A Rad51-independent pathway promotes single-strand template repair in gene editing

English version

Autoři: Danielle N. Gallagher ^aff001; Nhung Pham ^aff002; Annie M. Tsai ^aff001; Abigail N. Janto ^aff001; Jihyun Choi ^aff001; Grzegorz Ira ^aff002; James E. Haber ^aff001; Nicolas V. Janto ^aff001
Působiště autorů: Department of Biology and Rosenstiel Basic Medical Sciences Research Center, Brandeis University, Waltham, MA, United States of America ^aff001; Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX, United States of America ^aff002
Vyšlo v časopise: A Rad51-independent pathway promotes single-strand template repair in gene editing. PLoS Genet 16(10): e32767. doi:10.1371/journal.pgen.1008689
Kategorie: Research Article
doi: https://doi.org/10.1371/journal.pgen.1008689

Souhrn

The Rad51/RecA family of recombinases perform a critical function in typical repair of double-strand breaks (DSBs): strand invasion of a resected DSB end into a homologous double-stranded DNA (dsDNA) template sequence to initiate repair. However, repair of a DSB using single stranded DNA (ssDNA) as a template, a common method of CRISPR/Cas9-mediated gene editing, is Rad51-independent. We have analyzed the genetic requirements for these Rad51-independent events in Saccharomyces cerevisiae by creating a DSB with the site-specific HO endonuclease and repairing the DSB with 80-nt single-stranded oligonucleotides (ssODNs), and confirmed these results by Cas9-mediated DSBs in combination with a bacterial retron system that produces ssDNA templates in vivo. We show that single strand template repair (SSTR), is dependent on Rad52, Rad59, Srs2 and the Mre11-Rad50-Xrs2 (MRX) complex, but unlike other Rad51-independent recombination events, independent of Rdh54. We show that Rad59 acts to alleviate the inhibition of Rad51 on Rad52’s strand annealing activity both in SSTR and in single strand annealing (SSA). Gene editing is Rad51-dependent when double-stranded oligonucleotides of the same size and sequence are introduced as templates. The assimilation of mismatches during gene editing is dependent on the activity of Msh2, which acts very differently on the 3’ side of the ssODN which can anneal directly to the resected DSB end compared to the 5’ end. In addition DNA polymerase Polδ’s 3’ to 5’ proofreading activity frequently excises a mismatch very close to the 3’ end of the template. We further report that SSTR is accompanied by as much as a 600-fold increase in mutations in regions adjacent to the sequences directly undergoing repair. These DNA polymerase ζ-dependent mutations may compromise the accuracy of gene editing.

Klíčová slova:

DNA recombination – DNA repair – Mismatch repair – Non-homologous end joining – Polymerase chain reaction – Single strand annealing – DNA annealing – DNA polymerase

Zdroje

1. Moore JK, Haber JE. Cell cycle and genetic requirements of two pathways of nonhomologous end-joining repair of double-strand breaks in Saccharomyces cerevisiae. Mol Cell Biol. 1996; 16(5):2164–2173. doi: 10.1128/mcb.16.5.2164 8628283

2. Boulton SJ, Jackson SP. Saccharomyces cerevisiae Ku70 potentiates illegitimate DNA double-strand break repair and serves as a barrier to error-prone DNA repair pathways. EMBO J. 1996; 15(18): 5093–5103. 8890183

3. Lemos BR, Kaplan AC, Bae JE, Ferrazzoli AE, Kuo J, et al. CRISPR/Cas9 cleavages in budding yeast reveal templated insertions and strand-specific insertion/deletion profiles. Proc Natl Acad Sci USA. 2018; 115(9) E2040–E2047 doi: 10.1073/pnas.1716855115 29440496

4. Ma JL, Kim EM, Haber JE. Yeast Mre11 and Rad1 proteins define a Ku-independent mechanism to repair double-strand breaks lacking overlapping end sequences. Mol Cell Biol. 2003; 23(23): 8820–8828. doi: 10.1128/mcb.23.23.8820-8828.2003 14612421

