Protein O-GlcNAcylation is the only known type of glycosylation that primarily happens in the nucleus and the cytoplasm. O-GlcNAcylation plays extremely important roles in many cellular events, and its dysregulation is related to multiple diseases. Integrating bioorthogonal chemistry and multiplexed proteomics, we systematically and site-specifically studied the distribution and dynamics of protein O-GlcNAcylation in the nucleus and the cytoplasm in human cells. The results demonstrated that O-GlcNAcylated proteins with different functions had distinct patterns of the distributions in these two cellular compartments, and the distributions of glycoproteins varied site-specifically. For example, for transcriptional repressor p66-alpha (GATAD2A), its glycoform with the site of T329 is much highly enriched in the nucleus while the glycoform with the site of S625 or S629 is equally distributed in both compartments. Moreover, the degradation of O-GlcNAcylated proteins in the nucleus and the cytoplasm are markedly different with the median half-life of O-GlcNAcylated proteins in the cytoplasm being about two times longer than that in the nucleus, and the same protein in different locations may have different dynamics. For instance, transcription factors in the nucleus have significantly shorter half-lives than the same glycoproteins in the cytoplasm. Kinases in the two compartments are more dynamic than other groups of proteins. The spatial and temporal analysis of protein O-GlcNAcylation aids in a better understanding of the properties and functions of this important modification and glycoproteins.

Spatial and temporal proteomics reveals distinct distribution and degradation of O-GlcNAcylated proteins

ACS Spring Meeting 2021

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