactions of OGT with mitotic structures such as the spindle and with mitotic proteins. We hypothesize that OGT forms complexes with various proteins during M phase progression that then targets OGT the spindle and midbody. I plan to use multiple techniques such as using FRAP (Fluorescent Recovery after Photo-bleaching) on GFP-OGT to measure the kinetics of OGT at spindle/midbody, quantitative proteomics to identify OGT targeting proteins, and finally identifying the function of Aurora Kinase B in targeting OGT to mitotic substrates and structures. Our expected contribution is significant because we expect to find insight into the regulation of OGT by mitotic targeting proteins and how these interactions are uncoupled in diseases such as cancer.