5. McVey M, Lee SE. MMEJ repair of double strand breaks (director’s cut): deleted sequences and alternative endings. Trends Genet. 2008; 24(11): 529–538. doi: 10.1016/j.tig.2008.08.007 18809224

6. Waters CA, Strande NT, Pryor JM, Strom CN, Mieczkowski P, Burkhalter MD, et al. The fidelity of the ligation step determines how ends are resolved during non-homologous end joining. Nat Commun. 2014; 5: 4286. doi: 10.1038/ncomms5286 24989324

7. Haber JE. A life investigating pathways that repair broken chromosomes. Annu Rev Genet. 2016; 50: 1–28. doi: 10.1146/annurev-genet-120215-035043 27732795

8. Clever B, Interthal H, Schmuckli-Maurer J, King J, Sigrist M, Heyer WD. Recombinational repair in yeast: functional interactions between Rad51 and Rad54 proteins. EMBO J. 1997; 16(9): 2535–2544. doi: 10.1093/emboj/16.9.2535 9171366

9. Hays SL, Firmenich AA, Berg P. Complex formation in yeast double-strand break repair: participation of Rad51, Rad52, Rad55, and Rad57 proteins. Proc Natl Acad Sci USA. 1995; 92(15): 6925–6929. doi: 10.1073/pnas.92.15.6925 7624345

10. Jiang H, Xie Y, Houston P, Stemke-Hale K, Mortensen UH, Rothstein R, et al. Direct association between the yeast Rad51 and Rad54 recombination proteins. J Biol Chem. 1996; 271(52): 33181–33186. doi: 10.1074/jbc.271.52.33181 8969173

11. Mazin AV, Bornarth CJ, Solinger JA, Heyer WD, Kowalczykowski SC. Rad54 protein is targeted to pairing loci by the Rad51 nucleoprotein filament. Mol Cell. 2000; 6(3): 583–592. doi: 10.1016/s1097-2765(00)00057-5 11030338

12. Petukhova G, Stratton S, Sung P. Catalysis of homologous DNA pairing by yeast Rad51 and Rad54 proteins. Nature. 1998; 393(6680): 91–94. doi: 10.1038/30037 9590697

13. Sung P. Yeast Rad55 and Rad57 proteins form a heterodimer that functions with replication protein A to promote DNA strand exchange by Rad51 recombinase. Genes Dev. 1997; 11(9): 1111–1121. doi: 10.1101/gad.11.9.1111 9159392

14. Lao JP, Oh SD, Shinohara M, Shinohara A, Hunter N. Rad52 promotes postinvasion steps of meiotic double-strand-break repair. Mol Cell. 2008; 29(4): 517–524. doi: 10.1016/j.molcel.2007.12.014 18313389

15. Ozenberger BA, Roeder GS. A unique pathway of double-strand break repair operates in tandemly repeated genes. Mol Cell Biol. 1991; 11(3): 1222–1231. doi: 10.1128/mcb.11.3.1222 1996088

16. Fishman-Lobell J, Haber JE. Removal of nonhomologous DNA ends in double-strand break recombination: the role of the yeast ultraviolet repair gene RAD1. Science. 1992; 258(5081): 480–484. doi: 10.1126/science.1411547 1411547

17. McDonald JP, Rothstein R. Unrepaired heteroduplex DNA in Saccharomyces cerevisiae is decreased in Rad1 Rad52-independent recombination. Genetics. 1994; 137(2): 393–405. 8070653

18. Ivanov EL, Sugawara N, Fishman-Lobell J, Haber JE. Genetic requirements for the single-strand annealing pathway of double-strand break repair in Saccharomyces cerevisiae. Genetics. 1996; 142(3): 693–704. 8849880

19. Sugawara N, Ira G, Haber JE. DNA length dependence of the single-strand annealing pathway and the role of Saccharomyces cerevisiae RAD59 in double-strand break repair. Mol Cell Biol. 2000; 20(14): 5300–5309. doi: 10.1128/mcb.20.14.5300-5309.2000 10866686

