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Replicational bypass of DNA damage



 
  


     Mutations primarily accumulate from the misincorporation of nucleotides opposite damaged bases during DNA replication [1].  Although DNA pol d is major replicative polymerase, and synthesises all of the leading strand and most of the lagging strand of SV40-based templates [2], two additional polymerases (DNA pols zeta (z) and eta (h)) allow DNA synthesis past sites of base damage.

 
 
  

 
 

Error-free bypass

 

     In yeast, the DNA damage inducible RAD30 gene encodes for DNA pol h which is capable of efficiently bypassing CPDs [3, 4]  (see UV-induced photoproducts).  As pol h, incorporates two adenines opposite the thymine-thymine dimer, this bypass is considered to be "error-free" [4], and accounts for > 95% of bypass events in yeast [5]. 

     The human homologue of DNA pol h has been identified as the product of the XP variant (XPV) gene [6, 7] (see xeroderma pigmentosum).  Like its yeast counterpart, the human DNA pol h bypasses thymine-thymine CPDs (the predominant CPD species) in an error-free manner, but is incapable of bypassing 6-4 PPs [6].  Mutations in the XPV gene result in an impaired (reduced 10-26 fold) and error-prone bypass of damaged bases [8-10], presumably a result of the "error-prone" lesion bypass pathway catalysed by DNA pol z.
 

 
  

 
 

Error-prone bypass

 

      The yeast DNA pol z consists of the products of two genes, Rev3 and Rev7 and plays an essential role in UV-induced mutagenesis (reviewed in [9, 11, 12]).  This complex has limited processivity (adding less than four nucleotides per binding), and lacks a proofreading 3'-5' exonuclease activity but can replicate past CPDs with approximately 10% of the efficiency of that for undamaged DNA, even though CPDs inhibit DNA replication by DNA pol d up to 10,000 fold [13, 14].  Deletion of any of the Rev genes results in normal viability in the absence of DNA damage, and moderately reduced viability following treatment with DNA damaging agents; but a great reduction in the rates of both spontaneous and DNA-damage induced mutagenesis [15-19].  This suggests that this pathway, although not essential for viability, plays a significant role in mutagenesis. 

     A human homologue of Rev3 (designated Rev3L) has been identified [20, 21].  Like its yeast counterpart, Rev3L is not essential for viability, and downregulation of Rev3L, via antisense mRNA, resulted in a decrease in UV-induced mutagenesis [20], suggesting that a similar "error-prone" lesion bypass pathway exists in humans.  As in yeast, this pathway appears to be responsible for most UV-induced mutations, even though it is only a minor contributor to translesion synthesis under normal conditions. 

     The question of why a mutagenic bypass system persists in higher eukaryotes remains unresolved.  However, as the DNA pol h mediated "error-free" pathway is incapable of transcribing past 6-4 PP, it is possible that the "error-prone" DNA pol z pathway is maintained for this purpose, which would account for the increased mutagenicity of 6-4 PPs relative to CPDs [22].  As 6-4 PPs are induced at only 1/4 to 1/3 the frequency of CPDs, and repaired at approximately 5 times their rate [22-26], these lesions may be infrequent enough that a mutagenic bypass is preferable to a blockage of replication.  Unfortunately, under conditions where the DNA pol h mediated "error-free" pathway is inactivated (as in the case of XP-V individuals) or saturated (as a result of deficient NER in classical XP groups, or in normal cells following high UV exposures), the increased contribution of the "error-prone" pathway to DNA synthesis past CPDs can lead to the introduction of significant numbers of mutations, and ultimately to carcinogenesis. 

 
  

 
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References:
 

1. McGregor, W.G., et al., Cell cycle-dependent strand bias for UV-induced mutations in the transcribed strand of excision repair-proficient human fibroblasts but not in repair-deficient cells. Mol Cell Biol, 1991. 11(4): p. 1927-34.

2. Waga, S., G. Bauer, and B. Stillman, Reconstitution of complete SV40 DNA replication with purified replication factors. J Biol Chem, 1994. 269(14): p. 10923-34.

3. McDonald, J.P., A.S. Levine, and R. Woodgate, The Saccharomyces cerevisiae RAD30 gene, a homologue of Escherichia coli dinB and umuC, is DNA damage inducible and functions in a novel error-free postreplication repair mechanism. Genetics, 1997. 147(4): p. 1557-68.

4. Johnson, R.E., S. Prakash, and L. Prakash, Efficient bypass of a thymine-thymine dimer by yeast DNA polymerase, Pol eta. Science, 1999. 283(5404): p. 1001-4.

5. Baynton, K., A. Bresson-Roy, and R.P. Fuchs, Analysis of damage tolerance pathways in Saccharomyces cerevisiae: a requirement for Rev3 DNA polymerase in translesion synthesis. Mol Cell Biol, 1998. 18(2): p. 960-6.

