A novel glycoproteomics workflow reveals dynamic O-GlcNAcylation of COPγ1 as a candidate regulator of protein trafficking

Published

Journal Article

Copyright © 2018 Cox, Luo, Smith, Bisnett, Soderblom and Boyce. O-linked ß-N-acetylglucosamine (O-GlcNAc) is an abundant and essential intracellular form of protein glycosylation in animals and plants. In humans, dysregulation of O-GlcNAcylation occurs in a wide range of diseases, including cancer, diabetes, and neurodegeneration. Since its discovery more than 30 years ago, great strides have been made in understanding central aspects of O-GlcNAc signaling, including identifying thousands of its substrates and characterizing the enzymes that govern it. However, while many O-GlcNAcylated proteins have been reported, only a small subset of these change their glycosylation status in response to a typical stimulus or stress. Identifying the functionally important O-GlcNAcylation changes in any given signaling context remains a significant challenge in the field. To address this need, we leveraged chemical biology and quantitative mass spectrometry methods to create a new glycoproteomics workflow for profiling stimulus-dependent changes in O-GlcNAcylated proteins. In proof-of-principle experiments, we used this new workflow to interrogate changes in O-GlcNAc substrates in mammalian protein trafficking pathways. Interestingly, our results revealed dynamic O-GlcNAcylation of COPγ1, an essential component of the coat protein I (COPI) complex that mediates Golgi protein trafficking. Moreover, we detected 11 O-GlcNAc moieties on COPγ1 and found that this modification is reduced by a model secretory stress that halts COPI trafficking. Our results suggest that O-GlcNAcylation may regulate the mammalian COPI system, analogous to its previously reported roles in other protein trafficking pathways. More broadly, our glycoproteomics workflow is applicable to myriad systems and stimuli, empowering future studies of O-GlcNAc in a host of biological contexts.

Full Text

Duke Authors

Cited Authors

  • Cox, NJ; Luo, PM; Smith, TJ; Bisnett, BJ; Soderblom, EJ; Boyce, M

Published Date

  • October 15, 2018

Published In

Volume / Issue

  • 9 / OCT

Electronic International Standard Serial Number (EISSN)

  • 1664-2392

Digital Object Identifier (DOI)

  • 10.3389/fendo.2018.00606

Citation Source

  • Scopus