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Cited by 8DNA Repair MechanismsGenomics and Chromatin DynamicsDNA and Nucleic Acid Chemistry

RTF2 controls replication repriming and ribonucleotide excision at the replisome

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Conti, Brooke A., Ruiz, Penelope D., Broton, Cayla, Blobel, Nicolas J., Kottemann, Molly C., Sridhar, Sunandini, Lach, Francis P., Wiley, Tom F. +3 more·2024·Nature Communications
DOI 10.1038/s41467-024-45947-zPMC PMC10908796
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Abstract

DNA replication through a challenging genomic landscape is coordinated by the replisome, which must adjust to local conditions to provide appropriate replication speed and respond to lesions that hinder its progression. We have previously shown that proteasome shuttle proteins, DNA Damage Inducible 1 and 2 (DDI1/2), regulate Replication Termination Factor 2 (RTF2) levels at stalled replisomes, allowing fork stabilization and restart. Here, we show that during unperturbed replication, RTF2 regulates replisome localization of RNase H2, a heterotrimeric enzyme that removes RNA from RNA-DNA heteroduplexes. RTF2, like RNase H2, is essential for mammalian development and maintains normal replication speed. However, persistent RTF2 and RNase H2 at stalled replication forks prevent efficient replication restart, which is dependent on PRIM1, the primase component of DNA polymerase α-primase. Our data show a fundamental need for RTF2-dependent regulation of replication-coupled ribonucleotide removal and reveal the existence of PRIM1-mediated direct replication restart in mammalian cells.

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  • In humans, RTFDC1 is required for appropriate localization of RNAseH2 to the replication fork to remove ribonucleotides from nascent DNA via ribonucleotide excision repair.

    “However, more recent work has also shown that in humans, RTFDC1 is required for appropriate localization of RNAseH2 to the replication fork to remove ribonucleotides from nascent DNA via the ribonucleotide excision repair pathway70.”

    From Variant-to-function mapping of late-onset Alzheimer’s disease GWAS signals in human microglial cell models implicates <i>RTFDC1</i> at the <i>CASS4</i> locus (2024) · cites this paper as [70]

    The Results section (page not provided) shows that RTF2 loss reduces RNase H2 at replication forks and increases DNA-embedded ribonucleotides; the Introduction says this localization is necessary for proper ribonucleotide excision repair. However, the decisive RTF2-loss experiments used mouse embryonic fibroblasts. The passages do not establish the claim specifically for human RTFDC1. (LLM, conf=0.84)

