BRCA2 is ubiquitinated in vivo and interacts with USP11, a deubiquitinating enzyme that exhibits prosurvival function in the cellular response to DNA damage.
Individuals carrying a germ line mutation of the breast cancer susceptibility gene BRCA2 are predisposed to breast, ovarian, and other types of cancer. The BRCA2 protein has been proposed to function in the repair of DNA double-strand breaks. Using an immunopurification-mass spectrometry approach to identify novel proteins that associate with the BRCA2 gene product, we found that a deubiquitinating enzyme, USP11, formed specific complexes with BRCA2. Moreover, BRCA2 was constitutively ubiquitinated in vivo in the absence of detectable proteasomal degradation. Mitomycin C (MMC) led to decreased BRCA2 protein levels associated with increased ubiquitination, consistent with proteasome-dependent degradation. While BRCA2 could be deubiquitinated by USP11 in transient overexpression assays, a catalytically inactive USP11 mutant had no effect on BRCA2 ubiquitination or protein levels. Antagonism of USP11 function either through expression of this mutant or through RNA interference increased cellular sensitivity to MMC in a BRCA2-dependent manner. All of these results imply that BRCA2 expression levels are regulated by ubiquitination in the cellular response to MMC-induced DNA damage and that USP11 participates in DNA damage repair functions within the BRCA2 pathway independently of BRCA2 deubiquitination.
Derald H. Ruttenberg Cancer Center, Mount Sinai School of Medicine, One Gustave L. Levy Place, Box 1130, New York, NY 10029, USA.
Alan R
AR
Schoenfeld
Sarah
S
Apgar
Georgia
G
Dolios
Rong
R
Wang
Stuart A
SA
Aaronson
D000595:Amino Acid Sequence; D000818:Animals; D024682:BRCA2 Protein / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002470:Cell Survival; D003546:Cysteine Endopeptidases / Q000378:metabolism; D004249:DNA Damage; D006801:Humans; D046911:Macromolecular Substances; D051379:Mice; D016685:Mitomycin / Q000378:metabolism; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000378:metabolism; D019384:Nucleic Acid Synthesis Inhibitors / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex; D034622:RNA Interference; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D013869:Thiolester Hydrolases / Q000235:genetics / Q000378:metabolism*; D025801:Ubiquitin / Q000378:metabolism*; D015854:Up-Regulation
D024682:BRCA2 Protein; D046911:Macromolecular Substances; D009097:Multienzyme Complexes; D019384:Nucleic Acid Synthesis Inhibitors; D011993:Recombinant Fusion Proteins; C101359:USP11 protein, human; D025801:Ubiquitin; D016685:Mitomycin; D013869:Thiolester Hydrolases; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex