BRCA1 Tumor Suppressor Research Articles

ANALYSIS OF THE mTOR PATHWAY IN OVARIES OF WOMEN WITH BRCA MUTATIONS

Mutations in the tumor suppressor genes BRCA1 and BRCA2 predispose women to cancer and are associated with diminished ovarian reserve through a known impairment in DNA repair function. The PI3K/AKT/mTOR pathway regulates primordial follicle activation, and its dysregulation is implicated in human cancers including BRCA-related ovarian cancers. We sought to investigate whether mTOR activity is upregulated in follicles of women with BRCA mutations, suggesting a potential factor contributing to diminished ovarian reserve.

BRCA2 BRC missense variants disrupt RAD51-dependent DNA repair.

Pathogenic mutations in the BRCA2 tumor suppressor gene predispose to breast, ovarian, pancreatic, prostate, and other cancers. BRCA2 maintains genome stability through homology-directed repair (HDR) of DNA double-strand breaks (DSBs) and replication fork protection. Nonsense or frameshift mutations leading to truncation of the BRCA2 protein are typically considered pathogenic; however, missense mutations resulting in single amino acid substitutions can be challenging to functionally interpret. The majority of missense mutations in BRCA2 have been classified as Variants of Uncertain Significance (VUS) with unknown functional consequences. In this study, we identified three BRCA2 VUS located within the BRC repeat region to determine their impact on canonical HDR and fork protection functions. We provide evidence that S1221P and T1980I, which map to conserved residues in the BRC2 and BRC7 repeats, compromise the cellular response to chemotherapeutics and ionizing radiation, and display deficits in fork protection. We further demonstrate biochemically that S1221P and T1980I disrupt RAD51 binding and diminish the ability of BRCA2 to stabilize RAD51-ssDNA complexes. The third variant, T1346I, located within the spacer region between BRC2 and BRC3 repeats, is fully functional. We conclude that T1346I is a benign allele, whereas S1221P and T1980I are hypomorphic disrupting the ability of BRCA2 to fully engage and stabilize RAD51 nucleoprotein filaments. Our results underscore the importance of correctly classifying BRCA2 VUS as pathogenic variants can impact both future cancer risk and guide therapy selection during cancer treatment.

Implicações éticas dos testes genéticos de suscetibilidade ao câncer de mama

Abstract Breast cancer is a public health problem because it is the malignant neoplasm with the highest incidence in women worldwide. The hereditary form corresponds to about 5% to 10% of all cases and is directly related to the inheritance of genetic mutations. The main ones occur in the BRCA1 and BRCA2 tumor suppressor genes. The identification of these mutations is extremely important because of the high risk of breast cancer development in this population, allowing differentiated screening strategies and the adoption of risk reduction measures. However, reflections on the ethical aspects related to the indiscriminate use of genetic testing are important and necessary. The objective of this study was to evaluate the knowledge and opinion of physicians of an oncology reference center on the indication of genetic tests for susceptibility to breast cancer given the ethical dilemmas to which they are submitted in medical practice.

Management Strategies of Breast Cancer Patients with BRCA1 and BRCA2 Pathogenic Germline Variants.

Most of breast cancer cases are sporadic; however, 15–20% are associated with family history, and some are inherited. Among those, deleterious mutations in BRCA1 and BRCA2 tumor suppressor genes are the most commonly encountered pathogenic germline variants (PGVs). Given the availability and affordability of multi-gene panel sequencing technologies, testing for PGVs is commonly practiced. With our enhanced understanding of cancer genetics and specific molecular alterations, the better acceptance of risk-directed screening and prevention, and the recent introduction of novel targeted therapies, management of BRCA-positive breast cancers is taking a new direction, focusing more on risk-reducing interventions, including mastectomy and salpingo-oophorectomy, and incorporating special treatment regimens, including platinum-based chemotherapy, and the recently-introduced PARP (poly (ADP)-ribose polymerase) inhibitors. Given the recent advances in reproductive technology and molecular medicine, younger women with PGVs may have the option of embryo selection through preimplantation genetic testing and diagnosis, thus preventing the potential transmission of the implicated genes to the next generations. In this review, we cover the clinical implications of identifying a pathogenic germline mutation in BRCA1 and BRCA2 genes in breast cancer patients, and their relatives, across the continuum of care – from cancer prevention and early detection, through active treatment and up to survivorship issues.

