Crosslinking of DNA

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In genetics, crosslinking of DNA occurs when various exogenous or endogenous agents react with two different positions in the DNA. This can occur in either the same strand (intrastrand crosslink) or in the opposite strands of the DNA (interstrand crosslink). Crosslinks also occur between DNA and protein. DNA replication is blocked by crosslinks, which cause replication arrest and cell death if the crosslink is not repaired.

The RAD51 family plays a role in repair.[1]

Crosslinking agents[edit]


Alkylating agents such as 1, 3-bis(2-chloroethyl)-1-nitrosourea (BCNU, carmustine)) and nitrogen mustard which are used in chemotherapy can cross link with DNA at N7 position of guanine on the opposite strands forming interstrand crosslinks.[2]

Cisplatin (cis-diamminedichloroplatinum(II)) and its derivatives forms DNA cross links as monoadduct, interstrand crosslink, intrastrand crosslink or DNA protein crosslink. Mostly it acts on the adjacent N-7 guanine forming 1, 2 intrastrand crosslink.[3][4]

DNA damage induced by ionizing radiation is similar to that of oxidative stress, and these lesions have been implicated in aging and cancer. Biological effects of single-base damage by radiation or oxidation, such as 8-oxoguanine and thymine glycol, have been extensively studied. Recently the focus has shifted to some of the more complex lesions. Tandem DNA lesions are formed at a substantial frequency by ionizing radiation and metal-catalyzed H2O2 reactions. Under anoxic conditions, the predominant double-base lesion is a species in which the C8 of guanine is linked to the 5-methyl group of an adjacent 3'-thymine (G[8,5- Me]T).[5]


  • Nitrous acid is formed in the stomach from dietary sources of nitrites. It induces formation of interstrand DNA crosslinks at the aminogroup of exocyclic N2 of guanine at CG sequences.
  • Reactive chemicals such as malondialdehyde are formed endogenously as the product of lipid peroxidation. They create etheno adducts formed by aldehyde which undergo rearrangements to form crosslinks on opposite strands.[6]
  • Psoralens are natural compounds (furocoumarins) present in plants. These compounds get activated in the presence of UV - A. They form covalent adducts with pyrimidines. Covalent adducts are formed by linking 3, 4 (pyrone) or 4', 5’ (furan) edge of psoralen to 5, 6 double bond of thymine. Psoralens can form two types of monoadducts and one diadduct (an interstrand crosslink) reacting with thymine.[7] The crosslinking reaction by Psoralens targets TA sequences intercalating in DNA and linking one base of the DNA with the one below it. Psoralen adducts cause replication arrest and are used in the treatment of psoriasis and vitiligo.
  • Aldehydes such as acrolein and crotonaldehyde found in tobacco smoke or automotive exhaust can form DNA interstrand crosslinks in DNA. Guanine adducts of DNA can also react with protein. A Schiff base formation between protein and aldehyde causes this DNA protein interstrand link
  • Formaldehyde (HCHO) induces protein-DNA and protein-protein crosslinks, and is a common reagent of choice for molecular biology experiments.[8] These crosslinks may be reversed by incubation at 70 °C.[9]

Repair of DNA crosslinks[edit]

DNA crosslinks generally cause loss of overlapping sequence information from the two strands of DNA. Therefore, accurate repair of the damage depends on retrieving the lost information from an undamaged homologous chromosome in the same cell. Retrieval can occur by pairing with a sister chromosome produced during a preceding round of replication. In a diploid cell retrieval may also occur by pairing with a non-sister homologous chromosome, as occurs especially during meiosis.[10] Once pairing has occurred, the crosslink can be removed and correct information introduced into the damaged chromosome by the process of homologous recombinational repair.

Treatment of E. coli with psoralen-plus-UV light (PUVA) produces interstrand crosslinks in the cells’ DNA. Cole et al.[11] and Sinden and Cole[12] presented evidence that an homologous recombinational repair process requiring the products of genes uvrA, uvrB, and recA can remove these crosslinks in E. coli. This process appears to be quite efficient. Even though one or two unrepaired crosslinks are sufficient to inactivate a cell, a wild-type bacterial cell can repair and therefore recover from 53 to 71 psoralen crosslinks. Eukaryotic yeast cells are also inactivated by one remaining crosslink, but wild type yeast cells can recover from 120 to 200 crosslinks.[13] In yeast, three pathways have a role in repair or toleration of crosslinks: homologous recombinational repair, nucleotide excision repair and translesion synthesis.[13]

Recombinational repair of DNA crosslinks also likely occurs in plants where it depends on gene rad51, a recA ortholog. In the plant Arabidopsis thaliana, mutants defective in a gene rad51 paralog XRCC3 are hypersensitive to mitomycin C, a crosslinking agent.[14] In rice (Oryza sativa), mutants with a defective RAD51C gene have increased sensitivity in somatic cells to mitomycin C.[15]

In humans, the leading cause of cancer deaths worldwide is lung cancer, including non small cell lung carcinoma (NSLC) which accounts for 85% of all lung cancer cases in the United States.[16] Individuals with NSLC are often treated with therapeutic platinum compounds (e.g. cisplatin, carboplatin or oxaliplatin) (see Lung cancer chemotherapy) that cause inter-strand DNA crosslinks. Among individuals with NSLC, low expression of BRCA1 in the primary tumor correlated with improved survival after platinum-containing chemotherapy.[17][18] This correlation implies that low BRCA1 in the cancer, and the consequent low level of DNA repair, causes vulnerability of the cancer to treatment by the DNA crosslinking agents. High BRCA1 may protect cancer cells by acting in the homologous recombinational repair pathway that removes the damages in DNA introduced by the platinum drugs. Taron et al.[17] and Papadaki et al.[18] concluded that the level of BRCA1 expression is a potentially important tool for tailoring chemotherapy in lung cancer management.


  1. ^ Gruver AM, Miller KA, Rajesh C, et al. (November 2005). "The ATPase motif in RAD51D is required for resistance to DNA interstrand crosslinking agents and interaction with RAD51C". Mutagenesis. 20 (6): 433–40. doi:10.1093/mutage/gei059. PMID 16236763. 
  2. ^ Ali-Osman F, Rairkar A, Young P (January 1995). "Formation and repair of 1,3-bis-(2-chloroethyl)-1-nitrosourea and cisplatin induced total genomic DNA interstrand crosslinks in human glioma cells". Cancer Biochem. Biophys. 14 (4): 231–41. PMID 7767897. 
  3. ^ Poklar N, Pilch DS, Lippard SJ, Redding EA, Dunham SU, Breslauer KJ (July 1996). "Influence of cisplatin intrastrand crosslinking on the conformation, thermal stability, and energetics of a 20-mer DNA duplex". Proc. Natl. Acad. Sci. U.S.A. 93 (15): 7606–11. doi:10.1073/pnas.93.15.7606. PMC 38793Freely accessible. PMID 8755522. 
  4. ^ Rudd GN, Hartley JA, Souhami RL (1995). "Persistence of cisplatin-induced DNA interstrand crosslinking in peripheral blood mononuclear cells from elderly and young individuals". Cancer Chemother. Pharmacol. 35 (4): 323–6. doi:10.1007/BF00689452. PMID 7828275. 
  5. ^ LC Colis; P Raychaudhury; AK Basu (2008). "Mutational specificity of gamma-radiation-induced guanine-thymine and thymine-guanine intrastrand cross-links in mammalian cells and translesion synthesis past the guanine-thymine lesion by human DNA polymerase eta". Biochemistry. 47 (6): 8070–8079. doi:10.1021/bi800529f. PMID 18616294. 
  6. ^ Mathews & Vanholde, Biochemistry, 2nd Edition. Benjamin Cummings Publication
  7. ^ Qi Wu, Laura A Christensen, Randy J Legerski & Karen M Vasquez, Mismatch repair participates in error-free processing of DNA interstrand crosslinks in human cells,EMBO reports 6, 6, 551–557 (2005).
  8. ^ Formaldehyde Crosslinking Experiments
  9. ^ Somashe Niranjanakumaria, Erika Lasdaa, Robert Brazasa, Mariano A. Garcia-Blanco. Reversible cross-linking combined with immunoprecipitation to study RNA–protein interactions in vivo. Methods. 2002 Feb;26(2):182-90. [1]
  10. ^ Harris Bernstein, Carol Bernstein and Richard E. Michod (2011). Meiosis as an Evolutionary Adaptation for DNA Repair. Chapter 19 pages 357-382 in “DNA Repair” (Inna Kruman editor). InTech Open Publisher. DOI: 10.5772/25117 ISBN 978-953-307-697-3
  11. ^ Cole RS, Levitan D, Sinden RR (1976). "Removal of psoralen interstrand cross-links from DNA of Escherichia coli: mechanism and genetic control". J. Mol. Biol. 103 (1): 39–59. doi:10.1016/0022-2836(76)90051-6. PMID 785009. 
  12. ^ Sinden RR, Cole RS (1978). "Repair of cross-linked DNA and survival of Escherichia coli treated with psoralen and light: effects of mutations influencing genetic recombination and DNA metabolism". J. Bacteriol. 136 (2): 538–47. PMC 218577Freely accessible. PMID 361714. 
  13. ^ a b Noll DM, Mason TM, Miller PS (2006). "Formation and repair of interstrand cross-links in DNA". Chem. Rev. 106 (2): 277–301. doi:10.1021/cr040478b. PMC 2505341Freely accessible. PMID 16464006. 
  14. ^ Bleuyard JY, White CI (2004). "The Arabidopsis homologue of Xrcc3 plays an essential role in meiosis". EMBO J. 23 (2): 439–49. doi:10.1038/sj.emboj.7600055. PMC 1271761Freely accessible. PMID 14726957. 
  15. ^ Kou Y, Chang Y, Li X, Xiao J, Wang S (2012). "The rice RAD51C gene is required for the meiosis of both female and male gametocytes and the DNA repair of somatic cells". J. Exp. Bot. 63 (14): 5323–35. doi:10.1093/jxb/ers190. PMC 3431001Freely accessible. PMID 22859673. 
  16. ^ Molina JR, Yang P, Cassivi SD, Schild SE, Adjei AA (2008). "Non-small cell lung cancer: epidemiology, risk factors, treatment, and survivorship". Mayo Clin. Proc. 83 (5): 584–94. doi:10.4065/83.5.584. PMC 2718421Freely accessible. PMID 18452692. 
  17. ^ a b Taron M, Rosell R, Felip E, Mendez P, Souglakos J, Ronco MS, Queralt C, Majo J, Sanchez JM, Sanchez JJ, Maestre J (2004). "BRCA1 mRNA expression levels as an indicator of chemoresistance in lung cancer". Hum. Mol. Genet. 13 (20): 2443–9. doi:10.1093/hmg/ddh260. PMID 15317748. 
  18. ^ a b Papadaki C, Sfakianaki M, Ioannidis G, Lagoudaki E, Trypaki M, Tryfonidis K, Mavroudis D, Stathopoulos E, Georgoulias V, Souglakos J (2012). "ERCC1 and BRAC1 mRNA expression levels in the primary tumor could predict the effectiveness of the second-line cisplatin-based chemotherapy in pretreated patients with metastatic non-small cell lung cancer". J Thorac Oncol. 7 (4): 663–71. doi:10.1097/JTO.0b013e318244bdd4. PMID 22425915. 

External links[edit]

  • PDB: 1AIO​ - Interactive structure for cisplatin and DNA adduct formation
  • PDB: 204D​ - Interactive structure for psoralen and crosslinked DNA
  • Psoralen Ultraviolet A Light Treatment [2]