title: PIP degron proteins, substrates of CRL4Cdt2, and not PIP boxes, interfere with DNA polymerase η and κ focus formation on UV damage.
abstract: Proliferating cell nuclear antigen (PCNA) is a well-known scaffold for many DNA replication and repair proteins, but how the switch between partners is regulated is currently unclear. Interaction with PCNA occurs via a domain known as a PCNA-Interacting Protein motif (PIP box). More recently, an additional specialized PIP box has been described, the « PIP degron », that targets PCNA-interacting proteins for proteasomal degradation via the E3 ubiquitin ligase CRL4(Cdt2). Here we provide evidence that CRL4(Cdt2)-dependent degradation of PIP degron proteins plays a role in the switch of PCNA partners during the DNA damage response by facilitating accumulation of translesion synthesis DNA polymerases into nuclear foci. We show that expression of a nondegradable PIP degron (Cdt1) impairs both Pol η and Pol κ focus formation on ultraviolet irradiation and reduces cell viability, while canonical PIP box-containing proteins have no effect. Furthermore, we identify PIP degron-containing peptides from several substrates of CRL4(Cdt2) as efficient inhibitors of Pol η foci formation. By site-directed mutagenesis we show that inhibition depends on a conserved threonine residue that confers high affinity for PCNA-binding. Altogether these findings reveal an important regulative role for the CRL4(Cdt2) pathway in the switch of PCNA partners on DNA damage.
authors.affiliation: Genome Surveillance and Stability Laboratory, Department of Molecular Bases of Human Diseases, CNRS-UPR1142, Institute of Human Genetics, 141, rue de la cardonille, 34396 Cedex 5, Montpellier, France and Replication and Genome Dynamics Laboratory, Department of Genome Dynamics, CNRS-UPR1142, Institute of Human Genetics, 141, rue de la cardonille, 34396 Cedex 5, Montpellier, France.
authors.forename: Nikolay
authors.initials: N
authors.lastname: Tsanov
authors.forename: Chames
authors.initials: C
authors.lastname: Kermi
authors.forename: Philippe
authors.initials: P
authors.lastname: Coulombe
authors.forename: Siem
authors.initials: S
authors.lastname: Van der Laan
authors.forename: Dana
authors.initials: D
authors.lastname: Hodroj
authors.forename: Domenico
authors.initials: D
authors.lastname: Maiorano
mesh_terms: D000818:Animals; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016923:Cell Death; D002460:Cell Line; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000737:chemistry; D004249:DNA Damage; D004260:DNA Repair; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D011495:Histone-Lysine N-Methyltransferase / Q000737:chemistry; D006801:Humans; D051379:Mice; D041681:NIH 3T3 Cells; D009687:Nuclear Proteins / Q000378:metabolism*; D054730:Protein Interaction Domains and Motifs; D059748:Proteolysis; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*; D014466:Ultraviolet Rays; D000099270:Y-Family DNA Polymerases
chemical_list: D018797:Cell Cycle Proteins; D050759:Cyclin-Dependent Kinase Inhibitor p21; D004259:DNA-Directed DNA Polymerase; D011495:Histone-Lysine N-Methyltransferase; D009687:Nuclear Proteins; D044767:Ubiquitin-Protein Ligases; C467199:CDT1 protein, human; C429530:DTL protein, human; C404302:POLK protein, human; D000099270:Y-Family DNA Polymerases; C112338:Rad30 protein