Enhancement of BRCA1 E3 ubiquitin ligase activity through direct interaction with the BARD1 protein.
The breast and ovarian cancer-specific tumor suppressor RING finger protein BRCA1 has been identified as an E3 ubiquitin (Ub) ligase through in vitro studies, which demonstrated that its RING finger domain can autoubiquitylate and monoubiquitylate histone H2A when supplied with Ub, E1, and UBC4 (E2). Here we report that the E3 ligase activity of the N-terminal 110 amino acid residues of BRCA1, which encodes a stable domain containing the RING finger, as well as that of the full-length BRCA1, was significantly enhanced by the BARD1 protein (residues 8-142), whose RING finger domain itself lacked Ub ligase activity in vitro. The results of mutagenesis studies indicate that the enhancement of BRCA1 E3 ligase activity by BARD1 depends on direct interaction between the two proteins. Using K48A and K63A Ub mutants, we found that BARD1 stimulated the formation of both Lys(48)- and Lys(63)-linked poly-Ub chains. However, the enhancement of BRCA1 autoubiquitylation by BARD1 mostly resulted in poly-Ub chains linked through Lys(63), which could potentially activate biological pathways other than BRCA1 degradation. We also found that co-expression of BRCA1 and BARD1 in living cells increased the abundance and stability of both proteins and that this depended on their ability to heterodimerize.
Xia
Yan
Y
Molecular and Cell Biology Laboratory, The Salk Institute for Biological Studies, La Jolla, California 92037, USA.
Pao
Gerald M
GM
Chen
Hong-Wu
HW
Verma
Inder M
IM
Hunter
Tony
T
D019313:BRCA1 Protein / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D019281:Dimerization; D004789:Enzyme Activation; D006801:Humans; D008025:Ligases / Q000378:metabolism*; D011485:Protein Binding; D015398:Signal Transduction; D025521:Tumor Suppressor Proteins; D044767:Ubiquitin-Protein Ligases; D016335:Zinc Fingers
D019313:BRCA1 Protein; D002352:Carrier Proteins; D025521:Tumor Suppressor Proteins; C102449:BARD1 protein, human; D044767:Ubiquitin-Protein Ligases; D008025:Ligases