title: Monoubiquitination of histone H2B is intrinsic to the Bre1 RING domain-Rad6 interaction and augmented by a second Rad6-binding site on Bre1.
abstract: Ubiquitin signaling on chromatin is linked to diverse aspects of genome regulation, including gene expression and DNA repair. The yeast RING E3 ligase Bre1 combines with the E2 Rad6 to monoubiquitinate histone H2B during transcription. Little is known about how Bre1 directs Rad6 toward transferring only a single ubiquitin to a specific lysine residue. Using a defined in vitro system, we show that the Bre1 RING domain interaction with Rad6 is minimally sufficient to monoubiquitinate nucleosomes at histone H2B Lys-123. In addition, we reveal a cluster of charged residues on the Bre1 RING domain that is critical for recognizing the nucleosome surface. Notably, a second Rad6 binding domain of Bre1 interacts with the E2 backside and potentiates ubiquitin transfer to the substrate. Taken together, our study establishes a molecular framework for how distinct RING and non-RING E3 elements cooperate to regulate E2 reactivity and substrate selection during gene expression.
authors.affiliation: From the Max F. Perutz Laboratories, Medical University of Vienna, Vienna Biocenter (VBC), Dr. Bohr-Gasse 9/3, 1030 Vienna, Austria.
authors.forename: Eleonora
authors.initials: E
authors.lastname: Turco
authors.affiliation: From the Max F. Perutz Laboratories, Medical University of Vienna, Vienna Biocenter (VBC), Dr. Bohr-Gasse 9/3, 1030 Vienna, Austria.
authors.forename: Laura D
authors.initials: LD
authors.lastname: Gallego
authors.affiliation: From the Max F. Perutz Laboratories, Medical University of Vienna, Vienna Biocenter (VBC), Dr. Bohr-Gasse 9/3, 1030 Vienna, Austria.
authors.forename: Maren
authors.initials: M
authors.lastname: Schneider
authors.affiliation: From the Max F. Perutz Laboratories, Medical University of Vienna, Vienna Biocenter (VBC), Dr. Bohr-Gasse 9/3, 1030 Vienna, Austria alwin.koehler@mfpl.ac.at.
authors.forename: Alwin
authors.initials: A
authors.lastname: Köhler
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D006657:Histones / Q000378:metabolism*; D015151:Immunoblotting; D008239:Lysine / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D009707:Nucleosomes / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D025801:Ubiquitin / Q000378:metabolism; D044763:Ubiquitin-Conjugating Enzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination
chemical_list: C472168:Bre1 protein, S cerevisiae; D006657:Histones; D009707:Nucleosomes; D029701:Saccharomyces cerevisiae Proteins; D025801:Ubiquitin; C471606:RAD6 protein, S cerevisiae; D044763:Ubiquitin-Conjugating Enzymes; D008239:Lysine
keywords: Bre1; Chromatin; E3 Ubiquitin Ligase; Histone; Rad6; Ubiquitin; Ubiquitin-conjugating Enzyme (E2 Enzyme)