Cellular regulation of ribonucleotide reductase in eukaryotes

Seminars in Cell and Developmental Biology

Volumn 30, Issue , 2014, Pages 97-103

  Guarino, Estrella ^a   Salguero, Israel ^a,b   Kearsey, Stephen E ^a  

^a UNIVERSITY OF OXFORD   (United Kingdom)

^b UNIVERSITY OF CAMBRIDGE   (United Kingdom)

Author keywords

Deoxyribonucleotide pools; DNA repair; DNA synthesis; Genome stability

Indexed keywords

DEOXYRIBONUCLEOSIDE TRIPHOSPHATE; RIBONUCLEOTIDE REDUCTASE; DEOXYRIBONUCLEOTIDE;

CELL ORGANELLE; CELLULAR DISTRIBUTION; DNA DAMAGE; DNA SYNTHESIS; EUKARYOTE; NONHUMAN; REGULATORY MECHANISM; REVIEW; ACTIVE TRANSPORT; ANIMAL; BIOSYNTHESIS; CELL CYCLE; DNA REPAIR; DNA REPLICATION; FEEDBACK SYSTEM; GENOMIC INSTABILITY; HUMAN; PHYSIOLOGY;

EUKARYOTA;

ACTIVE TRANSPORT, CELL NUCLEUS; ANIMALS; CELL CYCLE; DEOXYRIBONUCLEOTIDES; DNA REPAIR; DNA REPLICATION; FEEDBACK, PHYSIOLOGICAL; GENOMIC INSTABILITY; HUMANS; RIBONUCLEOTIDE REDUCTASES;

EID: 84901618448     PISSN: 10849521     EISSN: 10963634     Source Type: Journal    
DOI: 10.1016/j.semcdb.2014.03.030     Document Type: Review

References (102)

