title: Ubc9 mediates nuclear localization and growth suppression of BRCA1 and BRCA1a proteins.
abstract: BRCA1 gene mutations are responsible for hereditary breast and ovarian cancers. In sporadic breast tumors, BRCA1 dysfunction or aberrant subcellular localization is thought to be common. BRCA1 is a nuclear-cytoplasm shuttling protein and the reason for cytoplasmic localization of BRCA1 in young breast cancer patients is not yet known. We have previously reported BRCA1 proteins unlike K109R and cancer-predisposing mutant C61G to bind Ubc9 and modulate ER-α turnover. In the present study, we have examined the consequences of altered Ubc9 binding and knockdown on the subcellular localization and growth inhibitory function of BRCA1 proteins. Our results using live imaging of YFP, GFP, RFP-tagged BRCA1, BRCA1a and BRCA1b proteins show enhanced cytoplasmic localization of K109 R and C61G mutant BRCA1 proteins in normal and cancer cells. Furthermore, down-regulation of Ubc9 in MCF-7 cells using Ubc9 siRNA resulted in enhanced cytoplasmic localization of BRCA1 protein and exclusive cytoplasmic retention of BRCA1a and BRCA1b proteins. These mutant BRCA1 proteins were transforming and impaired in their capacity to inhibit growth of MCF-7 and CAL51 breast cancer cells. Interestingly, cytoplasmic BRCA1a mutants showed more clonogenicity in soft agar and higher levels of expression of Ubc9 than parental MCF7 cells. This is the first report demonstrating the physiological link between cytoplasmic mislocalization of mutant BRCA1 proteins, loss of ER-α repression, loss of ubiquitin ligase activity and loss of growth suppression of BRCA1 proteins. Thus, binding of BRCA1 proteins to nuclear chaperone Ubc9 provides a novel mechanism for nuclear import and control of tumor growth.
authors.affiliation: Cancer Biology Program, Department of OB/GYN, Morehouse School of Medicine, Georgia Cancer Center for Excellence, Grady Health System, Atlanta, Georgia 30303-3031, USA.
authors.forename: Yunlong
authors.initials: Y
authors.lastname: Qin
authors.forename: Jingyao
authors.initials: J
authors.lastname: Xu
authors.forename: Kartik
authors.initials: K
authors.lastname: Aysola
authors.forename: Nurjahan
authors.initials: N
authors.lastname: Begum
authors.forename: Vaishali
authors.initials: V
authors.lastname: Reddy
authors.forename: Yuli
authors.initials: Y
authors.lastname: Chai
authors.forename: William E
authors.initials: WE
authors.lastname: Grizzle
authors.forename: Edward E
authors.initials: EE
authors.lastname: Partridge
authors.forename: E Shyam P
authors.initials: ES
authors.lastname: Reddy
authors.forename: Veena N
authors.initials: VN
authors.lastname: Rao
mesh_terms: D021581:Active Transport, Cell Nucleus; D000818:Animals; D017209:Apoptosis; D019313:BRCA1 Protein / Q000235:genetics / Q000378:metabolism*; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D019556:COS Cells; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism*; D049109:Cell Proliferation; D002470:Cell Survival; D002522:Chlorocebus aethiops; D003593:Cytoplasm / Q000378:metabolism; D047628:Estrogen Receptor alpha / Q000378:metabolism; D005260:Female; D006801:Humans; D008856:Microscopy, Fluorescence; D017354:Point Mutation; D011485:Protein Binding; D034622:RNA Interference; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014162:Transfection; D044763:Ubiquitin-Conjugating Enzymes / Q000235:genetics / Q000378:metabolism*; D000098843:Ubiquitin-Conjugating Enzyme UBC9
chemical_list: D019313:BRCA1 Protein; C492913:BRCA1 protein, human; C506487:ESR1 protein, human; D047628:Estrogen Receptor alpha; D011993:Recombinant Fusion Proteins; D044763:Ubiquitin-Conjugating Enzymes; D000098843:Ubiquitin-Conjugating Enzyme UBC9