Writing and Erasing O-GlcNAc on Casein Kinase 2 Alpha Alters thePhosphoproteome

O-GlcNAc is an essential carbohydrate modificationthat intersects with phosphorylation signaling pathways via crosstalkon protein substrates or by direct modification of the kinases thatwrite the phosphate modification. Casein kinase 2 alpha (CK2 alpha), thecatalytic subunit of the ubiquitously expressed and constitutivelyactive kinase CK2, is modified by O-GlcNAc, but the effect of thismodification on the phosphoproteome in cells is unknown. Here, weapply complementary targeted O-GlcNAc editors, nanobody-OGTand -splitOGA, to selectively write and erase O-GlcNAc from atagged CK2 alpha to measure the effects on the phosphoproteome in cells. These tools effectively and selectively edit the Ser347glycosite on CK2 alpha. Using quantitative phosphoproteomics, we report 51 phosphoproteins whose enrichment changes as a functionof editing O-GlcNAc on CK2 alpha, including HDAC1, HDAC2, ENSA, SMARCAD1, and PABPN1. Specific phosphosites on HDAC1Ser393 and HDAC2 Ser394, both reported CK2 substrates, are significantly enhanced by O-GlcNAcylation of CK2 alpha. These data willpropel future studies on the crosstalk between O-GlcNAc and phosphorylation

Automated summary

This study utilized novel targeted O-GlcNAc editing tools to investigate the effect of O-GlcNAc modification on the CK2α phosphoproteome. The results showed that O-GlcNAc modification significantly enhanced specific phosphosites on HDAC1 and HDAC2.