1. Bebenek K., Roberts J.D., Kunkel T.A. The effects of dNTP pool imbalances on frameshift fidelity during DNA replication. J Biol Chem 1992, 267:3589-3596.
2. Chabes A., Georgieva B., Domkin V., Zhao X., Rothstein R., Thelander L. Survival of DNA damage in yeast directly depends on increased dNTP levels allowed by relaxed feedback inhibition of ribonucleotide reductase. Cell 2003, 112:391-401.
3. Gon S., Napolitano R., Rocha W., Coulon S., Fuchs R.P. Increase in dNTP pool size during the DNA damage response plays a key role in spontaneous and induced-mutagenesis in Escherichia coli. Proc Natl Acad Sci U S A 2011, 108:19311-19316.
4. Holmberg C., Fleck O., Hansen H.A., Liu C., Slaaby R., Carr A.M., et al. Ddb1 controls genome stability and meiosis in fission yeast. Genes Dev 2005, 19:853-862.
5. Mathews C.K. DNA precursor metabolism and genomic stability. FASEB J 2006, 20:1300-1314.
6. Moss J., Tinline-Purvis H., Walker C.A., Folkes L.K., Stratford M.R., Hayles J., et al. Break-induced ATR and Ddb1-Cul4(Cdt)(2) ubiquitin ligase-dependent nucleotide synthesis promotes homologous recombination repair in fission yeast. Genes Dev 2010, 24:2705-2716.
7. Ahluwalia D., Schaaper R.M. Hypermutability and error catastrophe due to defects in ribonucleotide reductase. Proc Natl Acad Sci U S A 2013, 110:18596-18601.
8. Alvino G.M., Collingwood D., Murphy J.M., Delrow J., Brewer B.J., Raghuraman M.K. Replication in hydroxyurea: it's a matter of time. Mol Cell Biol 2007, 27:6396-6406.
9. Koc A., Wheeler L.J., Mathews C.K., Merrill G.F. Hydroxyurea arrests DNA replication by a mechanism that preserves basal dNTP pools. J Biol Chem 2004, 279:223-230.
10. Sabatinos S.A., Green M.D., Forsburg S.L. Continued DNA synthesis in replication checkpoint mutants leads to fork collapse. Mol Cell Biol 2012, 32:4986-4997.
11. Bester A.C., Roniger M., Oren Y.S., Im M.M., Sarni D., Chaoat M., et al. Nucleotide deficiency promotes genomic instability in early stages of cancer development. Cell 2011, 145:435-446.
12. Bonate P.L., Arthaud L., Cantrell W.R., Stephenson K., Secrist J.A., Weitman S. Discovery and development of clofarabine: a nucleoside analogue for treating cancer. Nat Rev Drug Discov 2006, 5:855-863.
13. Shao J., Zhou B., Chu B., Yen Y. Ribonucleotide reductase inhibitors and future drug design. Curr Cancer Drug Targets 2006, 6:409-431.
14. Hakansson P., Dahl L., Chilkova O., Domkin V., Thelander L. The Schizosaccharomyces pombe replication inhibitor Spd1 regulates ribonucleotide reductase activity and dNTPs by binding to the large Cdc22 subunit. J Biol Chem 2006, 281:1778-1783.
15. Hakansson P., Hofer A., Thelander L. Regulation of mammalian ribonucleotide reduction and dNTP pools after DNA damage and in resting cells. J Biol Chem 2006, 281:7834-7841.
16. Reichard P. Interactions between deoxyribonucleotide and DNA synthesis. Annu Rev Biochem 1988, 57:349-374.
17. Hofer A., Crona M., Logan D.T., Sjoberg B.M. DNA building blocks: keeping control of manufacture. Crit Rev Biochem Mol Biol 2012, 47:50-63.
18. Nordlund P., Reichard P. Ribonucleotide reductases. Annu Rev Biochem 2006, 75:681-706.
19. Lundin D., Gribaldo S., Torrents E., Sjoberg B.M., Poole A.M. Ribonucleotide reduction - horizontal transfer of a required function spans all three domains. BMC Evol Biol 2010, 10:383.
20. Jordan A., Reichard P. Ribonucleotide reductases. Annu Rev Biochem 1998, 67:71-98.
21. Zhang Z., Yang K., Chen C.C., Feser J., Huang M. Role of the C terminus of the ribonucleotide reductase large subunit in enzyme regeneration and its inhibition by Sml1. Proc Natl Acad Sci U S A 2007, 104:2217-2222.
22. Fairman J.W., Wijerathna S.R., Ahmad M.F., Xu H., Nakano R., Jha S., et al. Structural basis for allosteric regulation of human ribonucleotide reductase by nucleotide-induced oligomerization. Nat Struct Mol Biol 2011, 18:316-322.
23. Logan D.T. Closing the circle on ribonucleotide reductases. Nat Struct Mol Biol 2011, 18:251-253.