20. Malkova A, Ross L, Dawson D, Hoekstra MF, Haber JE. Meiotic recombination initiated by a double-strand break in rad50 delta yeast cells otherwise unable to initiate meiotic recombination. Genetics. 1996; 143(2): 741–754. 8725223

21. Signon L, Malkova A, Naylor ML, Klein H, Haber JE. Genetic requirements for RAD51- and RAD54-independent break-induced replication repair of a chromosomal double-strand break. Mol Cell Biol. 2001; 21(6): 2048–2056. doi: 10.1128/MCB.21.6.2048-2056.2001 11238940

22. Le S, Moore JK, Haber JE, Greider CW. Rad50 and Rad51 define two pathways that collaborate to maintain telomeres in the absence of telomerase. Genetics. 1999; 152(1): 143–152. 10224249

23. Teng SC, Chang J, McCowan B, Zakian VA. Telomerase-independent lengthening of yeast telomeres occurs by an abrupt Rad50p-dependent Rif-inhibited recombinational process. Mol Cell. 2000; 6(4): 947–952. doi: 10.1016/s1097-2765(05)00094-8 11090632

24. Chen Q, Ijpma A, Greider CW. Two survivor pathways that allow growth in the absence of telomerase are generated by distinct telomere recombination events. Mol Cell Biol. 2001; 21(5): 1819–1827. doi: 10.1128/MCB.21.5.1819-1827.2001 11238918

25. Lydeard JR, Jain S, Yamaguchi M, Haber JE. Break-induced replication and terlomerase-independent telomere maintenance require Pol32. Nature. 2007; 448(7155): 820–823. doi: 10.1038/nature06047 17671506

26. Ira G, Haber JE. Characterization of RAD51-independent break-induced replication that acts preferentially with short homologous sequences. Mol Cell Biol. 2002; 22(18): 6384–6392. doi: 10.1128/mcb.22.18.6384-6392.2002 12192038

27. Sasanuma H, Furihata Y, Shinohara M, Shinohara A. Remodeling of the Rad51 DNA strand-exchange protein by the Srs2 helicase. Genetics. 2013; 194(4): 859–872. doi: 10.1534/genetics.113.150615 23770697

28. Shen B, Zhang J, Wu H, Wang J, Ma K, Li Z, et al. Generation of gene-modified mice via Cas9/RNA-mediated gene targeting. Cell Res. 2013; 23(5): 720–723. doi: 10.1038/cr.2013.46 23545779

29. Mali P, Yang L, Esvelt KM, Aach J, Guell M, DiCarlo JE, et al. RNA-guided human genome engineering via Cas9. Science. 2013; 339(6121): 823–826. doi: 10.1126/science.1232033 23287722

30. DiCarlo JE, Norville JE, Mali P, Rios X, Aach J, Church GM. Genome engineering in Saccharomyces cerevisiae using CRISPR-Cas systems. Nucleic Acids Res. 2013; 41(7): 4336–4343. doi: 10.1093/nar/gkt135 23460208

31. Gallagher DN, Haber JE. Repair of a site-specific DNA cleavage: old-school lessons for Cas9-mediated gene editing. ACS Chem Biol. 2018; 13(2): 397–405. doi: 10.1021/acschembio.7b00760 29083855

32. Knott GJ, Doudna JA. CRISPR-Cas guides the future of genetic engineering. Science. 2018; 361(6405): 866–869. doi: 10.1126/science.aat5011 30166482

33. Storici F, Snipe JR, Chan GK, Gordenin DA, Resnick MA. Conservative repair of a chromosomal single-strand DNA through two steps of annealing. Mol Cell Biol. 2006; 26(20): 7654–7657. doi: 10.1128/MCB.00672-06 16908537

34. Davis L, Maizels N. Two distinct pathways support gene correction by single-stranded donors at DNA nicks. Cell Rep. 2016; 17(7): 1872–1881. doi: 10.1016/j.celrep.2016.10.049 27829157

35. Paix A, Folkmann A, Seydoux G. Precision genome editing using CRISPR-Cas9 and linear repair templates in C. elegans. Methods. 2017; 15: 86–93. doi: 10.1016/j.ymeth.2017.03.023 28392263

