Alternative titles; symbols
HGNC Approved Gene Symbol: RPA2
Cytogenetic location: 1p35.3 Genomic coordinates (GRCh38): 1:27,891,524-27,914,797 (from NCBI)
Replication protein A (see RPA1; 179835) is a 3-subunit single-stranded DNA (ssDNA)-binding protein that has been isolated from human cells and found to be essential for in vitro replication of the papovavirus SV40. Erdile et al. (1990) cloned the 32-kD subunit of RPA from a HeLa cell DNA library. The open reading frame predicted a protein of 270 amino acids, including the N-terminal methionine residue. The N-terminal region was particularly rich in glycine and serine.
To investigate the functions of RPA32 in multiple DNA repair pathways, Mer et al. (2000) determined the 3-dimensional solution structure of the C-terminal region of human RPA32 (amino acids 172 to 270), both free and in complex with a 16-amino acid peptide fragment encompassing the RPA-binding region of UNG2 (607752) (amino acids 73 to 88). High-resolution nuclear magnetic resonance imaging demonstrated that the interaction interfaces of UNG2, XPA (191525), and RAD52 (600392) with RPA32 are similar. The authors proposed a competition-based protein switch mechanism to assemble requisite proteins at sites of DNA damage.
The function of the ATR (601215)-ATRIP (606605) protein kinase complex is crucial for the cellular response to replication stress and DNA damage. Zou and Elledge (2003) demonstrated that the RPA complex, which associates with single-stranded DNA (ssDNA), is required for recruitment of ATR to sites of DNA damage and for ATR-mediated CHK1 (603078) activation in human cells. In vitro, RPA stimulates the binding of ATRIP to single-stranded DNA. The binding of ATRIP to RPA-coated single-stranded DNA enables the ATR-ATRIP complex to associate with DNA and stimulates phosphorylation of the RAD17 (603139) protein that is bound to DNA. Furthermore, Ddc2, the budding yeast homolog of ATRIP, is specifically recruited to double-stranded DNA breaks in an RPA-dependent manner. A checkpoint-deficient mutant of RPA, rfa1-t11, is defective for recruiting Ddc2 to single-stranded DNA both in vivo and in vitro. Zou and Elledge (2003) concluded that RPA-coated single-stranded DNA is the critical structure at sites of DNA damage that recruits the ATR-ATRIP complex and facilitates its recognition of substrates for phosphorylation and the initiation of checkpoint signaling.
Activation-induced cytidine deaminase (AID; 605257) is a ssDNA deaminase required for somatic hypermutation and class switch recombination of immunoglobulin genes. Class switch recombination involves transcription through switch regions, which generates ssDNA within R loops. Chaudhuri et al. (2004) characterized the mechanism of AID targeting to in vitro transcribed substrates harboring somatic hypermutation motifs. They showed that the targeting activity of AID is due to RPA, a ssDNA-binding heterochimeric protein involved in replication, recombination, and repair. The 32-kD subunit of RPA interacts specifically with AID from activated B cells in a manner that seems to be dependent on posttranslational AID modification. Chaudhuri et al. (2004) concluded that RPA is implicated as a novel factor involved in immunoglobulin diversification, and proposed that B cell-specific AID-RPA complexes preferentially bind to ssDNA of small transcription bubbles at somatic hypermutation hotspots, leading to AID-mediated deamination and RPA-mediated recruitment of DNA repair proteins.
Lee et al. (2010) found that a dimeric protein phosphatase-4 (PPP4) complex containing PPP4C (602035) and PPP4R2 (613822) dephosphorylated RPA2, regulating its role in the DNA double-stranded break response. PPP4C efficiently dephosphorylated phospho-RPA2 in vitro. Silencing PPP4R2 in human cell lines, or introduction of a PPP4R2 mutant unable to interact with PPP4C, altered the kinetics and pattern of RPA2 phosphorylation. Depletion of PPP4R2 impeded homologous recombination via inefficient loading of the essential factor RAD51, causing an extended G2-M checkpoint and hypersensitivity to DNA damage. Cells expressing phosphomimetic RPA2 mutants had a comparable phenotype, suggesting that PPP4-mediated dephosphorylation of RPA2 is necessary for efficient DNA damage response.
Using PCR amplification of genomic DNA from rodent-human hybrid cell lines, Umbricht et al. (1993) mapped the REPA2 gene to chromosome 1. By PCR amplification of somatic cell hybrids of chromosome 1 and by fluorescence in situ hybridization, Umbricht et al. (1994) mapped RPA2 to 1p35.
Chaudhuri, J., Khuong, C., Alt, F. W. Replication protein A interacts with AID to promote deamination of somatic hypermutation targets. Nature 430: 992-998, 2004. [PubMed: 15273694] [Full Text: https://doi.org/10.1038/nature02821]
Erdile, L. F., Wold, M. S., Kelly, T. J. The primary structure of the 32-kDa subunit of human replication protein A. J. Biol. Chem. 265: 3177-3182, 1990. [PubMed: 2406247]
Lee, D.-H., Pan, Y., Kanner, S., Sung, P., Borowiec, J. A., Chowdhury, D. A PP4 phosphatase complex dephosphorylates RPA2 to facilitate DNA repair via homologous recombination. Nature Struct. Molec. Biol. 17: 365-372, 2010. [PubMed: 20154705] [Full Text: https://doi.org/10.1038/nsmb.1769]
Mer, G., Bochkarev, A., Gupta, R., Bochkareva, E., Frappier, L., Ingles, C. J., Edwards, A. M., Chazin, W. J. Structural basis for the recognition of DNA repair proteins UNG2, XPA, and RAD52 by replication factor RPA. Cell 103: 449-456, 2000. [PubMed: 11081631] [Full Text: https://doi.org/10.1016/s0092-8674(00)00136-7]
Umbricht, C. B., Erdile, L. F., Jabs, E. W., Kelly, T. J. Cloning, overexpression, and genomic mapping of the 14-kDa subunit of human replication protein A. J. Biol. Chem. 268: 6131-6138, 1993. [PubMed: 8454588]
Umbricht, C. B., Griffin, C. A., Hawkins, A. L., Grzeschik, K. H., O'Connell, P., Leach, R., Green, E. D., Kelly, T. J. High-resolution genomic mapping of the three human replication protein A genes (RPA1, RPA2, and RPA3). Genomics 20: 249-257, 1994. [PubMed: 8020972] [Full Text: https://doi.org/10.1006/geno.1994.1161]
Zou, L., Elledge, S. J. Sensing DNA damage through ATRIP recognition of RPA-ssDNA complexes. Science 300: 1542-1548, 2003. [PubMed: 12791985] [Full Text: https://doi.org/10.1126/science.1083430]