24. Elledge S.J., Davis R.W. Identification of the DNA damage-responsive element of RNR2 and evidence that four distinct cellular factors bind it. Mol Cell Biol 1989, 9:5373-5386.
25. Elledge S.J., Davis R.W. DNA damage induction of ribonucleotide reductase. Mol Cell Biol 1989, 9:4932-4940.
26. Elledge S.J., Davis R.W. Two genes differentially regulated in the cell cycle and by DNA-damaging agents encode alternative regulatory subunits of ribonucleotide reductase. Genes Dev 1990, 4:740-751.
27. Huang M., Zhou Z., Elledge S.J. The DNA replication and damage checkpoint pathways induce transcription by inhibition of the Crt1 repressor. Cell 1998, 94:595-605.
28. Wang P.J., Chabes A., Casagrande R., Tian X.C., Thelander L., Huffaker T.C. Rnr4p, a novel ribonucleotide reductase small-subunit protein. Mol Cell Biol 1997, 17:6114-6121.
29. Sanvisens N., de Llanos R., Puig S. Function and regulation of yeast ribonucleotide reductase: cell cycle, genotoxic stress, and iron bioavailability. Biomed J 2013, 36:51-58.
30. Tsaponina O., Barsoum E., Astrom S.U., Chabes A. Ixr1 is required for the expression of the ribonucleotide reductase Rnr1 and maintenance of dNTP pools. PLoS Genet 2011, 7:e1002061.
31. Shen C., Lancaster C.S., Shi B., Guo H., Thimmaiah P., Bjornsti M.A. TOR signaling is a determinant of cell survival in response to DNA damage. Mol Cell Biol 2007, 27:7007-7017.
32. Tomar R.S., Zheng S., Brunke-Reese D., Wolcott H.N., Reese J.C. Yeast Rap1 contributes to genomic integrity by activating DNA damage repair genes. EMBO J 2008, 27:1575-1584.
33. Mazumder A., Tummler K., Bathe M., Samson L.D. Single-cell analysis of ribonucleotide reductase transcriptional and translational response to DNA damage. Mol Cell Biol 2013, 33:635-642.
34. Maqbool Z., Kersey P.J., Fantes P.A., McInerny C.J. MCB-mediated regulation of cell cycle-specific cdc22+ transcription in fission yeast. Mol Genet Genomics 2003, 269:765-775.
35. Harris P., Kersey P.J., McInerny C.J., Fantes P.A. Cell cycle, DNA damage and heat shock regulate suc22+ expression in fission yeast. Mol Gen Genet 1996, 252:284-291.
36. Hogan C.J., Aligianni S., Durand-Dubief M., Persson J., Will W.R., Webster J., et al. Fission yeast Iec1-ino80-mediated nucleosome eviction regulates nucleotide and phosphate metabolism. Mol Cell Biol 2010, 30:657-674.
37. Gomez-Escoda B., Ivanova T., Calvo I.A., Alves-Rodrigues I., Hidalgo E., Ayte J. Yox1 links MBF-dependent transcription to completion of DNA synthesis. EMBO Rep 2011, 12:84-89.
38. Saitoh S., Chabes A., McDonald W.H., Thelander L., Yates J.R., Russell P. Cid13 is a cytoplasmic poly(A) polymerase that regulates ribonucleotide reductase mRNA. Cell 2002, 109:563-573.
39. Johansson E., Hjortsberg K., Thelander L. Two YY-1-binding proximal elements regulate the promoter strength of the TATA-less mouse ribonucleotide reductase R1 gene. J Biol Chem 1998, 273:29816-29821.
40. Chabes A.L., Bjorklund S., Thelander L. S Phase-specific transcription of the mouse ribonucleotide reductase R2 gene requires both a proximal repressive E2F-binding site and an upstream promoter activating region. J Biol Chem 2004, 279:10796-10807.
41. Zhang Y.W., Jones T.L., Martin S.E., Caplen N.J., Pommier Y. Implication of checkpoint kinase-dependent up-regulation of ribonucleotide reductase R2 in DNA damage response. J Biol Chem 2009, 284:18085-18095.
42. D'Angiolella V., Donato V., Forrester F.M., Jeong Y.T., Pellacani C., Kudo Y., et al. Cyclin F-mediated degradation of ribonucleotide reductase M2 controls genome integrity and DNA repair. Cell 2012, 149:1023-1034.
43. Chabes A.L., Pfleger C.M., Kirschner M.W., Thelander L. Mouse ribonucleotide reductase R2 protein: a new target for anaphase-promoting complex-Cdh1-mediated proteolysis. Proc Natl Acad Sci U S A 2003, 100:3925-3929.
44. Nakano K., Balint E., Ashcroft M., Vousden K.H. A ribonucleotide reductase gene is a transcriptional target of p53 and p73. Oncogene 2000, 19:4283-4289.
