Generation of an Unbiased Interactome for the Tetratricopeptide Repeat Domain of O-GlcNAc Transferase Indicates a Role for the Enzyme in Intellectual Disability

Abstract

The O-GIcNAc transferase (OGT) is localized to the nucleus and cytoplasm where it regulates nucleocytoplasmic proteins by modifying serine and threonine residues with a non-extended monosaccharide, {beta}-N-Acetyl-Glucosamine (O-GlcNAc). With thousands of known O-GlcNAc modified proteins but only one OGT encoded in the mammalian genome, a prevailing question is how OGT selects its substrates. Prior work has indicated that the N-terminal tetratricopeptide repeat (TPR) domain of OGT, rather than its C-terminal catalytic domain, is responsible for subcellular targeting and substrate selection. An additional impetus for exploring the OGT TPR domain interactome is the fact that missense mutations in OGT associated with X-linked intellectual disability (XLID) are primarily localized to the TPR domain without substantial impact on activity or stability of the enzyme. Therefore, we adapted the BioID labeling method to identify interactors of a TPR-BirA* fusion protein in HeLa cells. We identified 115 high confidence interactors representing both known and novel O-GlcNAc modified proteins and OGT interactors. The TPR interactors are highly enriched in processes in which OGT has a known role (e.g. chromatin remodeling, cellular survival of heat stress, circadian rhythm), as well as processes i n which OGT has yet to be implicated (e.g. pre-mRNA processing). Importantly, the identified TPR interactors are involved in several disease states but most notably are highly enriched in pathologies featuring intellectual disability. These proteins represent candidate interactors that may underlie the mechanism by which mutations in OGT lead to XLID. Furthermore, the identified interactors provide additional evidence of the importance of the TPR domain for OGT targeting and/or substrate selection. Thus, this defined interactome for the TPR domain of OGT serves as a jumping off point for future research exploring the role of OGT, the TPR domain, and its protein interactors in multiple cellular processes and disease mechanisms, including intellectual disability.