title: BRCA1-associated protein 1 interferes with BRCA1/BARD1 RING heterodimer activity.
abstract: The breast and ovarian tumor suppressor BRCA1 constitutes a RING heterodimer E3 ligase with BARD1. BRCA1-associated protein 1 (BAP1) is a ubiquitin COOH-terminal hydrolase that was initially identified as a protein that bound to the RING finger domain of BRCA1. However, how BAP1 contributes to the E3 activity of BRCA1/BARD1 is unclear. Here, we report that BAP1 interacts with BARD1 to inhibit the E3 ligase activity of BRCA1/BARD1. Domains comprised by residues 182-365 of BAP1 interact with the RING finger domain of BARD1, and surface plasmon resonance spectroscopy (BIAcore) analyses showed that BAP1 interferes with the BRCA1/BARD1 association. The perturbation resulted in inhibition of BRCA1 autoubiquitination and NPM1/B23 ubiquitination by BRCA1/BARD1. Although BAP1 was capable of deubiquitinating the polyubiquitin chains mediated by BRCA1/BARD1 in vitro, a catalytically inactive mutant of BAP1, C91S, still inhibited the ubiquitination in vitro and in vivo, implicating a second mechanism of action. Importantly, inhibition of BAP1 expression by short hairpin RNA resulted in hypersensitivity of the cells to ionizing irradiation and in retardation of S-phase progression. Together, these results suggest that BAP1 and BRCA1/BARD1 coordinately regulate ubiquitination during the DNA damage response and the cell cycle.
authors.affiliation: Division of Breast and Endocrine Surgery, Institute of Advanced Medical Science, and Department of Radiology, St. Marianna University School of Medicine, Kawasaki, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Nishikawa
authors.forename: Wenwen
authors.initials: W
authors.lastname: Wu
authors.forename: Ayaka
authors.initials: A
authors.lastname: Koike
authors.forename: Ryoko
authors.initials: R
authors.lastname: Kojima
authors.forename: Hiromichi
authors.initials: H
authors.lastname: Gomi
authors.forename: Mamoru
authors.initials: M
authors.lastname: Fukuda
authors.forename: Tomohiko
authors.initials: T
authors.lastname: Ohta
mesh_terms: D019313:BRCA1 Protein / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002461:Cell Line, Transformed; D015536:Down-Regulation; D006367:HeLa Cells; D006801:Humans; D007259:Infrared Rays; D000090243:Nucleophosmin; D034741:RNA, Small Interfering / Q000235:genetics; D016196:S Phase / Q000502:physiology / Q000528:radiation effects; D025521:Tumor Suppressor Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D025801:Ubiquitin / Q000378:metabolism; D043222:Ubiquitin Thiolesterase / Q000235:genetics / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000037:antagonists & inhibitors / Q000378:metabolism*
chemical_list: C400134:BAP1 protein, human; D019313:BRCA1 Protein; C492913:BRCA1 protein, human; C000716967:NPM1 protein, human; D034741:RNA, Small Interfering; D025521:Tumor Suppressor Proteins; D025801:Ubiquitin; D000090243:Nucleophosmin; C102449:BARD1 protein, human; D044767:Ubiquitin-Protein Ligases; D043222:Ubiquitin Thiolesterase