36. Bothmer A, Phadke T, Barrera LA, Margulies CM, Lee CS, Buquicchio F, et al. Characterization of the interplay between DNA repair and CRISPR/Cas9-induced DNA lesions at an endogenous locus. Nat Commun. 2017; 8: 13905 doi: 10.1038/ncomms13905 28067217

37. Hicks WM, Yamaguchi M, Haber JE. Real-time analysis of double-strand DNA break repair by homologous recombination. Proc Natl Acad Sci USA. 2011; 108(8): 3108–3115. doi: 10.1073/pnas.1019660108 21292986

38. Davis L, Maizels N. Homology-directed repair of DNA nicks via pathways distinct from canonical double-strand break repair. Proc Natl Acad Sci USA. 2014; 111(10): 924–932. http://doi.org/10.1073/pnas.1400236111 24556991

39. Keskin H, Shen Y, Huang F, Patel M, Yang T, Ashley K, et al. Transcript-RNA-templated DNA recombination and repair. Nature. 2014; 515(7527): 436–439. doi: 10.1038/nature13682 25186730

40. Godin S, Wier A, Kabbinavar F, Bratton-Palmer DS, Ghodke H, Van Houten B, et al. The Shu complex interacts with Rad51 through the Rad51 paralogues Rad55-Rad57 to mediate error-free recominbation. Nucleic Acids Res. 2013; 41(8): 4525–34 doi: 10.1093/nar/gkt138 23460207

41. Symington LS. Mechanism and regulation of DNA end resection in eukaryotes. Crit Rev Biochem Mol Biol. 2016; 51(3): 195–212. doi: 10.3109/10409238.2016.1172552 27098756

42. Ferrari M, Dibitetto D, De Gregorio G, Eapen VV, Rawal CC, Lazzro F, et al. Functional interplay between the 53BP1-ortholog Rad9 and the Mre11 complex regulates resection, end-tethering and repair of a double strand break. PLoS Genet. 2015; 11(1) doi: 10.1371/journal.pgen.1004928 25569305

43. Cassani C, Gobbini E, Vertemara J, Wang W, Marsella A, Sung P, et al. Structurally distinct Mre11 domains mediate MRX functions in resection, end-tethering and DNA damage resistance. Nucleic Acids Res. 2018; 46(6): 2990–3008 doi: 10.1093/nar/gky086 29420790

44. Krejci L, Van Komen S, Li Y, Villemain J, Reddy MS, Klein H, et al. DNA helicase Srs2 disrupts the Rad51 presynaptic filament. Nature. 2003; 423(6937): 305–309. doi: 10.1038/nature01577 12748644

45. Veaute X, Jeusset J, Soustelle C, Kowalczykowski SC, Le Cam E, Fabre F. The Srs2 helicase prevents recombination by disrupting Rad51 nucleoprotein filaments. Nature. 2003; 423(6937): 309–312. doi: 10.1038/nature01585 12748645

46. Mimitou EP & Symington LS. Sae2, Exo1 and Sgs1 collaborate in DNA double-strand break processing. Nature. 2008; 455(7214): 770–774. doi: 10.1038/nature07312 18806779

47. Zhu Z, Chung WH, Shim EY, Lee SE, Ira G. Sgs1 helicase and two nucleases Dna2 and Exo1 resect DNA double-strand break ends. Cell. 2008; 134(6): 981–994. doi: 10.1016/j.cell.2008.08.037 18805091

48. Chen X, Cui D, Papusha A, Zhang X, Chu CD, Tang J, et al. The Fun30 nucleosome remodeler promotes resection of DNA double-strand break ends. Nature. 2012; 484(7417): 576–580. doi: 10.1038/nature11355 22960743

49. Costelloe T, Louge R, Tomimatsu N, Mukherjee B, Martini E, Khadaroo B, et al. The yeast Fun30 and human SMARCAD1 chromatin remodelers promote DNA end resection. Nature. 2012; 489(7417): 581–584. doi: 10.1038/nature11353 22960744

