title: Mechanism of polyubiquitination by human anaphase-promoting complex: RING repurposing for ubiquitin chain assembly.
abstract: Polyubiquitination by E2 and E3 enzymes is a predominant mechanism regulating protein function. Some RING E3s, including anaphase-promoting complex/cyclosome (APC), catalyze polyubiquitination by sequential reactions with two different E2s. An initiating E2 ligates ubiquitin to an E3-bound substrate. Another E2 grows a polyubiquitin chain on the ubiquitin-primed substrate through poorly defined mechanisms. Here we show that human APC's RING domain is repurposed for dual functions in polyubiquitination. The canonical RING surface activates an initiating E2-ubiquitin intermediate for substrate modification. However, APC engages and activates its specialized ubiquitin chain-elongating E2 UBE2S in ways that differ from current paradigms. During chain assembly, a distinct APC11 RING surface helps deliver a substrate-linked ubiquitin to accept another ubiquitin from UBE2S. Our data define mechanisms of APC/UBE2S-mediated polyubiquitination, reveal diverse functions of RING E3s and E2s, and provide a framework for understanding distinctive RING E3 features specifying ubiquitin chain elongation.
authors.affiliation: Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
authors.forename: Nicholas G
authors.initials: NG
authors.lastname: Brown
authors.affiliation: Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA; Department of Microbiology, Immunology and Biochemistry, University of Tennessee Health Sciences Center, Memphis, TN 38163, USA.
authors.forename: Edmond R
authors.initials: ER
authors.lastname: Watson
authors.affiliation: Research Institute of Molecular Pathology (IMP), 1030 Vienna, Austria.
authors.forename: Florian
authors.initials: F
authors.lastname: Weissmann
authors.affiliation: Research Institute of Molecular Pathology (IMP), 1030 Vienna, Austria.
authors.forename: Marc A
authors.initials: MA
authors.lastname: Jarvis
authors.affiliation: Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA; Howard Hughes Medical Institute, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
authors.forename: Ryan
authors.initials: R
authors.lastname: VanderLinden
authors.affiliation: Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
authors.forename: Christy R R
authors.initials: CRR
authors.lastname: Grace
authors.affiliation: Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
authors.forename: Jeremiah J
authors.initials: JJ
authors.lastname: Frye
authors.affiliation: Research Institute of Molecular Pathology (IMP), 1030 Vienna, Austria.
authors.forename: Renping
authors.initials: R
authors.lastname: Qiao
authors.affiliation: Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany; Department of 3D Electron Cryomicroscopy, Institute of Microbiology and Genetics, Georg-August Universität, 37077 Göttingen, Germany.
authors.forename: Prakash
authors.initials: P
authors.lastname: Dube
authors.affiliation: Research Institute of Molecular Pathology (IMP), 1030 Vienna, Austria.
authors.forename: Georg
authors.initials: G
authors.lastname: Petzold
authors.affiliation: Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
authors.forename: Shein Ei
authors.initials: SE
authors.lastname: Cho
authors.affiliation: Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
authors.forename: Omar
authors.initials: O
authors.lastname: Alsharif
authors.affiliation: Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
authors.forename: Ju
authors.initials: J
authors.lastname: Bao
authors.affiliation: Research Institute of Molecular Pathology (IMP), 1030 Vienna, Austria.
authors.forename: Iain F
authors.initials: IF
authors.lastname: Davidson
authors.affiliation: Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
authors.forename: Jie J
authors.initials: JJ
authors.lastname: Zheng
authors.affiliation: Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
authors.forename: Amanda
authors.initials: A
authors.lastname: Nourse
authors.affiliation: NE-CAT, Bldg. 436E, Department of Chemistry and Chemical Biology, Cornell University, Argonne, IL 60439, USA.
authors.forename: Igor
authors.initials: I
authors.lastname: Kurinov
authors.affiliation: Research Institute of Molecular Pathology (IMP), 1030 Vienna, Austria. Electronic address: jan-michael.peters@imp.ac.at.
authors.forename: Jan-Michael
authors.initials: JM
authors.lastname: Peters
authors.affiliation: Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany; Department of 3D Electron Cryomicroscopy, Institute of Microbiology and Genetics, Georg-August Universität, 37077 Göttingen, Germany. Electronic address: hstark1@gwdg.de.
authors.forename: Holger
authors.initials: H
authors.lastname: Stark
authors.affiliation: Department of Structural Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA; Howard Hughes Medical Institute, St. Jude Children's Research Hospital, Memphis, TN 38105, USA. Electronic address: brenda.schulman@stjude.org.
authors.forename: Brenda A
authors.initials: BA
authors.lastname: Schulman
mesh_terms: D000595:Amino Acid Sequence; D064198:Apc11 Subunit, Anaphase-Promoting Complex-Cyclosome / Q000378:metabolism*; D064190:Apc2 Subunit, Anaphase-Promoting Complex-Cyclosome / Q000378:metabolism*; D064192:Apc4 Subunit, Anaphase-Promoting Complex-Cyclosome / Q000378:metabolism; D059447:Cell Cycle Checkpoints; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D046908:Peptide Biosynthesis, Nucleic Acid-Independent; D025821:Polyubiquitin / Q000096:biosynthesis* / Q000235:genetics; D017434:Protein Structure, Tertiary; D044763:Ubiquitin-Conjugating Enzymes / Q000378:metabolism*; D054875:Ubiquitination / Q000502:physiology*
chemical_list: D064192:Apc4 Subunit, Anaphase-Promoting Complex-Cyclosome; D025821:Polyubiquitin; C544962:Ube2S protein, human; D044763:Ubiquitin-Conjugating Enzymes; D064198:Apc11 Subunit, Anaphase-Promoting Complex-Cyclosome; D064190:Apc2 Subunit, Anaphase-Promoting Complex-Cyclosome