BRCA1-directed, enhanced and aberrant homologous recombination: mechanism and potential treatment strategies

Abstract

Despite intense studies, questions still remain regarding the molecular mechanisms leading to the development of hereditary breast and ovarian cancers. Research focused on elucidating the role of the breast cancer susceptibility gene 1 (BRCA1) in the DNA damage response may be of the most critical importance to understanding these processes. The BRCA1 protein has an N-terminal RING domain possessing E3 ubiquitinligase activity and a C-terminal BRCT domain involved in binding specific phosphoproteins. These domains are involved directly or indirectly in DNA double-strand break (DSB) repair. As the two terminal domains of BRCA1 represent two separate entities, understanding how these domains communicate and are functionally altered in regards to DSB repair is critical for understanding the development of BRCA1-related breast and ovarian cancers and for developing novel therapeutics. Herein, we review recent findings of how altered functions of these domains might lead to cancer through a mechanism of increased aberrant homologous recombination and possible implications for the development of BRCA1 inhibitors.

Figure 1

Effects of BRCA1 RING and BRCT domain mutations on homologous recombination. (A) The BRCA1 protein consists of an N-terminal RING domain possessing E3 ubiquitin-ligase activity and two tandem C-terminal BRCT repeats that bind phosphoproteins such as Abraxas-RAP80 (BRCA1-A complex). (B) Mutations in the RING domain that disrupt the ubiquitin-ligase activity of BRCA1, but not BARD1 binding, do not affect HR., (C) Certain mutations in the BRCT binding pocket that disrupt interactions with phosphoproteins, or silencing of the BRCA1 A complex, results in aberrant, hyper-HR (HHR).– (D) Mutations in both the RING and BRCT domains diminish HHR, suggesting a dependence of ubiquitinylation specifically in HHR but not in HR. ↑, increase; ↓, decrease; ↔, no change; ?, unknown; *, no effect, or decreased NHEJ.

Figure 2

Possible involvement of BRCA1 and PML nuclear bodies in the temporal processing of DSBs during homologous recombination. Top portion (above the arrow) depicts the spatio-temporal process of normal HR whereas the bottom (below the arrow) depicts HHR. BRCA1 is likely playing a critical structural role in the temporal ‘handing-over’ process during HR in which the N- and C-terminal domains, along with the internal domain, communicate and coordinate the various steps and ensure a timely execution and conclusion of the repair process. The anchoring of BRCA1 to repair centers via phosphorylation, ubiquitinylation and sumoylation, and subsequent processing of these post-translational modifications, drives the DSB repair process from the time ionizing radiation (IR) damage occurs until it is repaired. PML-NBs are believed to be ‘factories’ for the (dis)assembly of DNA repair proteins such as BRCA1 and RAD51. Silencing of the BRCA1 A complex, or the expression of specific BRCA1 BRCT mutants (e.g., K1702M or M1775R), results in extensive DNA resection at the DSB and ‘stalled’ repair late in the recombination process (with accumulated levels of RPA and RAD51– associated with ssDNA) that seems to occur in close proximity to the PML-NBs. Excessive resection might occur because of the inability of BRCA1 BRCT mutants to bind CtIP and recruit MRN in the initial stages of resection thereby allowing other nuclease complexes such as Exo1-BLM-Dna2 uncontrolled access to DNA ends. Ub, ubiquitin; Dub, deubiquitinase; dynamic interactions between the RING, BRCT, SQ cluster and coiled-coil (CC) domains.

Figure 3

Potential BRCA1 therapeutic targets. BRCA1 with its RING, tandem BRCT, and overlapping SQ cluster and coiled-coil domains are indicated. Although the BRCT domain and, to a lesser extent, the RING domain, have been the focus of inhibitor design, others such as the coiled-coil domain may also be viable targets. The Zn²⁺ binding sites of the RING domain can be non-specifically inhibited by platinum compounds listed here in order of their affinity for the domain. Extensive exploration of phosphopeptides that bind to the BRCT domain has resulted in the peptide shown which has a K_i of 40 nM. Structural representations are of the BRCA1 RING and BRCT domains co-crystallized with the RING domain of BARD1 and a BACH1 phosphopeptide, respectively, and were adapted from PDB entry codes 1JM7 and 1T29, using the PyMOL Molecular Graphics System (Schrödinger, LLC).