    Experimental — accuracy not yet measured

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  1. [1]Cortez D Replication-coupled DNA repair Mol. Cell 2019 74 866 876 10.1016/j.molcel.2019.04.027 31173722 PMC6557297 · DOI
  2. [2]Zeman MK Cimprich KA Causes and consequences of replication stress Nat. Cell Biol. 2014 16 2 9 10.1038/ncb2897 24366029 PMC4354890 · DOI · Paper page
  3. [3]Conti BA Smogorzewska A Mechanisms of direct replication restart at stressed replisomes DNA Repair 2020 95 102947 10.1016/j.dnarep.2020.102947 32853827 PMC7669714 · DOI
  4. [4]Kottemann MC Conti BA Lach FP Smogorzewska A Removal of RTF2 from stalled replisomes promotes maintenance of genome integrity Mol. Cell 2018 69 24 35.e25 10.1016/j.molcel.2017.11.035 29290612 PMC6467090 · DOI · Paper page
  5. [5]Inagawa T Schizosaccharomyces pombe Rtf2 mediates site-specific replication termination by inhibiting replication restart Proc. Natl. Acad. Sci. USA 2009 106 7927 7932 10.1073/pnas.0812323106 19416828 PMC2683088 · DOI
  6. [6]Budden AM Schizosaccharomyces pombe Rtf2 is important for replication fork barrier activity of RTS1 via splicing of Rtf1 Elife 2023 12 e78554 10.7554/eLife.78554 37615341 PMC10473836 · DOI
  7. [7]Cerritelli SM Crouch RJ Ribonuclease H: the enzymes in eukaryotes Febs J. 2009 276 1494 1505 10.1111/j.1742-4658.2009.06908.x 19228196 PMC2746905 · DOI
  8. [8]Kellner V Luke B Molecular and physiological consequences of faulty eukaryotic ribonucleotide excision repair Embo J. 2020 39 e102309 10.15252/embj.2019102309 31833079 PMC6996501 · DOI
  9. [9]Eder PS Walder RY Walder JA Substrate specificity of human RNase H1 and its role in excision repair of ribose residues misincorporated in DNA Biochimie 1993 75 123 126 10.1016/0300-9084(93)90033-O 8389211 · DOI
  10. [10]Chon H Contributions of the two accessory subunits, RNASEH2B and RNASEH2C, to the activity and properties of the human RNase H2 complex Nucleic Acids Res. 2009 37 96 110 10.1093/nar/gkn913 19015152 PMC2615623 · DOI
  11. [12]Kind B Altered spatio-temporal dynamics of RNase H2 complex assembly at replication and repair sites in Aicardi-Goutières syndrome Hum. Mol. Genet. 2014 23 5950 5960 10.1093/hmg/ddu319 24986920 · DOI
  12. [13]Reijns MA Enzymatic removal of ribonucleotides from DNA is essential for mammalian genome integrity and development Cell 2012 149 1008 1022 10.1016/j.cell.2012.04.011 22579044 PMC3383994 · DOI · Paper page
  13. [14]Alabert C Nascent chromatin capture proteomics determines chromatin dynamics during DNA replication and identifies unknown fork components Nat. Cell Biol. 2014 16 281 293 10.1038/ncb2918 24561620 PMC4283098 · DOI
  14. [15]Wessel SR Mohni KN Luzwick JW Dungrawala H Cortez D Functional analysis of the replication fork proteome identifies BET proteins as PCNA regulators Cell Rep. 2019 28 3497 3509.e3494 10.1016/j.celrep.2019.08.051 31553917 PMC6878991 · DOI
  15. [16]Dungrawala H The replication checkpoint prevents two types of fork collapse without regulating replisome stability Mol. Cell 2015 59 998 1010 10.1016/j.molcel.2015.07.030 26365379 PMC4575883 · DOI
  16. [17]Heuzé J RNase H2 degrades toxic RNA:DNA hybrids behind stalled forks to promote replication restart Embo J. 2023 42 e113104 10.15252/embj.2022113104 37855233 PMC10690446 · DOI
  17. [18]Murante RS Henricksen LA Bambara RA Junction ribonuclease: an activity in Okazaki fragment processing Proc. Natl. Acad. Sci. USA 1998 95 2244 2249 10.1073/pnas.95.5.2244 9482870 PMC19307 · DOI
  18. [19]Turchi JJ Huang L Murante RS Kim Y Bambara RA Enzymatic completion of mammalian lagging-strand DNA replication Proc. Natl. Acad. Sci. USA 1994 91 9803 9807 10.1073/pnas.91.21.9803 7524089 PMC44905 · DOI
