A mitotic GlcNAcylation/phosphorylation signaling complex alters the posttranslational state of the cytoskeletal protein vimentin.
O-linked beta-N-acetylglucosamine (O-GlcNAc) is a highly dynamic intracellular protein modification responsive to stress, hormones, nutrients, and cell cycle stage. Alterations in O-GlcNAc addition or removal (cycling) impair cell cycle progression and cytokinesis, but the mechanisms are not well understood. Here, we demonstrate that the enzymes responsible for O-GlcNAc cycling, O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA) are in a transient complex at M phase with the mitotic kinase Aurora B and protein phosphatase 1. OGT colocalized to the midbody during telophase with Aurora B. Furthermore, these proteins coprecipitated with each other in a late mitotic extract. The complex was stable under Aurora inhibition; however, the total cellular levels of O-GlcNAc were increased and the localization of OGT was decreased at the midbody after Aurora inhibition. Vimentin, an intermediate filament protein, is an M phase substrate for both Aurora B and OGT. Overexpression of OGT or OGA led to defects in mitotic phosphorylation on multiple sites, whereas OGT overexpression increased mitotic GlcNAcylation of vimentin. OGA inhibition caused a decrease in vimentin late mitotic phosphorylation but increased GlcNAcylation. Together, these data demonstrate that the O-GlcNAc cycling enzymes associate with kinases and phosphatases at M phase to regulate the posttranslational status of vimentin.
Slawson
Chad
C
Department of Biological Chemistry, The Johns Hopkins School of Medicine, Baltimore, MD 21205, USA.
Lakshmanan
T
T
Knapp
Spencer
S
Hart
Gerald W
GW
D000117:Acetylglucosamine / Q000378:metabolism*; D064107:Aurora Kinase B; D064127:Aurora Kinases; D002455:Cell Division; D048749:Cytokinesis; D006367:HeLa Cells; D006801:Humans; D018613:Microscopy, Confocal; D015877:Miosis; D008938:Mitosis; D008954:Models, Biological; D017351:N-Acetylglucosaminyltransferases / Q000378:metabolism*; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D015398:Signal Transduction; D014746:Vimentin / Q000378:metabolism*
D014746:Vimentin; D017351:N-Acetylglucosaminyltransferases; C449880:O-GlcNAc transferase; C576921:AURKB protein, human; D064107:Aurora Kinase B; D064127:Aurora Kinases; D017346:Protein Serine-Threonine Kinases; D000117:Acetylglucosamine