Alternative titles; symbols
HGNC Approved Gene Symbol: RAD18
Cytogenetic location: 3p25.3 Genomic coordinates (GRCh38) : 3:8,877,075-8,963,472 (from NCBI)
Postreplication repair functions in gap filling of a daughter strand on replication of damaged DNA. In the yeast Saccharomyces cerevisiae, the rad18 protein, together with the rad6 protein (UBE2A; 312180), plays a pivotal role in the process (review by Tateishi et al., 2000). Human RAD18 is an E3 ubiquitin ligase that ubiquitinates PCNA (176740), a key component of DNA replication and repair factors, and is recruited with PCNA to sites of DNA damage (Centore et al., 2012).
Tateishi et al. (2000) cloned a RAD18 cDNA by screening a human placenta cDNA library with a human EST clone that encodes a peptide with homology to the N terminus of S. cerevisiae rad18. Human RAD18 encodes a deduced 495-amino acid protein that shares 20% sequence identity and 42% similarity with yeast rad18. Human RAD18 localizes in the nucleus. In vivo the human RAD18 protein binds to the human homologs of the yeast rad6 protein through a conserved ring finger motif. Stable transformants with the human RAD18 gene mutated in this motif became sensitive to UV, methyl methanesulfonate, and mitomycin C, and were found to be defective in the replication of UV-damaged DNA. Thus, human RAD18 is a functional homolog of yeast rad18.
The RAD6 (see 179095) pathway is central to post-replicative DNA repair in eukaryotic cells. Two principal elements of this pathway are the ubiquitin-conjugating enzymes RAD6 and the MMS2 (603001)-UBC13 (603679) heterodimer, which are recruited to chromatin by the RING finger proteins RAD18 and RAD5 (608048), respectively. Hoege et al. (2002) showed that UBC9 (601661), a small ubiquitin-related modifier (SUMO)-conjugating enzyme, is also affiliated with this pathway and that PCNA (176740), a DNA polymerase sliding clamp involved in DNA synthesis and repair, is a substrate. PCNA is monoubiquitinated through RAD6 and RAD18, modified by lys63-linked multiubiquitination, which additionally requires MMS2, UBC13, and RAD5, and is conjugated to SUMO by UBC9. All 3 modifications affect the same lysine residue of PCNA, K164, suggesting that they label PCNA for alternative functions. Hoege et al. (2002) demonstrated that these modifications differentially affect resistance to DNA damage, and that damage-induced PCNA ubiquitination is elementary for DNA repair and occurs at the same conserved residue in yeast and humans.
Ulrich and Jentsch (2000) demonstrated that RAD18 and RAD5 play a central role in mediating physical contacts between the members of the RAD6 pathway. RAD5 recruits the UBC13-MMS2 complex to DNA by means of its RING finger domain. Moreover, RAD5 association with RAD18 brings UBC13-MMS2 into contact with the RAD6-RAD18 complex. Interaction between the 2 RING finger proteins thus promotes the formation of a heteromeric complex in which the 2 distinct ubiquitin-conjugating activities of RAD6 and UBC13-MMS2 can be closely coordinated. Ulrich and Jentsch (2000) found that while UBC13 and MMS2 are largely cytosolic proteins, DNA damage triggers their redistribution to the nucleus.
By gene targeting, Tateishi et al. (2003) constructed Rad18 knockout mouse embryonic stem cells. These cells grew at a normal rate but were hypersensitive to DNA-damaging agents and showed defective postreplication repair. The mutation rate of knockout cells was normal; however, spontaneous sister chromatid exchange occurred with twice the normal frequency and was 3-fold higher following mild DNA damage. Stable transformation and gene targeting to specific loci occurred with significantly higher frequency in Rad18 knockout cells than in wildtype cells. Tateishi et al. (2003) concluded that dysfunction of Rad18 increases the frequency of homologous as well as illegitimate recombinations and that Rad18 contributes to the maintenance of genomic stability through postreplication repair.