6. Masutani, C., et al., Xeroderma pigmentosum variant (XP-V) correcting protein from HeLa cells has a thymine dimer bypass DNA polymerase activity. Embo J, 1999. 18(12): p. 3491-501.

7. Masutani, C., et al., The XPV (xeroderma pigmentosum variant) gene encodes human DNA polymerase eta. Nature, 1999. 399(6737): p. 700-4.

8. Svoboda, D.L., L.P. Briley, and J.M. Vos, Defective bypass replication of a leading strand cyclobutane thymine dimer in xeroderma pigmentosum variant cell extracts. Cancer Res, 1998. 58(11): p. 2445-8.

9. McGregor, W.G., et al., Abnormal, error-prone bypass of photoproducts by xeroderma pigmentosum variant cell extracts results in extreme strand bias for the kinds of mutations induced by UV light. Mol Cell Biol, 1999. 19(1): p. 147-54.

10. Cordonnier, A.M., A.R. Lehmann, and R.P. Fuchs, Impaired translesion synthesis in xeroderma pigmentosum variant extracts. Mol Cell Biol, 1999. 19(3): p. 2206-11.

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12. Burgers, P.M., Eukaryotic DNA polymerases in DNA replication and DNA repair. Chromosoma, 1998. 107(4): p. 218-27.

13. O'Day, C.L., P.M. Burgers, and J.S. Taylor, PCNA-induced DNA synthesis past cis-syn and trans-syn-I thymine dimers by calf thymus DNA polymerase delta in vitro. Nucleic Acids Res, 1992. 20(20): p. 5403-6.

14. Nelson, J.R., C.W. Lawrence, and D.C. Hinkle, Thymine-thymine dimer bypass by yeast DNA polymerase zeta. Science, 1996. 272(5268): p. 1646-9.

15. Quah, S.K., R.C. von Borstel, and P.J. Hastings, The origin of spontaneous mutation in Saccharomyces cerevisiae. Genetics, 1980. 96(4): p. 819-39.

16. Roche, H., R.D. Gietz, and B.A. Kunz, Specificities of the Saccharomyces cerevisiae rad6, rad18, and rad52 mutators exhibit different degrees of dependence on the REV3 gene product, a putative nonessential DNA polymerase. Genetics, 1995. 140(2): p. 443-56.

17. Roche, H., R.D. Gietz, and B.A. Kunz, Specificity of the yeast rev3 delta antimutator and REV3 dependency of the mutator resulting from a defect (rad1 delta) in nucleotide excision repair. Genetics, 1994. 137(3): p. 637-46.

18. Kalinowski, D.P., F.W. Larimer, and M.J. Plewa, Analysis of spontaneous frameshift mutations in REV1 and rev1-1 strains of Saccharomyces cerevisiae. Mutat Res, 1995. 331(1): p. 149-59.

19. Lawrence, C., The RAD6 DNA repair pathway in Saccharomyces cerevisiae: what does it do, and how does it do it? Bioessays, 1994. 16(4): p. 253-8.

20. Gibbs, P.E., et al., A human homolog of the Saccharomyces cerevisiae REV3 gene, which encodes the catalytic subunit of DNA polymerase zeta. Proc Natl Acad Sci U S A, 1998. 95(12): p. 6876-80.

21. Xiao, W., et al., Identification, chromosomal mapping and tissue-specific expression of hREV3 encoding a putative human DNA polymerase zeta. Carcinogenesis, 1998. 19(5): p. 945-9.

22. Zdzienicka, M.Z., et al., (6-4) photoproducts and not cyclobutane pyrimidine dimers are the main UV-induced mutagenic lesions in Chinese hamster cells. Mutat Res, 1992. 273(1): p. 73-83.

23. Mitchell, D.L. and R.S. Nairn, The biology of the (6-4) photoproduct. Photochem Photobiol, 1989. 49(6): p. 805-19.

24. Cleaver, J.E., et al., The relative biological importance of cyclobutane and (6-4) pyrimidine-pyrimidone dimer photoproducts in human cells: evidence from a xeroderma pigmentosum revertant. Photochem Photobiol, 1988. 48(1): p. 41-9.

25. Broughton, B.C., et al., Relationship between pyrimidine dimers, 6-4 photoproducts, repair synthesis and cell survival: studies using cells from patients with trichothiodystrophy. Mutat Res, 1990. 235(1): p. 33-40.

26. Szymkowski, D.E., C.W. Lawrence, and R.D. Wood, Repair by human cell extracts of single (6-4) and cyclobutane thymine-thymine photoproducts in DNA. Proceedings of the National Academy of Sciences of the United States of America, 1993. 90(21): p. 9823-7.
 
 



 
 
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