title: Cdc6 stability is regulated by the Huwe1 ubiquitin ligase after DNA damage.
abstract: The Cdc6 protein is an essential component of pre-replication complexes (preRCs), which assemble at origins of DNA replication during the G1 phase of the cell cycle. Previous studies have demonstrated that, in response to ionizing radiation, Cdc6 is ubiquitinated by the anaphase promoting complex (APC(Cdh1)) in a p53-dependent manner. We find, however, that DNA damage caused by UV irradiation or DNA alkylation by methyl methane sulfonate (MMS) induces Cdc6 degradation independently of p53. We further demonstrate that Cdc6 degradation after these forms of DNA damage is also independent of cell cycle phase, Cdc6 phosphorylation of the known Cdk target residues, or the Cul4/DDB1 and APC(Cdh1) ubiquitin E3 ligases. Instead Cdc6 directly binds a HECT-family ubiquitin E3 ligase, Huwe1 (also known as Mule, UreB1, ARF-BP1, Lasu1, and HectH9), and Huwe1 polyubiquitinates Cdc6 in vitro. Degradation of Cdc6 in UV-irradiated cells or in cells treated with MMS requires Huwe1 and is associated with release of Cdc6 from chromatin. Furthermore, yeast cells lacking the Huwe1 ortholog, Tom1, have a similar defect in Cdc6 degradation. Together, these findings demonstrate an important and conserved role for Huwe1 in regulating Cdc6 abundance after DNA damage.
authors.affiliation: Department of Biochemistry and Biophysics, School of Medicine and Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, NC 27599-7260, USA.
authors.forename: Jonathan R
authors.initials: JR
authors.lastname: Hall
authors.forename: Evelyn
authors.initials: E
authors.lastname: Kow
authors.forename: Kathleen R
authors.initials: KR
authors.lastname: Nevis
authors.forename: Chiajung Karen
authors.initials: CK
authors.lastname: Lu
authors.forename: K Scott
authors.initials: KS
authors.lastname: Luce
authors.forename: Qing
authors.initials: Q
authors.lastname: Zhong
authors.forename: Jeanette Gowen
authors.initials: JG
authors.lastname: Cook
mesh_terms: D064173:Anaphase-Promoting Complex-Cyclosome; D002384:Catalysis; D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000378:metabolism*; D018844:Cyclin-Dependent Kinases / Q000378:metabolism; D004249:DNA Damage; D006367:HeLa Cells; D006801:Humans; D009687:Nuclear Proteins / Q000378:metabolism*; D010766:Phosphorylation; D025821:Polyubiquitin / Q000378:metabolism; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D025521:Tumor Suppressor Proteins; D043743:Ubiquitin-Protein Ligase Complexes / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry / Q000378:metabolism*
chemical_list: C103706:CDC6 protein, human; D018797:Cell Cycle Proteins; D009687:Nuclear Proteins; D025521:Tumor Suppressor Proteins; D025821:Polyubiquitin; D043743:Ubiquitin-Protein Ligase Complexes; C502830:HUWE1 protein, human; D064173:Anaphase-Promoting Complex-Cyclosome; D044767:Ubiquitin-Protein Ligases; D018844:Cyclin-Dependent Kinases