PSY8-1 Management of HBOC: from the view of genomic medicine

HBOC is a tumor susceptibility syndrome caused by a germline pathogenic variant of tumor suppressor gene BRCA1 or BRCA2. Hereditary tumors due to pathogenic variants in a single gene account for 5-10% of all cancers, and HBOC can be a model for the future management of hereditary tumors. The reasons for this are as follows.

Ovarian tumors and genetic predisposition.

Summary of knowledge in the field of ovarian cancer and genetic predisposition. Ovarian tumors are usually diagnosed at advanced stages of the disease and the prognosis for these patients is generally poor. The 5-year overall survival rate, regardless of the histopathological type of tumor, is around 44%. Germline mutations causing hereditary tumor syndromes are predominantly involved in the development of epithelial ovarian tumors. The most common is hereditary breast and ovarian cancer syndrome, which is caused by germline mutations in the tumor suppressor genes BRCA1 and BRCA2. Several other tumor suppressor genes and oncogenes are known to be associated with ovarian tumors and cause other types of tumor syndromes. Inherited tumor syndromes include Lynch syndrome, Peutz-Jegers syndrome, Gorlin syndrome, Li-Fraumeni syndrome and others. The indication for genetic examination of germline mutations is given by a clinical geneticist on the basis of the recommendation of the attending physician. At present, every ovarian tumor, primary peritoneal tumor and tube tumor diagnosed at any age is indicated for genetic testing. Early identification of genes for hereditary cancer syndromes, thanks to rapidly developing molecular genetic methods, is an important step towards personalized treatment of ovarian cancer and preventive measures in families at risk. It is also important to note that a negative molecular genetic test result does not exclude genetic risk.

WRN rescues replication forks compromised by a BRCA2 deficiency: Predictions for how inhibition of a helicase that suppresses premature aging tilts the balance to fork demise and chromosomal instability in cancer.

Hereditary breast and ovarian cancers are frequently attributed to germline mutations in the tumor suppressor genes BRCA1 and BRCA2. BRCA1/2 act to repair double-strand breaks (DSBs) and suppress the demise of unstable replication forks. Our work elucidated a dynamic interplay between BRCA2 and the WRN DNA helicase/exonuclease defective in the premature aging disorder Werner syndrome. WRN and BRCA2 participate in complementary pathways to stabilize replication forks in cancer cells, allowing them to proliferate. Whether the functional overlap of WRN and BRCA2 is relevant to replication at gaps between newly synthesized DNA fragments, protection of telomeres, and/or metabolism of secondary DNA structures remain to be determined. Advances in understanding the mechanisms elicited during replication stress have prompted the community to reconsider avenues for cancer therapy. Insights from studies of PARP or topoisomerase inhibitors provide working models for the investigation of WRN's mechanism of action. We discuss these topics, focusing on the implications of the WRN-BRCA2 genetic interaction under conditions of replication stress.

Kinase Activity of PAR1b, Which Mediates Nuclear Translocation of the BRCA1 Tumor Suppressor, Is Potentiated by Nucleic Acid-Mediated PAR1b Multimerization.

PAR1b is a cytoplasmic serine/threonine kinase that controls cell polarity and cell–cell interaction by regulating microtubule stability while mediating cytoplasmic-to-nuclear translocation of BRCA1. PAR1b is also a cellular target of the CagA protein of Helicobacter pylori, which leads to chronic infection causatively associated with the development of gastric cancer. The CagA-PAR1b interaction inactivates the kinase activity of PAR1b and thereby dampens PAR1b-mediated BRCA1 phosphorylation, which reduces the level of nuclear BRCA1 and thereby leads to BRCAness and BRCAness-associated genome instability underlying gastric carcinogenesis. While PAR1b can multimerize within the cells, little is known about the mechanism and functional role of PAR1b multimerization. We found in the present study that PAR1b was multimerized in vitro by binding with nucleic acids (both single- and double-stranded DNA/RNA) via the spacer region in a manner independent of nucleic-acid sequences, which markedly potentiated the kinase activity of PAR1b. Consistent with these in vitro observations, cytoplasmic introduction of double-stranded DNA or expression of single-stranded RNA increased the PAR1b kinase activity in the cells. These findings indicate that the cytoplasmic DNA/RNA contribute to nuclear accumulation of BRCA1 by constitutively activating/potentiating cytoplasmic PAR1b kinase activity, which is subverted in gastric epithelial cells upon delivery of H. pylori CagA oncoprotein.