45. Tanaka H., Arakawa H., Yamaguchi T., Shiraishi K., Fukuda S., Matsui K., et al. A ribonucleotide reductase gene involved in a p53-dependent cell-cycle checkpoint for DNA damage. Nature 2000, 404:42-49.
46. Yamaguchi T., Matsuda K., Sagiya Y., Iwadate M., Fujino M.A., Nakamura Y., et al. p53R2-dependent pathway for DNA synthesis in a p53-regulated cell cycle checkpoint. Cancer Res 2001, 61:8256-8262.
47. Guittet O., Hakansson P., Voevodskaya N., Fridd S., Graslund A., Arakawa H., et al. Mammalian p53R2 protein forms an active ribonucleotide reductase in vitro with the R1 protein, which is expressed both in resting cells in response to DNA damage and in proliferating cells. J Biol Chem 2001, 276:40647-40651.
48. Pontarin G., Ferraro P., Bee L., Reichard P., Bianchi V. Mammalian ribonucleotide reductase subunit p53R2 is required for mitochondrial DNA replication and DNA repair in quiescent cells. Proc Natl Acad Sci U S A 2012, 109:13302-13307.
49. Chabes A., Domkin V., Thelander L. Yeast Sml1, a protein inhibitor of ribonucleotide reductase. J Biol Chem 1999, 274:36679-36683.
50. Zhao X., Georgieva B., Chabes A., Domkin V., Ippel J.H., Schleucher J., et al. Mutational and structural analyses of the ribonucleotide reductase inhibitor Sml1 define its Rnr1 interaction domain whose inactivation allows suppression of mec1 and rad53 lethality. Mol Cell Biol 2000, 20:9076-9083.
51. Zhao X., Muller E.G., Rothstein R. A suppressor of two essential checkpoint genes identifies a novel protein that negatively affects dNTP pools. Mol Cell 1998, 2:329-340.
52. Andreson B.L., Gupta A., Georgieva B.P., Rothstein R. The ribonucleotide reductase inhibitor, Sml1, is sequentially phosphorylated, ubiquitylated and degraded in response to DNA damage. Nucleic Acids Res 2010, 38:6490-6501.
53. Zhao X., Chabes A., Domkin V., Thelander L., Rothstein R. The ribonucleotide reductase inhibitor Sml1 is a new target of the Mec1/Rad53 kinase cascade during growth and in response to DNA damage. EMBO J 2001, 20:3544-3553.
54. Danielsson J., Liljedahl L., Barany-Wallje E., Sonderby P., Kristensen L.H., Martinez-Yamout M.A., et al. The intrinsically disordered RNR inhibitor Sml1 is a dynamic dimer. Biochemistry 2008, 47:13428-13437.
55. Gupta V., Peterson C.B., Dice L.T., Uchiki T., Racca J., Guo J.T., et al. Sml1p is a dimer in solution: characterization of denaturation and renaturation of recombinant Sml1p. Biochemistry 2004, 43:8568-8578.
56. Yao R., Zhang Z., An X., Bucci B., Perlstein D.L., Stubbe J., et al. Subcellular localization of yeast ribonucleotide reductase regulated by the DNA replication and damage checkpoint pathways. Proc Natl Acad Sci U S A 2003, 100:6628-6633.
57. Lee Y.D., Wang J., Stubbe J., Elledge S.J. Dif1 is a DNA-damage-regulated facilitator of nuclear import for ribonucleotide reductase. Mol Cell 2008, 32:70-80.
58. Wu X., Huang M. Dif1 controls subcellular localization of ribonucleotide reductase by mediating nuclear import of the R2 subunit. Mol Cell Biol 2008, 28:7156-7167.
59. Lee Y.D., Elledge S.J. Control of ribonucleotide reductase localization through an anchoring mechanism involving Wtm1. Genes Dev 2006, 20:334-344.
60. Zhang Z., An X., Yang K., Perlstein D.L., Hicks L., Kelleher N., et al. Nuclear localization of the Saccharomyces cerevisiae ribonucleotide reductase small subunit requires a karyopherin and a WD40 repeat protein. Proc Natl Acad Sci U S A 2006, 103:1422-1427.
61. Ainsworth W.B., Hughes B.T., Au W.C., Sakelaris S., Kerscher O., Benton M.G., et al. Cytoplasmic localization of Hug1p, a negative regulator of the MEC1 pathway, coincides with the compartmentalization of Rnr2p-Rnr4p. Biochem Biophys Res Commun 2013, 439:443-448.
62. Liu C., Powell K.A., Mundt K., Wu L., Carr A.M., Caspari T. Cop9/signalosome subunits and Pcu4 regulate ribonucleotide reductase by both checkpoint-dependent and -independent mechanisms. Genes Dev 2003, 17:1130-1140.