Using yeast 2-hybrid screening and coimmunoprecipitation analysis, Adams et al. (2005) showed that human BRCTX (SLF1; 618467) interacted with human RAD18. The C terminus of BRCTX and residues 285 to 495 at the C terminus of RAD18 mediated the interaction. BRCTX and RAD18 colocalized in nucleus.
Liu et al. (2012) found that mouse Brctx relocalized to sites of DNA damage from its normally diffuse nuclear localization in transfected HEK293T cells after irradiation. Mass spectrometric and coimmunoprecipitation analyses revealed that mouse Brctx interacted with RAD18 in transfected HEK293T cells. Both BRCT domains of Brctx and residues 439 to 470 of RAD18 were required for the interaction. Further analysis confirmed that the BRCT domains of Brctx bound specifically to phosphorylated RAD18, and interaction between the 2 proteins was critical for RAD18 function in the UV-induced damage repair pathway. Examination of DNA damage-induced Brctx foci formation in mouse embryonic fibroblasts revealed that Brctx acted downstream of Rnf8 (611685) and Rad18 in the DNA damage signal transduction pathway.
Hishida et al. (2009) examined the response of yeast cells to chronic low dose ultraviolet light (CLUV) and identified a key role for the RAD6-RAD18-RAD5 error-free postreplication repair (PRR) pathway in promoting cell growth and survival. They found that loss of the RAD6 error-free PRR pathway resulted in DNA damage checkpoint-induced G2 arrest in CLUV-exposed cells, whereas wildtype and nucleotide excision repair-deficient cells were largely unaffected. Cell cycle arrest in the absence of the RAD6 error-free PRR pathway was not caused by a repair defect or by the accumulation of ultraviolet-induced photoproducts. Hishida et al. (2009) observed increased replication protein A (RPA; see 179835)- and Rad52 (600392)-yellow fluorescent protein foci in the CLUV-exposed Rad18 (605256)-delta cells and demonstrated that Rad52-mediated homologous recombination is required for the viability of the Rad18-delta cells after release from CLUV-induced G2 arrest. These and other data presented suggested that, in response to environmental levels of ultraviolet exposure, the RAD6 error-free PRR pathway promotes replication of damaged templates without the generation of extensive single-stranded DNA regions. Thus this pathway is specifically important during chronic low dose ultraviolet exposure to prevent counterproductive DNA checkpoint activation and allow cells to proliferate normally.
Using genetic and physical approaches, Branzei et al. (2008) showed that in S. cerevisiae Rad18 is required for the formation of X-shaped sister chromatid junctions (SCJs) at damaged replication forks through a process involving PCNA (176740) polyubiquitylation and the ubiquitin-conjugating enzymes Mms2 (603001) and Ubc13 (603679). The Rad18-Mms2-mediated damage bypass through SCJs requires the SUMO-conjugating enzyme Ubc9 (601661) and sumoylated PCNA, and is coordinated with Rad51 (179617)-dependent recombination events. Branzei et al. (2008) proposed that the Rad18-Rad5-Mms2-dependent SCJs represent template switch events. They concluded that their results unmasked a role for PCNA ubiquitylation and sumoylation pathways in promoting transient damage-induced replication-coupled recombination events involving sister chromatids at replication forks.
Hu et al. (2013) delineated 2 pathways that spontaneously fuse inverted repeats to generate unstable chromosomal rearrangements in wildtype mouse embryonic stem cells. Gamma radiation induced a RECQL3 (604610)-regulated pathway that selectively fused identical, but not mismatched, repeats. By contrast, ultraviolet light induced a RAD18-dependent pathway that efficiently fused mismatched repeats. In addition, TREX2 (300370), a 3-prime-to-5-prime exonuclease, suppressed identical repeat fusion but enhanced mismatched repeat fusion, clearly separating these pathways. TREX2 associated with UBC13 and enhanced PCNA ubiquitination in response to ultraviolet light, consistent with its being a novel member of error-free postreplication repair. RAD18 and TREX2 also suppressed replication fork stalling in response to nucleotide depletion. Replication fork stalling induced fusion for identical and mismatched repeats, implicating faulty replication as a causal mechanism for both pathways.
