title: Genetic analysis of BRCA1 ubiquitin ligase activity and its relationship to breast cancer susceptibility.
abstract: The N-terminus of the Breast Cancer-1 predisposition protein (BRCA1) associates with the BRCA1-associated RING domain-1 protein (BARD1) to form a heterodimer, which exhibits ubiquitin ligase activity that is abrogated by known cancer-associated BRCA1 missense mutations. The majority of missense substitutions identified in patients with a personal or a family history of disease have not been followed in pedigrees, nor there is a functional understanding of their impact. We have examined, by extensive missense substitution, the interaction of BRCA1 with components that contribute to its ubiquitin ligase activity, BARD1 and the E2 ubiquitin-conjugating enzyme, UbcH5a. Selection from a randomly generated library of BRCA1 missense mutations for variants that inhibit the interaction with these components identified substitutions in residues found altered in patient DNA, indicating a correlation between loss of component-binding and propensity to disease development. We further show that the BRCA1:E2 interaction is sensitive to substitutions in all structural elements of the BRCA1 N-terminus, whereas the BARD1 interaction is sensitive to a subset of BRCA1 substitutions, which also inhibit E2-binding. Patient variants that inhibit the BRCA1:E2 interaction show loss of ubiquitin ligase activity and correlate with disease susceptibility and theoretical predictions of pathogenicity. These data link the loss of ubiquitin ligase activity, through loss of E2-binding, to the majority of non-polymorphic patient variants described within the N-terminus of BRCA1 and illustrate the likely significant role of BRCA1 ubiquitin ligase activity in tumour suppression.
authors.affiliation: Department of Medical and Molecular Genetics, Kings College London School of Medicine at Guy's, Kings and St Thomas' Hospitals, Guy's Hospital, 8th Floor Guy's Tower, St Thomas' Street, London SE1 9RT, UK. jo.morris@genetics.kcl.ac.uk
authors.forename: Joanna R
authors.initials: JR
authors.lastname: Morris
authors.forename: Laurent
authors.initials: L
authors.lastname: Pangon
authors.forename: Chris
authors.initials: C
authors.lastname: Boutell
authors.forename: Toyomasa
authors.initials: T
authors.lastname: Katagiri
authors.forename: Nicholas H
authors.initials: NH
authors.lastname: Keep
authors.forename: Ellen
authors.initials: E
authors.lastname: Solomon
mesh_terms: D019313:BRCA1 Protein / Q000235:genetics* / Q000378:metabolism; D019281:Dimerization; D005260:Female; D020022:Genetic Predisposition to Disease; D006801:Humans; D033862:Iron-Binding Proteins / Q000235:genetics / Q000378:metabolism; D008297:Male; D046912:Multiprotein Complexes / Q000235:genetics* / Q000378:metabolism; D020125:Mutation, Missense; D009369:Neoplasms / Q000201:enzymology / Q000235:genetics*; D011485:Protein Binding / Q000235:genetics; D017434:Protein Structure, Tertiary / Q000235:genetics; D025521:Tumor Suppressor Proteins / Q000235:genetics* / Q000378:metabolism; D044763:Ubiquitin-Conjugating Enzymes / Q000235:genetics / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000235:genetics* / Q000378:metabolism
chemical_list: D019313:BRCA1 Protein; D033862:Iron-Binding Proteins; D046912:Multiprotein Complexes; D025521:Tumor Suppressor Proteins; C097308:UBE2D1 protein, human; D044763:Ubiquitin-Conjugating Enzymes; C102449:BARD1 protein, human; D044767:Ubiquitin-Protein Ligases