BRCA1 haploinsufficiency promotes chromosomal amplification under Fenton reaction-based carcinogenesis through ferroptosis-resistance

Germline-mutation in BRCA1 tumor suppressor gene is an established risk for carcinogenesis not only in females but also in males. Deficiency in the repair of DNA double-strand breaks is hypothesized as a responsible mechanism for carcinogenesis. However, supporting data is insufficient both in the mutation spectra of cancers in the patients with BRCA1 germline-mutation and in murine knockout/knock-in models of Brca1 haploinsufficiency. Furthermore, information on the driving force toward carcinogenesis in BRCA1 mutation carriers is lacking. Here we applied Fenton reaction-based renal carcinogenesis to a rat heterozygously knockout model of BRCA1 haploinsufficiency (mutant [MUT] model; L63X/+). Rat MUT model revealed significant promotion of renal cell carcinoma (RCC) induced by ferric nitrilotriacetate (Fe-NTA). Array-based comparative genome hybridization of the RCCs identified significant increase in chromosomal amplification, syntenic to those in breast cancers of BRCA1 mutation carriers, including c-Myc, in comparison to those in the wild-type. Subacute-phase analysis of the kidney after repeated Fe-NTA treatment in the MUT model revealed dysregulated iron metabolism with mitochondrial malfunction assessed by expression microarray and electron microscopy, leading to renal tubular proliferation with iron overload. In conclusion, we for the first time demonstrate that biallelic wild-type BRCA1 provides more robust protection for mitochondrial metabolism under iron-catalyzed oxidative stress, preventing the emergence of neoplastic cells with chromosomal amplification. Our results suggest that oxidative stress via excess iron is a major driving force for carcinogenesis in BRCA1 haploinsufficiency, which can be a target for cancer prevention and therapeutics.

Aging accelerates while multiparity delays tumorigenesis in mouse models of high-grade serous carcinoma

ObjectivesThe “incessant ovulation” hypothesis links increased risk for tubo-ovarian high-grade serous carcinoma (HGSC) due to more ovulations and reduced risk conferred by pre-menopausal exposures like oral contraceptive use, multiparity, and breastfeeding. However, most women diagnosed with HGSC are postmenopausal, implying age is a major risk factor for HGSC. Our mouse model for HGSC, based on tamoxifen (TAM)-induced somatic inactivation of the Brca1, Trp53, Rb1, and Nf1 (BPRN) tumor suppressor genes in oviductal epithelium, recapitulates key genetic, histopathologic, and biological features of human HGSCs. We aimed to credential the model for future efforts to define biological and risk modification factors in HGSC pathogenesis. MethodsBPRN mice were treated with TAM to induce tumors at defined ages and parity status. ResultsBPRN mice aged 9-months prior to tumor induction had markedly shorter survival than 6–8 week old mice induced to form tumors (median 46.5 weeks versus 61.5 weeks, log-rank test P = 0.0006). No significant differences in cancer phenotypes were observed between multiparous versus nulliparous BPRN mice. However, using a modified tumor model with one wild-type Nf1 allele (BPRNfl/+), nulliparous mice had more advanced tumors than multiparous mice (Mantel-Haenszel Chi-square test of association, P = 0.01). ConclusionsOur findings show aging is associated with significantly shortened survival post tumor induction in the BRPN model and multiparity delays development and/or progression of HGSC in certain genetic contexts. The findings support relevance of our mouse model to gain mechanistic insights into how known factors exert their protective effects and to test novel approaches for HGSC prevention.

BRCA2-DSS1 interaction is dispensable for RAD51 recruitment at replication-induced and meiotic DNA double strand breaks

The interaction between tumor suppressor BRCA2 and DSS1 is essential for RAD51 recruitment and repair of DNA double stand breaks (DSBs) by homologous recombination (HR). We have generated mice with a leucine to proline substitution at position 2431 of BRCA2, which disrupts this interaction. Although a significant number of mutant mice die during embryogenesis, some homozygous and hemizygous mutant mice undergo normal postnatal development. Despite lack of radiation induced RAD51 foci formation and a severe HR defect in somatic cells, mutant mice are fertile and exhibit normal RAD51 recruitment during meiosis. We hypothesize that the presence of homologous chromosomes in close proximity during early prophase I may compensate for the defect in BRCA2-DSS1 interaction. We show the restoration of RAD51 foci in mutant cells when Topoisomerase I inhibitor-induced single strand breaks are converted into DSBs during DNA replication. We also partially rescue the HR defect by tethering the donor DNA to the site of DSBs using streptavidin-fused Cas9. Our findings demonstrate that the BRCA2-DSS1 complex is dispensable for RAD51 loading when the homologous DNA is close to the DSB.