63. Nestoras K., Mohammed A.H., Schreurs A.S., Fleck O., Watson A.T., Poitelea M., et al. Regulation of ribonucleotide reductase by Spd1 involves multiple mechanisms. Genes Dev 2010, 24:1145-1159.
64. Liu C., Poitelea M., Watson A., Yoshida S.H., Shimoda C., Holmberg C., et al. Transactivation of Schizosaccharomyces pombe cdt2+ stimulates a Pcu4-Ddb1-CSN ubiquitin ligase. EMBO J 2005, 24:3940-3951.
65. Yoshida S.H., Al-Amodi H., Nakamura T., McInerny C.J., Shimoda C. The Schizosaccharomyces pombe cdt2(+) gene, a target of G1-S phase-specific transcription factor complex DSC1, is required for mitotic and premeiotic DNA replication. Genetics 2003, 164:881-893.
66. Havens C.G., Shobnam N., Guarino E., Centore R.C., Zou L., Kearsey S.E., et al. Direct role for proliferating cell nuclear antigen in substrate recognition by the E3 ubiquitin ligase CRL4Cdt2. J Biol Chem 2012, 287:11410-11421.
67. Salguero I., Guarino E., Shepherd M.E., Deegan T.D., Havens C.G., MacNeill S.A., et al. Ribonucleotide reductase activity is coupled to DNA synthesis via proliferating cell nuclear antigen. Curr Biol 2012, 22:720-726.
68. Bondar T., Ponomarev A., Raychaudhuri P. Ddb1 is required for the proteolysis of the Schizosaccharomyces pombe replication inhibitor Spd1 during S phase and after DNA damage. J Biol Chem 2004, 279:9937-9943.
69. Fleck O., Vejrup-Hansen R., Watson A., Carr A.M., Nielsen O., Holmberg C. Spd1 accumulation causes genome instability independently of ribonucleotide reductase activity but functions to protect the genome when deoxynucleotide pools are elevated. J Cell Sci 2013, 126:4985-4994.
70. Vejrup-Hansen R., Fleck O., Landvad K., Fahnøe U., Broendum S.S., Schreurs A.-S., et al. Spd2 assists Spd1 in modulation of RNR architecture but does not regulate deoxynucleotide pools. J Cell Sci 2014, [in press].
71. Arner E.S., Eriksson S. Mammalian deoxyribonucleoside kinases. Pharmacol Ther 1995, 67:155-186.
72. Bourdon A., Minai L., Serre V., Jais J.P., Sarzi E., Aubert S., et al. Mutation of RRM2B, encoding p53-controlled ribonucleotide reductase (p53R2), causes severe mitochondrial DNA depletion. Nat Genet 2007, 39:776-780.
73. Copeland W.C. Defects in mitochondrial DNA replication and human disease. Crit Rev Biochem Mol Biol 2012, 47:64-74.
74. Mathews C.K., Song S. Maintaining precursor pools for mitochondrial DNA replication. FASEB J 2007, 21:2294-2303.
75. Chimploy K., Song S., Wheeler L.J., Mathews C.K. Ribonucleotide reductase association with mammalian liver mitochondria. J Biol Chem 2013, 288:13145-13155.
76. Wang C., Liu Z. Arabidopsis ribonucleotide reductases are critical for cell cycle progression, DNA damage repair, and plant development. Plant Cell 2006, 18:350-365.
77. Garton S., Knight H., Warren G.J., Knight M.R., Thorlby G.J. Crinkled leaves 8 - a mutation in the large subunit of ribonucleotide reductase - leads to defects in leaf development and chloroplast division in Arabidopsis thaliana. Plant J 2007, 50:118-127.
78. Yoo S.C., Cho S.H., Sugimoto H., Li J., Kusumi K., Koh H.J., et al. Rice virescent3 and stripe1 encoding the large and small subunits of ribonucleotide reductase are required for chloroplast biogenesis during early leaf development. Plant Physiol 2009, 150:388-401.
79. Pontarin G., Fijolek A., Pizzo P., Ferraro P., Rampazzo C., Pozzan T., et al. Ribonucleotide reduction is a cytosolic process in mammalian cells independently of DNA damage. Proc Natl Acad Sci U S A 2008, 105:17801-17806.
80. Niida H., Katsuno Y., Sengoku M., Shimada M., Yukawa M., Ikura M., et al. Essential role of Tip60-dependent recruitment of ribonucleotide reductase at DNA damage sites in DNA repair during G1 phase. Genes Dev 2010, 24:333-338.
81. Odsbu I., Morigen Skarstad K. A reduction in ribonucleotide reductase activity slows down the chromosome replication fork but does not change its localization. PloS ONE 2009, 4:e7617.