  19. [20]Chen JZ Qiu J Shen B Holmquist GP Mutational spectrum analysis of RNase H(35) deficient Saccharomyces cerevisiae using fluorescence-based directed termination PCR Nucleic Acids Res. 2000 28 3649 3656 10.1093/nar/28.18.3649 10982888 PMC110751 · DOI
  20. [21]Ohle C Transient RNA-DNA hybrids are required for efficient double-strand break repair Cell 2016 167 1001 1013.e1007 10.1016/j.cell.2016.10.001 27881299 · DOI
  21. [22]Qiu J Qian Y Frank P Wintersberger U Shen B Saccharomyces cerevisiae RNase H(35) functions in RNA primer removal during lagging-strand DNA synthesis, most efficiently in cooperation with Rad27 nuclease Mol. Cell Biol. 1999 19 8361 8371 10.1128/MCB.19.12.8361 10567561 PMC84926 · DOI
  22. [23]Berti M Vindigni A Replication stress: getting back on track Nat. Struct. Mol. Biol. 2016 23 103 109 10.1038/nsmb.3163 26840898 PMC5125612 · DOI
  23. [24]Heller RC Marians KJ Replisome assembly and the direct restart of stalled replication forks Nat. Rev. Mol. Cell Biol. 2006 7 932 943 10.1038/nrm2058 17139333 · DOI · Paper page
  24. [25]Petermann E Helleday T Pathways of mammalian replication fork restart Nat. Rev. Mol. Cell Biol. 2010 11 683 687 10.1038/nrm2974 20842177 · DOI · Paper page
  25. [26]Yeeles JT Poli J Marians KJ Pasero P Rescuing stalled or damaged replication forks Cold Spring Harb. Perspect. Biol. 2013 5 a012815 10.1101/cshperspect.a012815 23637285 PMC3632063 · DOI
  26. [27]Taylor MRG Yeeles JTP The initial response of a eukaryotic replisome to DNA damage Mol. Cell 2018 70 1067 1080.e1012 10.1016/j.molcel.2018.04.022 29944888 PMC6024075 · DOI
  27. [28]Bianchi J PrimPol bypasses UV photoproducts during eukaryotic chromosomal DNA replication Mol. Cell 2013 52 566 573 10.1016/j.molcel.2013.10.035 24267451 PMC4228047 · DOI
  28. [29]García-Gómez S PrimPol, an archaic primase/polymerase operating in human cells Mol. Cell 2013 52 541 553 10.1016/j.molcel.2013.09.025 24207056 PMC3899013 · DOI
  29. [30]Mouron S Repriming of DNA synthesis at stalled replication forks by human PrimPol Nat. Struct. Mol. Biol. 2013 20 1383 1389 10.1038/nsmb.2719 24240614 · DOI
  30. [31]Schiavone D PrimPol is required for replicative tolerance of G Quadruplexes in Vertebrate Cells Mol. Cell 2016 61 161 169 10.1016/j.molcel.2015.10.038 26626482 PMC4712188 · DOI
  31. [32]Wan L hPrimpol1/CCDC111 is a human DNA primase-polymerase required for the maintenance of genome integrity EMBO Rep. 2013 14 1104 1112 10.1038/embor.2013.159 24126761 PMC3981091 · DOI
  32. [33]Vassilev LT Selective small-molecule inhibitor reveals critical mitotic functions of human CDK1 Proc. Natl. Acad. Sci. USA 2006 103 10660 10665 10.1073/pnas.0600447103 16818887 PMC1502288 · DOI
  33. [34]Alabert C Groth A Chromatin replication and epigenome maintenance Nat. Rev. Mol. Cell Biol. 2012 13 153 10.1038/nrm3288 22358331 · DOI
  34. [35]Cortez, D. Preventing replication fork collapse to maintain genome integrity. DNA repair. 10.1016/j.dnarep.2015.04.026 (2015). PMC4522347 25957489 · DOI · Paper page
  35. [36]Sasaki T An Rtf2 domain-containing protein influences Pre-mRNA splicing and is essential for embryonic development in Arabidopsis thaliana Genetics 2015 200 523 535 10.1534/genetics.115.176438 25819795 PMC4492377 · DOI
  36. [37]Dungrawala H Cortez D Purification of proteins on newly synthesized DNA using iPOND Meth. Mol. Biol. (Clifton, N.J.) 2015 1228 123 131 10.1007/978-1-4939-1680-1_10 PMC4384176 25311126 · DOI
  37. [38]Taglialatela A Restoration of Replication Fork Stability in BRCA1- and BRCA2-Deficient Cells by Inactivation of SNF2-Family Fork Remodelers Mol. Cell 2017 68 414 430.e418 10.1016/j.molcel.2017.09.036 29053959 PMC5720682 · DOI
  38. [39]Reijns MA The structure of the human RNase H2 complex defines key interaction interfaces relevant to enzyme function and human disease J. Biol. Chem. 2011 286 10530 10539 10.1074/jbc.M110.177394 21177854 PMC3060506 · DOI
