Proteomics

Dataset Information

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Spatiotemporal proximity labeling tools to track GlcNAc-modified functional protein hubs during cellular signaling


ABSTRACT: A fundamental mechanism that all eukaryotic cells use to adapt to their environment is dynamic protein modification with monosaccharide sugars. In humans, O-linked N-acetylglucosamine (O-GlcNAc) is rapidly added and removed on diverse protein sites as a response to fluctuating nutrient levels, stressors, and signaling cues. Two aspects elude methods to track O-GlcNAc response functions with chemical strategies: spatial control over subcellular locations and time control during labeling reactions. The objective of this study was to create intracellular proximity labeling tools to identify functional changes of O-GlcNAc patterns with spatiotemporal control. We developed a labeling strategy based on TurboID system for rapid protein biotin conjugation that was directed to O-GlcNAc protein modifications inside cells, a set of tools we call “GlycoID.” Localized variants to the nucleus and cytosol, nuc-GlycoID and cyt-GlycoID, label O-GlcNAc proteins and their interactomes in subcellular space. Labeling during insulin as well as serum stimulation revealed functional changes in O-GlcNAc proteins in as soon as 30 minutes of signaling. We demonstrate that the GlycoID strategy can capture O-GlcNAcylated “activity hubs” consisting of O-GlcNAc proteins and their associated protein-protein interactions. The ability to follow changes in O-GlcNAc hubs during physiological events like insulin stimulation poises these tools to be used for determining mechanisms of glycobiological cell regulation. Our datasets in HeLa cells will be a useful functional resource for O-GlcNAc-associated physiology.

INSTRUMENT(S): Orbitrap Fusion

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Permanent Cell Line Cell, Cell Culture

DISEASE(S): Mixed Disorder As Reaction To Stress

SUBMITTER: Charlie Fehl  

LAB HEAD: Charlie Fehl

PROVIDER: PXD033063 | Pride | 2022-10-15

REPOSITORIES: Pride

Dataset's files

Source:
Action DRS
allSpectra.CID.ITMS.iso_0.apl Other
allSpectra.CID.ITMS.iso_1.apl Other
allSpectra.CID.ITMS.iso_10.apl Other
allSpectra.CID.ITMS.iso_11.apl Other
allSpectra.CID.ITMS.iso_12.apl Other
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Publications

Spatiotemporal Proximity Labeling Tools to Track GlcNAc Sugar-Modified Functional Protein Hubs during Cellular Signaling.

Liu Yimin Y   Nelson Zachary M ZM   Reda Ali A   Fehl Charlie C  

ACS chemical biology 20220712 8


A fundamental mechanism that all eukaryotic cells use to adapt to their environment is dynamic protein modification with monosaccharide sugars. In humans, O-linked <i>N</i>-acetylglucosamine (O-GlcNAc) is rapidly added to and removed from diverse protein sites as a response to fluctuating nutrient levels, stressors, and signaling cues. Two aspects remain challenging for tracking functional O-GlcNAc events with chemical strategies: spatial control over subcellular locations and time control durin  ...[more]

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