82. Guarino E., Salguero I., Jimenez-Sanchez A., Guzman E.C. Double-strand break generation under deoxyribonucleotide starvation in Escherichia coli. J Bacteriol 2007, 189:5782-5786.
83. Guarino E., Jimenez-Sanchez A., Guzman E.C. Defective ribonucleoside diphosphate reductase impairs replication fork progression in Escherichia coli. J Bacteriol 2007, 189:3496-3501.
84. Sanchez-Romero M.A., Molina F., Jimenez-Sanchez A. Correlation between ribonucleoside-diphosphate reductase and three replication proteins in Escherichia coli. BMC Mol Biol 2010, 11:11.
85. Poli J., Tsaponina O., Crabbe L., Keszthelyi A., Pantesco V., Chabes A., et al. dNTP pools determine fork progression and origin usage under replication stress. EMBO J 2012, 31:883-894.
86. Kuzminov A. Inhibition of DNA synthesis facilitates expansion of low-complexity repeats: is strand slippage stimulated by transient local depletion of specific dNTPs?. BioEssays 2013, 35:306-313.
87. Sanvisens N., Bano M.C., Huang M., Puig S. Regulation of ribonucleotide reductase in response to iron deficiency. Mol Cell 2011, 44:759-769.
88. Puig S., Askeland E., Thiele D.J. Coordinated remodeling of cellular metabolism during iron deficiency through targeted mRNA degradation. Cell 2005, 120:99-110.
89. Smith M.L., Micali O.C., Hubbard S.P., Mir-Rashed N., Jacobson D.J., Glass N.L. Vegetative incompatibility in the het-6 region of Neurospora crassa is mediated by two linked genes. Genetics 2000, 155:1095-1104.
90. Smith R.P., Wellman K., Haidari L., Masuda H., Smith M.L. Nonself recognition through intermolecular disulfide bond formation of ribonucleotide reductase in neurospora. Genetics 2013, 193:1175-1183.
91. Smith R.P., Wellman K., Smith M.L. Trans-species activity of a nonself recognition domain. BMC Microbiol 2013, 13:63.
92. Zimanyi C.M., Ando N., Brignole E.J., Asturias F.J., Stubbe J., Drennan C.L. Tangled up in knots: structures of inactivated forms of E. coli class Ia ribonucleotide reductase. Structure 2012, 20:1374-1383.
93. McIntosh E.M. MCB elements and the regulation of DNA replication genes in yeast. Curr Genet 1993, 24:185-192.
94. Franzolin E., Pontarin G., Rampazzo C., Miazzi C., Ferraro P., Palumbo E., et al. The deoxynucleotide triphosphohydrolase SAMHD1 is a major regulator of DNA precursor pools in mammalian cells. Proc Natl Acad Sci U S A 2013, 110:14272-14277.
95. Laguette N., Sobhian B., Casartelli N., Ringeard M., Chable-Bessia C., Segeral E., et al. SAMHD1 is the dendritic- and myeloid-cell-specific HIV-1 restriction factor counteracted by Vpx. Nature 2011, 474:654-657.
96. Hrecka K., Hao C., Gierszewska M., Swanson S.K., Kesik-Brodacka M., Srivastava S., et al. Vpx relieves inhibition of HIV-1 infection of macrophages mediated by the SAMHD1 protein. Nature 2011, 474:658-661.
97. Goldstone D.C., Ennis-Adeniran V., Hedden J.J., Groom H.C., Rice G.I., Christodoulou E., et al. HIV-1 restriction factor SAMHD1 is a deoxynucleoside triphosphate triphosphohydrolase. Nature 2011, 480:379-382.
98. Rice G.I., Bond J., Asipu A., Brunette R.L., Manfield I.W., Carr I.M., et al. Mutations involved in Aicardi-Goutieres syndrome implicate SAMHD1 as regulator of the innate immune response. Nat Genet 2009, 41:829-832.
99. Chabes A., Stillman B. Constitutively high dNTP concentration inhibits cell cycle progression and the DNA damage checkpoint in yeast Saccharomyces cerevisiae. Proc Natl Acad Sci U S A 2007, 104:1183-1188.
100. Halazonetis T.D., Gorgoulis V.G., Bartek J. An oncogene-induced DNA damage model for cancer development. Science 2008, 319:1352-1355.
101. Nick McElhinny S.A., Watts B.E., Kumar D., Watt D.L., Lundstrom E.B., Burgers P.M., et al. Abundant ribonucleotide incorporation into DNA by yeast replicative polymerases. Proc Natl Acad Sci U S A 2010, 107:4949-4954.
102. Nick McElhinny S.A., Kumar D., Clark A.B., Watt D.L., Watts B.E., Lundstrom E.B., et al. Genome instability due to ribonucleotide incorporation into DNA. Nat Chem Biol 2010, 6:774-781.