Chemoproteomic and Transcriptomic Analysis Reveals that O-GlcNAc Regulates Mouse Embryonic Stem Cell Fate through the Pluripotency Network

Self-renewal and differentiation of embryonic stem cells (ESCs) are influenced by protein O-linked beta-N-acetylglucosamine (O-GlcNAc) modification, but the underlying mechanism remains incompletely understood. Herein, we report the identification of 979 O-GlcNAcylated proteins and 1340 modification sites in mouse ESCs (mESCs) by using a chemoproteomics method. In addition to OCT4 and SOX2, the third core pluripotency transcription factor (PTF) NANOG was found to be modified and functionally regulated by O-GlcNAc. Upon differentiation along the neuronal lineage, the O-GlcNAc stoichiometry at 123 sites of 83 proteins-several of which were PTFs-was found to decline. Transcriptomic profiling reveals 2456 differentially expressed genes responsive to OGT inhibition during differentiation, of which 901 are target genes of core PTFs. By acting on the core PTF network, suppression of O-GlcNAcylation upregulates neuron-related genes, thus contributing to mESC fate determination.

Automated summary

The O-GlcNAc modification is found to affect the self-renewal and differentiation of embryonic stem cells (ESCs), and the core pluripotency transcription factor NANOG is regulated by this modification. During neuronal lineage differentiation, the O-GlcNAc modification decreases, and suppressing O-GlcNAcylation can upregulate neuron-related genes, thus influencing the fate determination of ESCs.