By fluorescence in situ hybridization and radiation hybrid analysis, Tateishi et al. (2000) mapped the RAD18 gene to 3p25-p24, a region where deletions are often found in lung, breast, ovarian, and testicular cancers.
Adams, D. J., van der Weyden, L., Gergely, F. V., Arends, M. J., Ng, B. L., Tannahill, D., Kanaar, R., Markus, A., Morris, B. J., Bradley, A. BRCTx is a novel, highly conserved RAD18-interacting protein. Molec. Cell. Biol. 25: 779-788, 2005. [PubMed: 15632077] [Full Text: https://doi.org/10.1128/MCB.25.2.779-788.2005]
Branzei, D., Vanoli, F., Foiani, M. SUMOylation regulates Rad18-mediated template switch. Nature 456: 915-920, 2008. [PubMed: 19092928] [Full Text: https://doi.org/10.1038/nature07587]
Centore, R. C., Yazinski, S. A., Tse, A., Zou, L. Spartan/C1orf124, a reader of PCNA ubiquitylation and a regulator of UV-induced DNA damage response. Molec. Cell 46: 625-635, 2012. [PubMed: 22681887] [Full Text: https://doi.org/10.1016/j.molcel.2012.05.020]
Hishida, T., Kubota, Y., Carr, A. M., Iwasaki, H. RAD6-RAD18-RAD5-pathway-dependent tolerance to chronic low-dose ultraviolet light. Nature 457: 612-615, 2009. [PubMed: 19079240] [Full Text: https://doi.org/10.1038/nature07580]
Hoege, C., Pfander, B., Moldovan, G.-L., Pyrowolakis, G., Jentsch, S. RAD6-dependent DNA repair is linked to modification of PCNA by ubiquitin and SUMO. Nature 419: 135-141, 2002. [PubMed: 12226657] [Full Text: https://doi.org/10.1038/nature00991]
Hu, L., Kim, T. M., Son, M. Y., Kim, S.-A., Holland, C. L., Tateishi, S., Kim, D. H., Yew, P. R., Montagna, C., Dumitrache, L. C., Hasty, P. Two replication fork maintenance pathways fuse inverted repeats to rearrange chromosomes. Nature 501: 569-572, 2013. [PubMed: 24013173] [Full Text: https://doi.org/10.1038/nature12500]
Liu, T., Chen, H., Kim, H., Huen, M. S. Y., Chen, J., Huang, J. RAD18-BRCTx interaction is required for efficient repair of UV-induced DNA damage. DNA Repair 11: 131-138, 2012. [PubMed: 22036607] [Full Text: https://doi.org/10.1016/j.dnarep.2011.10.012]
Tateishi, S., Niwa, H., Miyazaki, J.-I., Fujimoto, S., Inoue, H., Yamaizumi, M. Enhanced genomic instability and defective postreplication repair in RAD18 knockout mouse embryonic stem cells. Molec. Cell. Biol. 23: 474-481, 2003. [PubMed: 12509447] [Full Text: https://doi.org/10.1128/MCB.23.2.474-481.2003]
Tateishi, S., Sakuraba, Y., Masuyama, S., Inoue, H., Yamaizumi, M. Dysfunction of human Rad18 results in defective postreplication repair and hypersensitivity to multiple mutagens. Proc. Nat. Acad. Sci. 97: 7927-7932, 2000. [PubMed: 10884424] [Full Text: https://doi.org/10.1073/pnas.97.14.7927]
Ulrich, H. D., Jentsch, S. Two RING finger proteins mediate cooperation between ubiquitin-conjugating enzymes in DNA repair. EMBO J. 19: 3388-3397, 2000. [PubMed: 10880451] [Full Text: https://doi.org/10.1093/emboj/19.13.3388]