title: S phase-specific proteolytic cleavage is required to activate stable DNA binding by the CDP/Cut homeodomain protein.
abstract: The CCAAT displacement protein (CDP), the homologue of the Drosophila melanogaster Cut protein, contains four DNA binding domains that function in pairs. Cooperation between Cut repeat 3 and the Cut homeodomain allows stable DNA binding to the ATCGAT motif, an activity previously shown to be upregulated in S phase. Here we showed that the full-length CDP/Cut protein is incapable of stable DNA binding and that the ATCGAT binding activity present in cells involves a 110-kDa carboxy-terminal peptide of CDP/Cut. A vector expressing CDP/Cut with Myc and hemagglutinin epitope tags at either end generated N- and C-terminal products of 90 and 110 kDa, suggesting that proteolytic cleavage was involved. In vivo pulse/chase labeling experiments confirmed that the 110-kDa protein was derived from the full-length CDP/Cut protein. Proteolytic processing was weak or not detectable in G(0) and G(1) but increased in populations of cells enriched in S phase, and the appearance of the 110-kDa protein coincided with the increase in ATCGAT DNA binding. Interestingly, the amino-truncated and the full-length CDP/Cut isoforms exhibited different transcriptional properties in a reporter assay. We conclude that proteolytic processing of CDP/Cut at the G(1)/S transition generates a CDP/Cut isoform with distinct DNA binding and transcriptional activities. These findings, together with the cleavage of the Scc1 protein at mitosis, suggest that site-specific proteolysis may play an important role in the regulation of cell cycle progression.
authors.affiliation: Molecular Oncology Group, McGill University Health Center, McGill University, Montreal, Quebec, Canada H3A 1A1.
authors.forename: N S
authors.initials: NS
authors.lastname: Moon
authors.forename: P
authors.initials: P
authors.lastname: Premdas
authors.forename: M
authors.initials: M
authors.lastname: Truscott
authors.forename: L
authors.initials: L
authors.lastname: Leduy
authors.forename: G
authors.initials: G
authors.lastname: Bérubé
authors.forename: A
authors.initials: A
authors.lastname: Nepveu
mesh_terms: D000818:Animals; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004331:Drosophila melanogaster; D018398:Homeodomain Proteins; D006801:Humans; D009687:Nuclear Proteins / Q000378:metabolism*; D011485:Protein Binding; D012097:Repressor Proteins / Q000378:metabolism*; D016196:S Phase; D014157:Transcription Factors; D015533:Transcriptional Activation
chemical_list: C080632:CUX1 protein, human; D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; D009687:Nuclear Proteins; D012097:Repressor Proteins; D014157:Transcription Factors