Identification of the nuclear localisation signal of O-GlcNAc transferase and its nuclear import regulation

Abstract

Nucleocytoplasmic O-GlcNAc transferase (OGT) attaches a single GlcNAc to hydroxyl groups of serine and threonine residues. Although the cellular localisation of OGT is important to regulate a variety of cellular processes, the molecular mechanisms regulating the nuclear localisation of OGT is unclear. Here, we characterised three amino acids (DFP; residues 451–453) as the nuclear localisation signal of OGT and demonstrated that this motif mediated the nuclear import of non-diffusible β-galactosidase. OGT bound the importin α5 protein, and this association was abolished when the DFP motif of OGT was mutated or deleted. We also revealed that O-GlcNAcylation of Ser389, which resides in the tetratricopeptide repeats, plays an important role in the nuclear localisation of OGT. Our findings may explain how OGT, which possesses a NLS, exists in the nucleus and cytosol simultaneously.

Crosstalk between phosphorylation and O‐GlcNAcylation: friend or foe

“…OGT has three isoforms that differ in the number of tetratricopeptide repeats (TPRs) they harbor at their N terminus, a well-described protein-protein interaction-mediating domain [49]. mOGT has an Nterminal mitochondrial localization sequence, whereas ncOGT contains a nuclear localization sequence between its TPRs and the catalytic domain located at the C terminus [51,52]. The OGT isoforms localize to different compartments of the cell.…”

“…Regulation may in part, therefore, be governed by (the number of) these TPR repeats. Deletion of this sequence prevents transport of ncOGT into the nucleus, resulting in an increase of cytosolic O-GlcNAcylation [51]. Short OGT (sOGT) and nucleocytoplasmic OGT (ncOGT) are found in the nucleus and cytoplasm, whereas mitochondrial OGT (mOGT) is found exclusively in mitochondria [50].…”

“…Localization of mOGT and ncOGT is directed by localization sequences. mOGT has an Nterminal mitochondrial localization sequence, whereas ncOGT contains a nuclear localization sequence between its TPRs and the catalytic domain located at the C terminus [51,52]. Deletion of this sequence prevents transport of ncOGT into the nucleus, resulting in an increase of cytosolic O-GlcNAcylation [51].…”

“…mOGT has an Nterminal mitochondrial localization sequence, whereas ncOGT contains a nuclear localization sequence between its TPRs and the catalytic domain located at the C terminus [51,52]. Deletion of this sequence prevents transport of ncOGT into the nucleus, resulting in an increase of cytosolic O-GlcNAcylation [51]. The localization sequence in ncOGT is also present in the other isoforms, thus sOGT likely also has a localization sequence whose relevance is yet to be reported.…”

Critical observations that shaped our understanding of the function(s) of intracellular glycosylation (O‐GlcNAc)

“…The first study demonstrated that Ser20 phosphorylation by checkpoint kinase 1 (Chk1) is essential for localization of OGT to midbodies during cytokinesis in HeLa cells , and the second study determined that phosphorylation by calcium/calmodulin‐dependent kinase (CaMK) II altered the pattern of O‐GlcNAcylation in hepatocytes . Three recent studies have identified several sites of O‐GlcNAcylation on OGT isoform 4 , two sites at the N terminus (Ser03, Ser04) of OGT (Isoform 1 and 3) , and one site at Ser389 on OGT (Isoform 1) . The latter site has been implicated in promoting nuclear import of OGT in HeLa cells.…”