50. Eapen VV, Sugawara N, Tsabar M, Wu WH, Haber JE. The Saccharomyces cerevisiae chromatin remodeler Fun30 regulates DNA end resection and checkpoint deactivation. Mol Cell Biol. 2012; 32(22): 4727–4740. doi: 10.1128/MCB.00566-12 23007155

51. Richardson CD, Kazane KR, Feng SJ, Zelin E, Bray NL, Schafer AJ, et al. CRISPR-Cas9 genome editing in human cells occurs via the Fanconi anemia pathway. Nat Genet. 2018; 50(8): 1132–1139. doi: 10.1038/s41588-018-0174-0 30054595 http://doi.org/10.1038/s41588-018-0174-0

52. Piazza A, Shah SS, Wright WD, Gore SK, Koszul R, Heyer WD. Dynamic Processing of Displacement Loops during Recombinational DNA Repair. Mol Cell. 2019;73(6):1255–1266.e4. doi: 10.1016/j.molcel.2019.01.005 30737186

53. Tsaponina O, Haber JE. Frequent Interchromosomal Template Switches during Gene Conversion in S. cerevisiae. Mol Cell. 2014;55(4):615–625. doi: 10.1016/j.molcel.2014.06.025 25066232

54. Anand R, Beach A, Li K, Haber JE. Rad51-mediated double-strand break repair and mismatch correction of divergent substrates. Nature. 2017; 544(7650): 377–380. doi: 10.1038/nature22046 28405019

55. Wu Y, Kantake N, Sugiyama T, Kowalczykowski SC. Rad51 protein controls Rad52-mediated DNA annealing. J Biol Chem. 2008; 283(21): 14883–14892. doi: 10.1074/jbc.M801097200 18337252

56. Shi I, Hallwyl SC, Seong C, Mortensen U, Rothstein R, Sung P. Role of the Rad52 amino-terminal DNA binding activity in DNA strand capture in homologous recombination. J Biol Chem. 2009; 284(48): 33275–33284. doi: 10.1074/jbc.M109.057752 19812039

57. Vaze MB, Pellicioli A, Lee SE, Ira G, Liberi G, Arbel-Eden A, et al. Recovery from checkpoint-mediated arrest after repair of a double-strand break requires Srs2 helicase. Mol Cell. 2002; 10(2); 373–385. doi: 10.1016/s1097-2765(02)00593-2 12191482

58. Rattray AJ, Shafer BK, McGill CB, Strathern JN. The roles of REV3 and RAD57 in double-strand-break-repair-induced mutagenesis of Saccharomyces cerevisiae. Genetics. 2002; 162(3): 1063–1077. 12454056

59. Sinha S, Li F, Villarreal D, Shim JH, Yoon S, Myung K, et al. Microhomology-mediated end joining induces hypermutagenesis at breakpoint junctions. PLOS Genet. 2017; 13(4): e1006714. doi: 10.1371/journal.pgen.1006714 28419093

60. Lee K, Ji JH, Yoon K, Che J, Seol JH, Lee SE, et al. Microhomology selection for microhomology mediated end joining in Saccharomyces cerevisiae. Genes. 2019; 10(4): E284. doi: 10.3390/genes10040284 30965655

61. Holbeck SL, Strathern JN. A role for Rev3 in mutagenesis during double-strand break repair in Saccharomyces cerevisiae. Genetics. 1997; 147(3): 1017–1024. 9383049

62. Hicks WM, Kim M, Haber JE. Increased mutagenesis and unique mutation signature associated with mitotic gene conversion. Science. 2010; 329(5987): 82–85. doi: 10.1126/science.1191125 20595613

63. Boeke JD, Trueheart J, Natsoulis G, Fink GR. 5-fluoroorotic acid as a selective agent in yeast molecular genetics. Methods Enzymol. 1987; 154: 164–175. doi: 10.1016/0076-6879(87)54076-9 3323810

64. Gillet-Markowska A, Louvel G and Fischer G. bz-rates: a web-tool to estimate mutation rates from fluctuation analysis. G3, 2015; 5(11): 2323–2327. doi: 10.1534/g3.115.019836 26338660

