O‐GlcNAcase is Cleaved Between Its Glycosidase and Histone Acetyltransferase Domains by Caspase‐3 During Apoptosis

Abstract

O-GlcNAc is a dynamic post-translational modification on myriad nucleocytoplasmic proteins, ranging from transcription factors, signaling proteins, to cytoskeletal proteins. O-GlcNAc is a nutrient sensor involved in the regulation of cellular activity, including transcription, response to stress, and protein-protein interactions. O-GlcNAcase, the O-GlcNAc removal enzyme, has been shown to be a substrate of caspase-3 in vitro. Here we identify the cleavage site of O-GlcNAcase by caspase-3. Using recombinant O-GlcNAcase and recombinant caspase-3, we have mapped the cleavage site by Edman sequencing. We find that the cleavage site is a non-canonical recognition site that occurs after Asp-413 of the tetrapeptide sequence SVVD in the human O-GlcNAcase. A point mutation, D413A, abrogates cleavage by caspase-3 in vitro. We also show that O-GlcNAcase is a substrate of caspase-3 during Fas-mediated apoptosis in vivo separating the two functional domains of this bi-functional enzyme These data suggest that O-GlcNAc cycling is affected by apoptosis induction and that O-GlcNAc and O-GlcNAcase itself are involved in the regulation of apoptosis. Supported by NIH grants CA42486 and HD13563 and G.W. H. receives a share of royalty received by the university on sales of the CTD 110.6 antibody. The terms of this arrangement are being managed by The Johns Hopkins University in accordance with its conflict of interest policies.