Abstract P3-12-24: Landscape of germline BRCA1 and BRCA2 mutations in breast cancer in Peru

Abstract BACKGROUND: Breast cancer (BC) is the most common cancer and the second leading cause of cancer-related deaths worldwide. Mutations in tumor suppressor genes BRCA1 and BRCA2 are the most common cause of hereditary breast and ovarian cancer. Additionally, carriers of germline BRCA1/2 mutations also have an elevated risk of other malignancies. Identification of mutations within these genes is essential for determining early cancer detection and risk-reducing strategies in carriers and establishing a therapeutic approach in breast cancer patients. The aims of this study were to evaluate the prevalence and spectrumof germline BRCA1 and BRCA2 variants in peruvian breast cancer patients. METHODS: Patients with HER2Neu negative BC referred to ONCOGENOMICS laboratory from 2019 to2021 to assessed poly (adenosine diphosphate-ribose) polymerase (PARP) inhibitor treatment were included in the study. Blood samples were collected to assess the status of germline BRCA1 and BRCA2. Next Generation Sequencing (NGS) was performed using the Ampliseq for Illumina BRCA panel and run on the Illumina MiSeq. The Human Genome Variation Society (HGVS) nomenclature guidelines (http://varnomen.hgvs.org/) were used to annotate identified variants and the ClinVar database (www.ncbi.nlm.nih.gov/clinvar/) was used to determine the clinical significance of all reported variants. Current American College of Medical Genetics (ACMG) guidelines were also used for further classification. RESULTS: A total of 155 HER2Neu negative BC cases were evaluated. Median age at BRCA1/2 evaluation was 51 (23-82) years, 78.1% (121) were negative HR status (triple-negative BC, TNBC); 21.9% (34) were positive HR status; 33.5% (52) were clinical stage III and 66.5% (103) were clinical stage IV. Most patients were from Lima-Callao (65.8%) followed by patients from the coast (23.9%), highlands (9%) and rainforest(1.3%). Only 18.7% of patients were from public health centers. The prevalence of pathogenic (P) and likely pathogenic (LP) variants in BRCA1/2 was 13.5% (21) (13 in BRCA1 and 8 in BRCA2). The pathogenic variant in BRCA1 c.2105dupT (p.Leu702Phefs*10) was found in 3 cases (2 Lima-Callao and 1 coast). All variants identified in BRCA1 were in TNBC. In BRCA2, half of pathogenic variants were found in TNBC. The prevalence of variants of uncertain significance (VUS) was 6.4% (10). In BRCA2, we identified 7 VUS and the variant c.5465A>T (p.Asn1822Ile) was found in 3 cases with positive HR status. Only one VUS was found in BRCA1. CONCLUSION: BRCA1 and BRCA2 germline mutations were identified in 13.5% of peruvian HER2Neu negative BC patients. A higher rate of P/LP variants was found in BRCA1. TNBC patients demostrated a higher prevalence in BRCA1 variants. Citation Format: Joseph A Pinto, Yomali Ferreyra, Alicia M Cock-Rada, Franco Doimi, Jhoysi Casas, Jhajaira Araujo, Gina Rosas, Leny Bravo, Carolina Belmar-López. Landscape of germline BRCA1 and BRCA2 mutations in breast cancer in Peru [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P3-12-24.

BARD1 is an ATPase activating protein for OLA1

OLA1 is a P-loop ATPase, implicated in centrosome duplication through the interactions with tumor suppressors BRCA1 and BARD1. Disruption of the interaction of OLA1 with BARD1 results in centrosome amplification. However, the molecular interplay and mechanism of the OLA1-BARD1 complex remain elusive. Here, we use a battery of biophysical, biochemical, and structural analyses to elucidate the molecular basis of the OLA1-BARD1 interaction. Our structural and enzyme kinetics analyses show this nucleotide-dependent interaction enhances the ATPase activity of OLA1 by increasing the turnover number (kcat). Unlike canonical GTPase activating proteins that act directly on the catalytic G domain, the BARD1 BRCT domain binds to the OLA1 TGS domain via a highly conserved BUDR motif. A cancer related mutation V695L on BARD1 is known to associate with centrosome abnormality. The V695L mutation reduces the BARD1 BRCT-mediated activation of OLA1. Crystallographic snapshot of the BRCT V695L mutant at 1.88 Å reveals this mutation perturbs the OLA1 binding site, resulting in reduced interaction. Altogether, our findings suggest the BARD1 BRCT domain serves as an ATPase activating protein to control OLA1 allosterically.

