The Ubiquitin-Interacting Motif–Containing Protein RAP80 Interacts with BRCA1 and Functions in DNA Damage Repair Response

Abstract

In this study, we examine the potential role of receptorassociated protein 80 (RAP80), a nuclear protein containing two ubiquitin-interacting motifs (UIM), in DNA damage response and double-strand break (DSB) repair. We show that following ionizing radiation and treatment with DNA-damaging agents, RAP80 translocates to discrete nuclear foci that colocalize with those of ;-H2AX. The UIMs and the region of amino acids 204 to 304 are critical for the relocalization of RAP80 to ionizing radiation-induced foci (IRIF). These observations suggest that RAP80 becomes part of a DNA repair complex at the sites of IRIF. We also show that RAP80 forms a complex with the tumor repressor BRCA1 and that this interaction is mediated through the BRCA1 COOH-terminal repeats of BRCA1. The UIMs are not required for the interaction of RAP80 with BRCA1. Knockdown of RAP80 in HEK293 cells significantly reduced DSB-induced homologydirected recombination (HDR). Moreover, inhibition of RAP80 expression by small interfering RNA increased radiosensitivity, whereas increased radioresistance was observed in human breast cancer MCF-7 cells with overexpression of RAP80. Taken together, our data suggest that RAP80 plays an important role in DNA damage response signaling and HDR-mediated DSB repair. We further show that RAP80 can function as a substrate of the ataxia-telangiectasia mutated protein kinase in vitro, which phosphorylates RAP80 at Ser 205 and Ser 402 . We show that this phosphorylation is not required for the migration of RAP80 to IRIF. [Cancer Res 2007;67(14):6647-56]

A small ubiquitin binding domain inhibits ubiquitin-dependent protein recruitment to DNA repair foci

“…18,19 53BP1 recognizes dimethylated histone H4 lysine 20 through its Tudor domain and histone H2A K15 monoubiquitinated by RNF168 via a newly identified ubiquitination-dependent recruitment (UDR) motif. 20,21 The ability of 53BP1 to bind efficiently also relies on chromatin reorganization mediated by ubiquitin-dependent removal of specific chromatin proteins in order to reveal dimethylated K20 of histone H4.…”

Degradation of Human RAP80 is Cell Cycle Regulated by Cdc20 and Cdh1 Ubiquitin Ligases

“…Receptor-associated protein 80 (RAP80) plays an important role in signal transduction in the DNA damage response by recruiting the BRCA1-A complex to DNA damage sites in a lysine 63-mediated polyubiquitin-binding dependent manner (6)(7)(8)(9)(10)(11)(12)(13)(14). The BRCA1-A complex contains many subunits, including a coiled coil domain-containing protein (CCDC98/ABRAXAS), the BRCA1/BRCA2-containing complex subunit 45/brain and reproductive organ-expressed protein (BRCC45/BRE), the BRCA1/BRCA2-containing complex subunit 36 (BRCC36), mediator of RAP80 interactions and targeting subunit of 40/new component of the BRCA1-A complex (MERIT40/NBA1), and breast cancer 1 (BRCA1; refs.…”

“…The BRCA1-A complex contains many subunits, including a coiled coil domain-containing protein (CCDC98/ABRAXAS), the BRCA1/BRCA2-containing complex subunit 45/brain and reproductive organ-expressed protein (BRCC45/BRE), the BRCA1/BRCA2-containing complex subunit 36 (BRCC36), mediator of RAP80 interactions and targeting subunit of 40/new component of the BRCA1-A complex (MERIT40/NBA1), and breast cancer 1 (BRCA1; refs. [6][7][8][9][10][11][12][13][14]. This recruitment is required for the activity of BRCA1 at the DNA damage checkpoint and for DNA repair.…”

Ubc13/Rnf8 ubiquitin ligases control foci formation of the Rap80/Abraxas/Brca1/Brcc36 complex in response to DNA damage

“…The UIM domains of Rap80 are required for the localization of Rap80 and Abraxas to IR-induced foci (IRIFs) (10,11,14,15). The fact that the UIMs of Rap80 bind to K63-linked polyubiquitin chains implies the existence of an upstream E3 ubiquitin ligase that should play a critical role in the DNA-damage response.…”