ABSTRACT
Protection of the stalled replication fork is crucial for responding to replication stress and minimizing its impact on chromosome instability, thus preventing diseases, including cancer. We found a new component, Abro1, in the protection of stalled replication fork integrity. Abro1 deficiency results in increased chromosome instability, and Abro1-null mice are tumor-prone. We show that Abro1 protects stalled replication fork stability by inhibiting DNA2 nuclease/WRN helicase-mediated degradation of stalled forks. Depletion of RAD51 prevents the DNA2/WRN-dependent degradation of stalled forks in Abro1-deficient cells. This mechanism is distinct from the BRCA2-dependent fork protection pathway, in which stable RAD51 filament formation prevents MRE11-dependent degradation of the newly synthesized DNA at stalled forks. Thus, our data reveal a new aspect of regulated protection of stalled replication forks that involves Abro1.
Subject(s)
DNA Replication , Genomic Instability , Nuclear Matrix-Associated Proteins/physiology , Ubiquitin-Specific Proteases/physiology , Animals , BRCA2 Protein/genetics , Cell Line , Cells, Cultured , DNA/biosynthesis , DNA Helicases/physiology , Endodeoxyribonucleases/physiology , MRE11 Homologue Protein/physiology , Mice, Knockout , Multifunctional Enzymes/physiology , Neoplasms, Experimental/genetics , Nuclear Matrix-Associated Proteins/genetics , Rad51 Recombinase/genetics , Stress, Physiological , Ubiquitin-Specific Proteases/geneticsABSTRACT
Germline mutations of BRCA1 confer hereditary susceptibility to breast and ovarian cancer. However, somatic mutation of BRCA1 is infrequent in sporadic breast cancers. The BRCA1 protein C terminus (BRCT) domains interact with multiple proteins and are required for BRCA1's tumor-suppressor function. In this study, we demonstrated that Abraxas, a BRCA1 BRCT domain-interacting protein, plays a role in tumor suppression. Abraxas exerts its function through binding to BRCA1 to regulate DNA repair and maintain genome stability. Both homozygous and heterozygous Abraxas knockout mice exhibited decreased survival and increased tumor incidence. The gene encoding Abraxas suffers from gene copy loss and somatic mutations in multiple human cancers including breast, ovarian, and endometrial cancers, suggesting that mutation and loss of function of Abraxas may contribute to tumor development in human patients.
Subject(s)
BRCA1 Protein/metabolism , Breast Neoplasms/genetics , Carrier Proteins/metabolism , Genomic Instability , Ovarian Neoplasms/genetics , 3T3 Cells , Animals , BRCA1 Protein/chemistry , Breast Neoplasms/pathology , Carrier Proteins/genetics , DNA Repair , Female , Germ-Line Mutation , HEK293 Cells , Homozygote , Humans , Mice , Ovarian Neoplasms/pathology , Protein Binding , Protein Structure, TertiaryABSTRACT
BRCC36-deubiquitinating enzyme (DUB) forms two different complexes through interactions with two different adaptor proteins Abraxas and ABRO1 in cells. Abraxas mainly localizes in the nucleus, mediating the interaction of BRCC36 with BRCA1. ABRO1 is mainly localized in the cytoplasm. Because it lacks the BRCA1-interacting motif, the ABRO1 complex does not interact with BRCA1. Both BRCC36-containing complexes contain common components including BRE and NBA1/MERIT40. Here, we found that the two complexes are assembled in a similar manner and NBA1 and BRE interaction is critical for maintaining the integrity of both of the complexes. Knockdown of NBA1 or BRE leads to decreased levels of components of the two BRCC36-containing complexes. We provided evidence that NBA1 interacts with BRE through a C-terminal conserved motif of the NBA1 protein and a C-terminal UEV domain of the BRE protein. Furthermore, the NBA1-BRE interaction is required for cellular resistance to ionizing irradiation and NBA1's role in recruiting BRCA1 to DNA damage sites. Together, these studies reveal critical interactions required for the formation and function of BRCC36-containing DUB complexes.