PARP Inhibitors: From the Mechanism of Action to Clinical Practice

Repairing damage and errors that occur in the DNA molecule is essential to maintain the integrity of the genome and cell viability. Deficits in DNA repair mechanisms lead to an increased risk of genetic instability and contribute to neoplastic transformation. Poly (ADP-ribose) polymerases (PARP) are a group of enzymes that play a key role in signalling and repairing DNA errors. The inhibition of its activity is a therapeutic strategy that takes advantage of the mechanism of synthetic lethality and that can be used in the treatment of tumours with specific defects in DNA repair pathways, namely in tumours with mutations in the tumour suppressor genes BRCA1 and BRCA2. There are several PARP inhibitors (iPARP), already approved by the USA Food and Drug Administration and the European Medicines Agency used in the treatment of breast, ovarian, pancreatic and prostate cancer. However, as with other target therapies, despite being well tolerated and widely used in the clinical practice, iPARP resistance is common and can be developed through various molecular mechanisms. In this article, we intend to make an updated review on iPARP and its main role in tumour cells, highlighting the several resistance mechanisms that have been recently revealed, as well as the current clinical applications and toxicity associated with this target therapy.

CRL4-DCAF8L1 Regulates BRCA1 and BARD1 Protein Stability.

BRCA1 is frequently down-regulated in breast cancer, the underlying mechanism is unclear. Here we identified DCAF8L1, an X-linked gene product, as a DDB1-Cullin associated Factor (DCAF) for CUL4 E3 ligases to target BRCA1 and BARD1 for proteasomal degradation. Forced expression of DCAF8L1 caused reduction of BRCA1 and BARD1, and impaired DNA damage repair function, conferring increased sensitivity to irradiation and DNA damaging agents, as well as Olaparib, a PARPi anticancer drug; while depletion of DCAF8L1 restored BRCA1 and suppressed the growth of its xenograft tumors. Furthermore, the expression of DCAF8L1 was induced in human H9 ES cells during transition from primed to naïve state when Xi chromosome was reactivated. Aberrant expression of DCAF8L1 was observed in human breast fibroadenoma and breast cancer. These findings suggest that CRL4DCAF8L1 is an important E3 ligase that may participate in the development of breast cancer, probably through regulating the stability of BRCA1 and BARD1 tumor suppressor, linking BRCA1 and X chromosome inactivation to breast carcinogenesis.

DNA fiber analyses to study functional importance of helicases and associated factors during replication stress.

Helicases, DNA translocases, nucleases and DNA-binding proteins play integral roles in protecting replication forks in human cells. Perturbations to replication fork dynamics can be caused by genetic loss of key factor(s) or exposure to replication stress inducing agents that perturb the nucleotide pool, stabilize unusual DNA secondary structures, or inhibit protein function (typically catalytic activity performed by a DNA polymerase, nuclease or helicase). DNA fiber analysis is a highly resourceful and facile experimental approach to study the molecular dynamics of replication forks in living cells. In this chapter, we provide a detailed list of reagents, equipment and experimental strategies to perform DNA fiber experiments. We have utilized these approaches to characterize the role of the Werner syndrome helicase (WRN) to protect replication forks in cells that are deficient in the tumor suppressor and genome stability factor BRCA2.

A BRCA1 Coiled-Coil Domain Variant Disrupting PALB2 Interaction Promotes the Development of Mammary Tumors and Confers a Targetable Defect in Homologous Recombination Repair.

