title: The BARD1 BRCT domain contributes to p53 binding, cytoplasmic and mitochondrial localization, and apoptotic function.
abstract: BARD1 is a breast cancer tumor suppressor with multiple domains and functions. BARD1 comprises a tandem BRCT domain at the C-terminus, and this sequence has been reported to target BARD1 to distinct subcellular locations such as nuclear DNA breakage sites and the centrosome through binding to regulatory proteins such as HP1 and OLA1, respectively. We now identify the BRCT domain as a binding site for p53. We first confirmed previous reports that endogenous BARD1 binds to p53 by immunoprecipitation assay, and further show that BARD1/p53 complexes locate at mitochondria suggesting a cellular location for p53 regulation of BARD1 apoptotic activity. We used a proximity ligation assay to map three distinct p53 binding sequences in human BARD1, ranging from weak (425-525) and modest (525-567) to strong (551-777 comprising BRCT domains). Deletion of the BRCT sequence caused major defects in the ability of BARD1 to (1) bind p53, (2) localize to the cytoplasm and mitochondria, and (3) induce Bax oligomerization and apoptosis. Our data suggest that BARD1 can move to mitochondria independent of p53, but subsequently associates with p53 to induce Bax clustering in part by decreasing mitochondrial Bcl-2 levels. We therefore identify a role for the BRCT domain in stimulating BARD1 nuclear export and mitochondrial localization, and in assembling mitochondrial BARD1/p53 complexes to regulate specific activities such as apoptotic function.
authors.affiliation: Centre for Cancer Research, The Westmead Millennium Institute for Medical Research, The University of Sydney, Westmead, NSW 2145, Australia.
authors.forename: Varsha
authors.initials: V
authors.lastname: Tembe
authors.affiliation: Centre for Cancer Research, The Westmead Millennium Institute for Medical Research, The University of Sydney, Westmead, NSW 2145, Australia.
authors.forename: Estefania
authors.initials: E
authors.lastname: Martino-Echarri
authors.affiliation: Centre for Cancer Research, The Westmead Millennium Institute for Medical Research, The University of Sydney, Westmead, NSW 2145, Australia.
authors.forename: Kamila A
authors.initials: KA
authors.lastname: Marzec
authors.affiliation: Centre for Cancer Research, The Westmead Millennium Institute for Medical Research, The University of Sydney, Westmead, NSW 2145, Australia.
authors.forename: Myth T S
authors.initials: MT
authors.lastname: Mok
authors.affiliation: Centre for Cancer Research, The Westmead Millennium Institute for Medical Research, The University of Sydney, Westmead, NSW 2145, Australia.
authors.forename: Kirsty M
authors.initials: KM
authors.lastname: Brodie
authors.affiliation: Centre for Cancer Research, The Westmead Millennium Institute for Medical Research, The University of Sydney, Westmead, NSW 2145, Australia.
authors.forename: Kate
authors.initials: K
authors.lastname: Mills
authors.affiliation: Centre for Cancer Research, The Westmead Millennium Institute for Medical Research, The University of Sydney, Westmead, NSW 2145, Australia.
authors.forename: Ying
authors.initials: Y
authors.lastname: Lei
authors.affiliation: Centre for Cancer Research, The Westmead Millennium Institute for Medical Research, The University of Sydney, Westmead, NSW 2145, Australia.
authors.forename: Anna
authors.initials: A
authors.lastname: DeFazio
authors.affiliation: Centre for Cancer Research, The Westmead Millennium Institute for Medical Research, The University of Sydney, Westmead, NSW 2145, Australia.
authors.forename: Helen
authors.initials: H
authors.lastname: Rizos
authors.affiliation: Electron Microscopy Unit, Institute for Clinical Pathology and Medical Research, Westmead Hospital, Westmead, NSW 2145, Australia.
authors.forename: Emma
authors.initials: E
authors.lastname: Kettle
authors.affiliation: Electron Microscopy Unit, Institute for Clinical Pathology and Medical Research, Westmead Hospital, Westmead, NSW 2145, Australia.
authors.forename: Ross
authors.initials: R
authors.lastname: Boadle
authors.affiliation: Centre for Cancer Research, The Westmead Millennium Institute for Medical Research, The University of Sydney, Westmead, NSW 2145, Australia. Electronic address: beric.henderson@sydney.edu.au.
authors.forename: Beric R
authors.initials: BR
authors.lastname: Henderson
mesh_terms: D000595:Amino Acid Sequence; D017209:Apoptosis; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D003593:Cytoplasm / Q000235:genetics / Q000378:metabolism*; D053960:DNA Breaks; D005260:Female; D006801:Humans; D061986:MCF-7 Cells; D008928:Mitochondria / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000235:genetics; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism*; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism*; D051028:bcl-2-Associated X Protein / Q000235:genetics / Q000378:metabolism
chemical_list: C494826:BAX protein, human; C000595707:BCL2 protein, human; D019253:Proto-Oncogene Proteins c-bcl-2; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; D025521:Tumor Suppressor Proteins; D051028:bcl-2-Associated X Protein; C102449:BARD1 protein, human; D044767:Ubiquitin-Protein Ligases
keywords: Apoptosis; BARD1; BCL-2; Mitochondria; p53