65. Bardwell AJ, Bardwell L, Tomkinson AE, Friedberg EC. Specific cleavage of model recombination and repair intermediates by the yeast Rad1-Rad10 DNA endonuclease. Science. 1994; 265(5181): 2082–2085. doi: 10.1126/science.8091230 8091230

66. Sharon E, Chen SA, Khosla NM, Smith JD, Pritchard JK, Fraser HB. Functional genetic variants revealed by massively parallel precise genome editing. Cell. 2018; 175(2): 544–557. doi: 10.1016/j.cell.2018.08.057 30245013

67. Hsu MY, Inouye M & Inouye S. Retron for the 67-base multicopy single-stranded DNA from Escherichia coli: a potential transposable element encoding both reverse transcriptase and Dam methylase functions. Proc. Natl. Acad. Sci. USA. 1990; 87(23): 9454–9458. doi: 10.1073/pnas.87.23.9454 1701261

68. Hsu MY, Eagle SG, Inouye M, Inouye S. Cell-free synthesis of the branched RNA-linked msDNA from retron-Ec67 of Escherichia coli. J Biol Chem. 1992; 267(20): 13823–13829. 1378431

69. Shimamoto T, Hsu MY, Inouye S, Inouye M. Reverse transcriptases from bacterial retrons require specific secondary structures at the 5’-end of the template for the cDNA priming reaction. J Biol Chem. 1993; 268(4): 2684–2692. 7679101

70. Miyata S, Ohshima A, Inouye S, Inouye M. In vivo production of a stable single-stranded cDNA in Saccharomyces cerevisiae by means of a bacterial retron. Proc. Natl. Acad. Sci. USA. 1192; 89(13): 5735–5739. doi: 10.1073/pnas.89.13.5735 1378616

71. Mirochnitchenko O, Inouye S, and Inouye M. Production of single- stranded DNA in mammalian cells by means of a bacterial retron. J Biol Chem. 1994: 269(4): 2380–2383. 7507924

72. Roy K, Smith J, Vonesch S, Lin G, Szu Tu C, Lederer AR, et al. Multiplexed precision genome editing with trackable genomic barcodes in yeast. Nat Biotechnol. 2018; 36: 512–520. doi: 10.1038/nbt.4137 29734294

73. Nickoloff JA, Chen EY, Heffron F. A 24-base-pair DNA sequence from the MAT locus stimulates intergenic recombination in yeast. Proc Natl Acad Sci USA. 1986; 83(20): 7831–7835. doi: 10.1073/pnas.83.20.7831 3020559

74. Shibata M, Nishimasu H, Kodera N, Hirano S, Ando T, Uchihashi T, et al. Real-space and real-time dynamics of CRISPR-Cas9 visualized by high-speed atomic force microscopy. Nat Commun. 2017; 8(1): 1430. doi: 10.1038/s41467-017-01466-8 29127285

75. Spell RM, Jinks-Robertson S. Examination of the roles of Sgs1 and Srs2 helicase in the enforcement of recombination fidelity in Saccharomyces cerevisiae. Genetics. 2004; 168(4): 1855–1865. doi: 10.1534/genetics.104.032771 15611162

76. Sugawara N, Goldfarb T, Studamire B, Alani E, Haber JE. Heteroduplex rejection during single-strand annealing requires Sgs1 helicase and mismatch repair proteins Msh2 and Msh6 but not Pms1. Proc Natl Acad Sci USA. 2004; 101(25): 9315–9320. doi: 10.1073/pnas.0305749101 15199178

77. Harmsen T, Klaasen S, van de Vrugt H, Te Riele H. DNA mismatch repair and oligonucleotide end-protection promote base-pair substitution distal from a CRISPR/Cas9-induced DNA break. Nucleic Acids Res. 2018; 46(6): 2945–2955. doi: 10.1093/nar/gky076 29447381

78. Bai Y, Symington LS. A Rad52 homolog is required for RAD51-independent mitotic recombination in Saccharomyces cerevisiae. Genes Dev. 1996; 10(16): 2025–2037. doi: 10.1101/gad.10.16.2025 8769646