The BRCA1 tumor suppressor gene encodes a multidomain protein for which several functions have been described. These include a key role in homologous recombination repair (HRR) of DNA double-strand breaks, which is shared with two other high-risk hereditary breast cancer suppressors, BRCA2 and PALB2. Although both BRCA1 and BRCA2 interact with PALB2, BRCA1 missense variants affecting its PALB2-interacting coiled-coil domain are considered variants of uncertain clinical significance (VUS). Using genetically engineered mice, we show here that a BRCA1 coiled-coil domain VUS, Brca1 p.L1363P, disrupts the interaction with PALB2 and leads to embryonic lethality. Brca1 p.L1363P led to a similar acceleration in the development of Trp53-deficient mammary tumors as Brca1 loss, but the tumors showed distinct histopathologic features, with more stable DNA copy number profiles in Brca1 p.L1363P tumors. Nevertheless, Brca1 p.L1363P mammary tumors were HRR incompetent and responsive to cisplatin and PARP inhibition. Overall, these results provide the first direct evidence that a BRCA1 missense variant outside of the RING and BRCT domains increases the risk of breast cancer. SIGNIFICANCE: These findings reveal the importance of a patient-derived BRCA1 coiled-coil domain sequence variant in embryonic development, mammary tumor suppression, and therapy response.See related commentary by Mishra et al., p. 6080.

WRN helicase safeguards deprotected replication forks in BRCA2-mutated cancer cells

The tumor suppressor BRCA2 protects stalled forks from degradation to maintain genome stability. However, the molecular mechanism(s) whereby unprotected forks are stabilized remains to be fully characterized. Here, we demonstrate that WRN helicase ensures efficient restart and limits excessive degradation of stalled forks in BRCA2-deficient cancer cells. In vitro, WRN ATPase/helicase catalyzes fork restoration and curtails MRE11 nuclease activity on regressed forks. We show that WRN helicase inhibitor traps WRN on chromatin leading to rapid fork stalling and nucleolytic degradation of unprotected forks by MRE11, resulting in MUS81-dependent double-strand breaks, elevated non-homologous end-joining and chromosomal instability. WRN helicase inhibition reduces viability of BRCA2-deficient cells and potentiates cytotoxicity of a poly (ADP)ribose polymerase (PARP) inhibitor. Furthermore, BRCA2-deficient xenograft tumors in mice exhibited increased DNA damage and growth inhibition when treated with WRN helicase inhibitor. This work provides mechanistic insight into stalled fork stabilization by WRN helicase when BRCA2 is deficient.

BRCA2 associates with MCM10 to suppress PRIMPOL-mediated repriming and single-stranded gap formation after DNA damage

The BRCA2 tumor suppressor protects genome integrity by promoting homologous recombination-based repair of DNA breaks, stability of stalled DNA replication forks and DNA damage-induced cell cycle checkpoints. BRCA2 deficient cells display the radio-resistant DNA synthesis (RDS) phenotype, however the mechanism has remained elusive. Here we show that cells without BRCA2 are unable to sufficiently restrain DNA replication fork progression after DNA damage, and the underrestrained fork progression is due primarily to Primase-Polymerase (PRIMPOL)-mediated repriming of DNA synthesis downstream of lesions, leaving behind single-stranded DNA gaps. Moreover, we find that BRCA2 associates with the essential DNA replication factor MCM10 and this association suppresses PRIMPOL-mediated repriming and ssDNA gap formation, while having no impact on the stability of stalled replication forks. Our findings establish an important function for BRCA2, provide insights into replication fork control during the DNA damage response, and may have implications in tumor suppression and therapy response.

BRCA1 prevents R-loop-associated centromeric instability

Centromeres are defined by chromatin containing the histone H3 variant CENP-A assembled onto repetitive α-satellite sequences, which are actively transcribed throughout the cell cycle. Centromeres play an essential role in chromosome inheritance and genome stability through coordinating kinetochores assembly during mitosis. Structural and functional alterations of the centromeres cause aneuploidy and chromosome aberrations which can induce cell death. In human cells, the tumor suppressor BRCA1 associates with centromeric chromatin in the absence of exogenous damage. While we previously reported that BRCA1 contributes to proper centromere homeostasis, the mechanism underlying its centromeric function and recruitment was not fully understood. Here, we show that BRCA1 association with centromeric chromatin depends on the presence of R-loops, which are non-canonical three-stranded structures harboring a DNA:RNA hybrid and are frequently formed during transcription. Subsequently, BRCA1 counteracts the accumulation of R-loops at centromeric α-satellite repeats. Strikingly, BRCA1-deficient cells show impaired localization of CENP-A, higher transcription of centromeric RNA, increased breakage at centromeres and formation of acentric micronuclei, all these features being R-loop-dependent. Finally, BRCA1 depletion reveals a Rad52-dependent hyper-recombination process between centromeric satellite repeats, associated with centromere instability and missegregation. Altogether, our findings provide molecular insights into the key function of BRCA1 in maintaining centromere stability and identity.