Defective BRCA1-mediated DNA end resection drives tandem duplication formation and FANCM synthetic lethality

Background

Double-strand breaks (DSBs) are the most toxic lesions that occur in DNA. Fortunately, cells possess a range of proteins involved in the repair of such damage. BRCA1 and BRCA2 are tumour suppressor proteins with essential roles in DSB repair by homologous recombination (HR). BRCA1 plays a role in DNA-end resection, interacting with CtIP and enhancing MRN activity, proteins involved in such process [1-4]. Later on, BRCA1 also interacts with enabling strand invasion and the completion of the homologous recombination process ensuring faithful DNA repair [5].

Because of their crucial role in DNA repair, mutations in BRCA1 and/or BRCA2 often lead to cancer. The genomes of BRCA1 and BRCA2-linked cancers exhibit similar features, such as genomic instability or PARPi sensitivity. However, they present different mutational signatures. The genomes of BRCA1-linked cancers show ‘Group 1’ non-homologous tandem duplications (TDs) whereas the loss of BRCA2 is not linked with TDs [6, 7].

The team of Dr. Scully took advantage of the Escherichia coli Tus/Ter replication fork barrier (RFB) to induce replication fork stalling on a mammalian chromosome [8] in order to study differences between Brca1 and Brca2 mutants. This tool permits the study of HR and other repair pathways at the Tus/Ter RFB. They demonstrated that Group 1 TD formation and Fancm synthetic lethality are produced by DNA-end resection defects.

Key Findings

BRCA1 Coiled-Coil (CC) mutant L1363P did not show Group 1 TD phenotype

The authors generated a series of Brca1 mutants in which the Coiled Coil domain was specifically affected, demonstrating their known HR defects and PARP1 sensitivity. They found that Brca1 CC mutants suppressed Tus/Ter-induced TDs. Therefore, they asked if cancer driven by BRCA1 CC mutations would accumulate TDs. In order to test it, they used a mouse model in which Trp53 and conditional alleles of Brca1 or Brca2 can be deleted. They performed whole genome sequencing (WGS) of the spleen tissue to analyse and compare the different tumours and study their TD configuration. As they hypothesised, the results indicated that Brca1 mutants showed a significant increase of Type 1 TDs, whereas Brca2 or Brca1 L1363P mutant (equivalent to the human L1407P) did not do so and exhibited very large TDs (>100 kb) (figure 6B from preprint).

Taken together, the authors elegantly showed that Brca1 L1363P mutant able to suppress Group 1 TD formation and that the Tus/Ter system could be widely used to predict TD formation.

Brca1 CC mutants tolerate Fancm deletion

Panday et al previously studied separation-of-function Fancm alleles to discriminate its functions. They showed that DNA end resection-defective Brca1^Δ11/hyg mES cells exhibited abundant TDs when depleted of FANCM. Exon 11 mutant Brca1 cells are synthetic lethal on a Fancm null background [9]. To better understand the relation between FANCM and BRCA1 the authors generated a series of Fancm^-/- in different Brca1 mutant backgrounds, observing that the lack of exon 11 in BRCA1 was the only condition to increase the formation of TDs. They discovered that Brca1^Δ11 Fancm ^-/- mutants presented clear growth defects when compared with other Brca1 mutants. In contrast, Brca1 CC mutants seemed to grow normally when FANCM was absent. Interestingly, this finding shows that the lack of FANCM is synthetic lethal with Brca1^Δ11 because of DNA-end resection defects.

What I like about this preprint

The preprint by Nilavar and colleagues is of particular interest to me, as their study aligns closely with my previous work, which aimed at understanding BRCA1 complexes and their functions in the context of cancer. Their Tus/Ter system allowed them to analyse and compare the genomes of BRCA1 and BRCA2 associated tumours, encountering differences that could be relevant for future treatments. One of the most challenging tasks in cancer research is the identification of novel therapeutic vulnerabilities. In this manuscript, the authors have used the Tus/Ter system as an elegant method to discern BRCA1-linked Group 1 TD formation. They also explained how Group 1 TD formation is accelerated when DNA-end resection is affected.

As they previously described, FANCM is a Group 1 TD co-suppressor [8, 9]. With different assays, they showed the synthetic sick phenotype of Brca1 Δ11 Fancm null cells, which could be of great interest for future treatments.

Questions for the authors

Q1: In figure 2C you show a very similar effects in CC-mutants and exon 11 mutant in terms of PALB2 binding. In the discussion, you mention that the exon 11 could bind PALB2. Could you speculate further on why the lack of exon 11 in BRCA1 have such a dramatic effect on PALB2 binding?

Q2: Having shown FANCM and BRCA1 Δ11 synthetic lethality, do you have evidence that this can be exploited therapeutically? For instance, does FANCM inhibition selectively impair the viability of BRCA1 Δ11 tumour models in vivo? Do you think this could be the next step in your research?

References

1. Ceppi, I., et al., Mechanism of BRCA1-BARD1 function in DNA end resection and DNA protection. Nature, 2024. 634(8033): p. 492-500.
2. Cruz-Garcia, A., A. Lopez-Saavedra, and P. Huertas, BRCA1 accelerates CtIP-mediated DNA-end resection. Cell Rep, 2014. 9(2): p. 451-9.
3. Daley, J.M., et al., Biochemical mechanism of DSB end resection and its regulation. DNA Repair (Amst), 2015. 32: p. 66-74.
4. Escribano-Diaz, C., et al., A cell cycle-dependent regulatory circuit composed of 53BP1-RIF1 and BRCA1-CtIP controls DNA repair pathway choice. Mol Cell, 2013. 49(5): p. 872-83.
5. Zhao, W., et al., Promotion of BRCA2-Dependent Homologous Recombination by DSS1 via RPA Targeting and DNA Mimicry. Mol Cell, 2015. 59(2): p. 176-87.
6. Menghi, F., et al., The Tandem Duplicator Phenotype Is a Prevalent Genome-Wide Cancer Configuration Driven by Distinct Gene Mutations. Cancer Cell, 2018. 34(2): p. 197-210 e5.
7. Nik-Zainal, S., et al., Landscape of somatic mutations in 560 breast cancer whole-genome sequences. Nature, 2016. 534(7605): p. 47-54.
8. Willis, N.A., et al., BRCA1 controls homologous recombination at Tus/Ter-stalled mammalian replication forks. Nature, 2014. 510(7506): p. 556-9.
9. Panday, A., et al., FANCM regulates repair pathway choice at stalled replication forks. Mol Cell, 2021. 81(11): p. 2428-2444 e6.

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