79. Davis AP, Symington LS. The Rad52-Rad59 complex interacts with Rad51 and replication protein A. DNA Repair. 2003; 2(10): 1127–1134. doi: 10.1016/s1568-7864(03)00121-6 13679150

80. Cortes-Ledesma F, Malagon F, Aguilera A. A novel yeast mutation, rad52-L89F, causes a specific defect in Rad51-independent recombination that correlates with a reduced ability of Rad520L89F to interact with Rad59. Genetics. 2004; 168(1): 553–557. doi: 10.1534/genetics.104.030551 15454565

81. Anand RP, Tsaponina O, Greenwell PW, Lee CS, Du W, Petes TD, et al. Chromosome rearrangements via template switching between diverged repeated sequences. Genes Dev. 2014; 28(21): 2394–2406. doi: 10.1101/gad.250258.114 25367035

82. Ivanov EL, Korolev VG, Fabre F. XRS2, a DNA repair gene of Saccharomyces cerevisiae, is needed for meitotic recombination. Genetics. 1992; 132(3): 651–664. 1468624

83. Gobbini E, Cassani C, Vertemara J, Wang W, Mambretti F, Casari E, et al. The MRX complex regulates Exo1 resection activity by altering DNA end structure. EMBO J. 2018; 37(16): e98588 doi: 10.15252/embj.201798588 29925516

84. Chen W, Jinks-Robertson S. The role of the mismatch repair machinery in regulating mitotic and meiotic recombination between diverged sequences in yeast. Genetics. 1999; 151(4): 1299–1313. 10101158

85. Spell RM, Jinks-Robertson S. Role of mismatch repair in the fidelity of RAD51- and RAD59-dependent recombination in Saccharomyces cerevisiae. Genetics. 2003;165(4):1733–1744. 14704162

86. Haber JE, Ray BL, Kolb JM, White CI. Rapid kinetics of mismatch repair of heteroduplex DNA that is formed during recombination in yeast. Proc Natl Acad Sci USA. 1993; 90(8): 3363–3367. doi: 10.1073/pnas.90.8.3363 8475081

87. Rothstein RJ. One-step gene disruption in yeast. Methods Enzymol. 1983; 101: 202–211. doi: 10.1016/0076-6879(83)01015-0 6310324

88. Anand R, Memisoglu G, Haber JE. Cas9-mediated gene editing in Saccharomyces cerevisiae. Protoc Exch. 2017; http://doi.org/10.1038/protex.2017.021a

89. Benjamini Yoav, Hochberg Yosef. Controlling the false discovery rate: a practical and powerful approach to multiple testing. Journal of the Royal Statistical Society, Series B. 1995; 5(1) 289–300

Článek A single Ho-induced double-strand break at the MAT locus is lethal in Candida glabrata

Článek Novel loci for childhood body mass index and shared heritability with adult cardiometabolic traits

Článek C. elegans CLASP/CLS-2 negatively regulates membrane ingression throughout the oocyte cortex and is required for polar body extrusion

Článek The O-GlcNAc transferase OGT is a conserved and essential regulator of the cellular and organismal response to hypertonic stress

Článek Chromosome separation during Drosophila male meiosis I requires separase-mediated cleavage of the homolog conjunction protein UNO

Článek How noncrossover homologs are conjoined and segregated in Drosophila male meiosis I: Stable but reversible homolog linkers require a novel Separase target protein

Článek Human ABCB1 with an ABCB11-like degenerate nucleotide binding site maintains transport activity by avoiding nucleotide occlusion

Článek Dbp5 associates with RNA-bound Mex67 and Nab2 and its localization at the nuclear pore complex is sufficient for mRNP export and cell viability

Článek RNA-directed DNA Methylation

Článek The DNA methylome of human sperm is distinct from blood with little evidence for tissue-consistent obesity associations

Článek A cautionary note on the use of unsupervised machine learning algorithms to characterise malaria parasite population structure from genetic distance matrices

Článek Selection for ancient periodic motifs that do not impart DNA bending