Docking of a specialized PIP Box onto chromatin-bound PCNA creates a degron for the ubiquitin ligase CRL4Cdt2.
Substrates of the E3 ubiquitin ligase CRL4(Cdt2), including Cdt1 and p21, contain a PCNA-binding motif called a PIP box. Upon binding of the PIP box to PCNA on chromatin, CRL4(Cdt2) is recruited and the substrate is ubiquitylated. Importantly, a PIP box cannot be sufficient for destruction, as most PIP box proteins are stable. Using Xenopus egg extracts, we identify two sequence elements in CRL4(Cdt2) substrates that promote their proteolysis: a specialized PIP box that confers exceptionally efficient PCNA binding and a basic amino acid 4 residues downstream of the PIP box, which recruits CRL4(Cdt2) to the substrate-PCNA complex. We also identify two mechanisms that couple CRL4(Cdt2)-dependent proteolysis to the chromatin-bound form of PCNA, ensuring that this proteolysis pathway is active only in S phase or after DNA damage. Thus, CRL4(Cdt2) recognizes an unusual degron, which is assembled specifically on chromatin via the binding of a specialized PIP box to PCNA.
Havens
Courtney G
CG
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA.
Walter
Johannes C
JC
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D002843:Chromatin / Q000378:metabolism*; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D045585:Flap Endonucleases / Q000235:genetics / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation; D008958:Models, Molecular; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D018809:Proliferating Cell Nuclear Antigen / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism*; D014981:Xenopus; D029867:Xenopus Proteins / Q000235:genetics / Q000378:metabolism*
C406819:Cdt1 protein, Xenopus; D018797:Cell Cycle Proteins; D002843:Chromatin; D050759:Cyclin-Dependent Kinase Inhibitor p21; D004268:DNA-Binding Proteins; D046912:Multiprotein Complexes; D018809:Proliferating Cell Nuclear Antigen; D029867:Xenopus Proteins; D044767:Ubiquitin-Protein Ligases; D045585:Flap Endonucleases