Distinct OGT-Binding Sites Promote HCF-1 Cleavage

• DOI: 10.1371/journal.pone.0136636
• Publication Date: 2015-08-25T18:19:23Z
• Authors: Tanja Bhuiyan; Patrice Waridel; Vaibhav Kapuria; Vincent Zoete; Winship Herr
• Keywords: Repetitive Sequences, Amino Acid; 0303 health sciences; Binding Sites; Transcription, Genetic; Science; Q; R; Glutamic Acid; N-Acetylglucosaminyltransferases; Protein Subunits; 03 medical and health sciences; Proteolysis; Medicine; Humans; Binding Sites; Cytokinesis/genetics; Glutamic Acid/metabolism; HeLa Cells; Host Cell Factor C1/genetics; Host Cell Factor C1/metabolism; Humans; N-Acetylglucosaminyltransferases/genetics; N-Acetylglucosaminyltransferases/metabolism; Protein Subunits/genetics; Protein Subunits/metabolism; Proteolysis; Repetitive Sequences, Amino Acid/genetics; Transcription, Genetic; Host Cell Factor C1; Research Article; Cytokinesis; HeLa Cells

Human HCF-1 (also referred to as HCFC-1) is a transcriptional co-regulator that undergoes a complex maturation process involving extensive O-GlcNAcylation and site-specific proteolysis. HCF-1 proteolysis results in two active, noncovalently associated HCF-1N and HCF-1C subunits that regulate distinct phases of the cell-division cycle. HCF-1 O-GlcNAcylation and site-specific proteolysis are both catalyzed by O-GlcNAc transferase (OGT), which thus displays an unusual dual enzymatic activity. OGT cleaves HCF-1 at six highly conserved 26 amino acid repeat sequences called HCF-1PRO repeats. Here we characterize the substrate requirements for OGT cleavage of HCF-1. We show that the HCF-1PRO-repeat cleavage signal possesses particular OGT-binding properties. The glutamate residue at the cleavage site that is intimately involved in the cleavage reaction specifically inhibits association with OGT and its bound cofactor UDP-GlcNAc. Further, we identify a novel OGT-binding sequence nearby the first HCF-1PRO-repeat cleavage signal that enhances cleavage. These results demonstrate that distinct OGT-binding sites in HCF-1 promote proteolysis, and provide novel insights into the mechanism of this unusual protease activity.