XRCC1 - Strategies for coordinating and assembling a versatile DNA damage response

Abstract

X-ray cross complementing protein 1 (XRCC1) is a DNA repair scaffold that supports base excision repair and single strand break repair, and is also a participant in other repair pathways. It also serves as an important co-transporter for several other repair proteins, including aprataxin and PNKP-like factor (APLF), and DNA Ligase 3α (LIG3). By combining highly specialized regions that help to organize specific repair functions with recruitment of additional enzymes whose contribution is dependent on the details of the damaged site, XRCC1 is able to handle an expanded range of problems that may arise as the repair progresses or in connection with other repair pathways with which it interfaces. This review discusses the interplay between these functions and considers some possible interactions that underlie its reported repair activities.

Keywords: Base excision repair; DNA repair; Single strand break repair; X-ray cross complementing protein 1; XRCC1.

Figure 1.. Schematic of XRCC1-supported DNA repair.

XRCC1 brings together enzymes focused on three overlapping but distinct DNA repair processes. The N-terminal, pol β-binding (PBB) domain recruits the primary BER polymerase, which can more generally support short patch repair. The central region responds to the PAR damage signal and supports long and short patch single strand break repair activities. The C-terminal DNA LIG3-binding BRCT2 domain brings together a ligase that is capable of interacting directly with DNA through its ZnF domain with a phosphorylation motif that can interact with the enzymes PNKP and APTX that sanitize the DNA termini at a break in order to create polymerase- or ligation-ready substrates. Linkers 1 and 2 contain important functionality that supports nuclear localization, non-specific dsDNA binding, and phosphorylation-dependent interactions with other repair proteins.

Figure 2.. XRCC1 plays an important role in repair protein nuclear localization.

A) XRCC1 contains a bipartite nuclear localization signal (NLS) in linker 1 that includes both major site binding (M) and minor site binding (m) motifs [21]. In addition to supporting nuclear localization of XRCC1, the NLS supports nuclear localization of at least two other XRCC1-binding repair proteins: LIG3 and APLF. B) The major and minor motifs of XRCC1 bind cooperatively to the cognate sites on Importinα (Impα). Impα also contains an unstructured N-terminal Impβ-binding motif (IBB) that remains bound to Impβ during the loading phase of the nuclear transport cycle. C) The bipartite NLS motif also facilitates intranuclear unloading of the XRCC1 cargo protein from Impα. A small fraction of the NLS exists with only the major site occupied. This allows binding of the nuclear unloading factor NUP50, which interacts with the region of Impα containing the minor site. D) Dissociation of the remaining major site motif allows rapid replacement by the N-terminal IBB domain, which has been freed due to intranuclear separation from Impβ. The unloading process helps to prevent export of incompletely unloaded cargo proteins.

Figure 3.. XRCC1 participates in BER by facilitating transfer of DNA damage.

DNA glycosylases and APE1 (green) scan DNA to detect damaged bases and abasic sites. Subsequent to encountering damage, the repair is initiated, and a metastable complex is formed. Encounter with XRCC1-pol β or free pol β facilitates damage release and transfer of the damaged DNA to pol β or to another more appropriate enzyme in the repair pathway (lower left). Occasional unfacilitated dissociation of the protective APE1 or DNA glycosylase from the damage intermediate complex results in an unprotected strand break, followed by PARP1 activation, and recruitment of XRCC1 via the BRCT1-PAR interaction (lower right). Highly basic regions of XRCC1 involved in DNA binding are indicated in blue. Abbreviations: APE1, apurinic/apyrimidinic endonuclease 1; PBB, XRCC1 N-terminal pol β-binding domain.

Figure 4.. Additional BER-related motifs.

A) The surface formed by the XRCC1 pol β-binding (PBB) domain (gray) in complex with pol β (green) (PDB: 3K75 [5]) possesses expanded binding capabilities, and has been shown to interact with acylprotein hydrolase (APEH). Post-translational modifications including ubiquitination of Lys168 and SUMOylation of Lys176 may also contribute to the binding interface. B) A truncated XRCC1 histone binding motif positioned immediately following the N-terminal pol β-binding (PBB) domain may support BER repair of histone-associated DNA. C) Modeled interaction of the XRCC1 binding motif (green) with H2A.Z (gray) obtained by mutating the corresponding YL1 residues in a reported YL1-H2A.Z complex (PDB: 5CHL, [29]). D) The structure shown in panel C in which the ribbon representation of H2A.Z is replaced by a surface rendering.

Figure 5.. XRCC1-APLF – an interaction between two DNA scaffolds.

The phosphopeptide motif located adjacent to the XRCC1 BRCT2 domain supports interaction with the APLF NHEJ scaffold, but the functional significance of this interaction remains unclear. From the perspective of SSBR, APLF adds additional recruitment capabilities provided by the PAR-binding Zn finger (PBZ) and histone chaperone domains (HCD) that may support additional damage recruitment capabilities beyond those provided by the XRCC1 BRCT1-PAR interaction. Such interactions could support recruitment of XRCC1/LIG3 to specific although as yet undetermined types of SSB. From the perspective of NHEJ, problems encountered at the final ligation step might be resolved by replacement of XRCC4-LIG4 with XRCC1-LIG3, which supports more challenging ligations that might arise. This interaction also may support the lower fidelity alt-NHEJ pathway if APLF is involved in alt-NHEJ, however such involvement has not been demonstrated and information about this pathway is currently limited. Other abbreviations: FHA, forkhead-associated domain; PBB – the XRCC1 N-terminal pol β-binding domain.