  39. [40]Zimmermann M CRISPR screens identify genomic ribonucleotides as a source of PARP-trapping lesions Nature 2018 559 285 289 10.1038/s41586-018-0291-z 29973717 PMC6071917 · DOI
  40. [41]Chon H RNase H2 roles in genome integrity revealed by unlinking its activities Nucleic Acids Res. 2013 41 3130 3143 10.1093/nar/gkt027 23355612 PMC3597693 · DOI
  41. [42]Benitez-Guijarro, M. et al. RNase H2, mutated in Aicardi-Goutières syndrome, promotes LINE-1 retrotransposition. Embo J.37. 10.15252/embj.201798506 (2018) PMC6068448 29959219 · DOI
  42. [43]Chen L R-ChIP Using Inactive RNase H Reveals Dynamic Coupling of R-loops with transcriptional pausing at gene promoters Mol. Cell 2017 68 745 757.e745 10.1016/j.molcel.2017.10.008 29104020 PMC5957070 · DOI
  43. [45]Niu H Potenski CJ Epshtein A Sung P Klein HL Roles of DNA helicases and Exo1 in the avoidance of mutations induced by Top1-mediated cleavage at ribonucleotides in DNA Cell Cycle (Georgetown, Tex.) 2016 15 331 336 10.1080/15384101.2015.1128594 26716562 PMC4943705 · DOI
  44. [46]Sparks JL Burgers PM Error-free and mutagenic processing of topoisomerase 1-provoked damage at genomic ribonucleotides Embo J. 2015 34 1259 1269 10.15252/embj.201490868 25777529 PMC4426484 · DOI
  45. [48]George JW Bessho M Bessho T Inactivation of XPF sensitizes cancer cells to gemcitabine J. Nucleic Acids 2019 2019 6357609 10.1155/2019/6357609 30941207 PMC6421022 · DOI
  46. [49]Iyer LM Koonin EV Leipe DD Aravind L Origin and evolution of the archaeo-eukaryotic primase superfamily and related palm-domain proteins: structural insights and new members Nucleic Acids Res. 2005 33 3875 3896 10.1093/nar/gki702 16027112 PMC1176014 · DOI
  47. [50]Quinet, A. et al. PRIMPOL-Mediated Adaptive Response Suppresses Replication Fork Reversal in BRCA-Deficient Cells. Mol. Cell. 10.1016/j.molcel.2019.10.008 (2019). PMC7007862 31676232 · DOI
  48. [51]Holzer S Structural basis for inhibition of human primase by arabinofuranosyl nucleoside analogues fludarabine and vidarabine ACS Chem. Biol. 2019 14 1904 1912 10.1021/acschembio.9b00367 31479243 PMC6757278 · DOI
  49. [52]Mirman Z Sasi NK King A Chapman JR de Lange T 53BP1-shieldin-dependent DSB processing in BRCA1-deficient cells requires CST-Polα-primase fill-in synthesis Nat. Cell Biol. 2022 24 51 61 10.1038/s41556-021-00812-9 35027730 PMC8849574 · DOI
  50. [53]Taylor MRG Yeeles JTP Dynamics of replication fork progression following helicase-polymerase uncoupling in eukaryotes J. Mol. Biol. 2019 431 2040 2049 10.1016/j.jmb.2019.03.011 30894292 PMC6525111 · DOI
  51. [54]Higuchi K Fate of DNA replication fork encountering a single DNA lesion during oriC plasmid DNA replication in vitro Genes Cells 2003 8 437 449 10.1046/j.1365-2443.2003.00646.x 12694533 · DOI
  52. [55]McInerney P O’Donnell M Functional uncoupling of twin polymerases: mechanism of polymerase dissociation from a lagging-strand block J. Biol. Chem 2004 279 21543 21551 10.1074/jbc.M401649200 15014081 · DOI
  53. [56]Williams JS Lujan SA Kunkel TA Processing ribonucleotides incorporated during eukaryotic DNA replication Nat. Rev. Mol. Cell Biol. 2016 17 350 363 10.1038/nrm.2016.37 27093943 PMC5445644 · DOI · Paper page
  54. [57]Kuchta RD Stengel G Mechanism and evolution of DNA primases Biochim. Biophys. Acta 2010 1804 1180 1189 10.1016/j.bbapap.2009.06.011 19540940 PMC2846230 · DOI
  55. [59]Cordoba, J. J., Mullins, E. A., Salay, L. E., Eichman, B. F. & Chazin, W. J. Flexibility and distributive synthesis regulate RNA priming and handoff in human DNA polymerase α-primase. J. Mol. Biol. 168330. 10.1016/j.jmb.2023.168330 (2023) PMC10872500 37884206 · DOI
  56. [60]Crow YJ Mutations in genes encoding ribonuclease H2 subunits cause Aicardi-Goutières syndrome and mimic congenital viral brain infection Nat. Genet. 2006 38 910 916 10.1038/ng1842 16845400 · DOI
  57. [61]Parry DA PRIM1 deficiency causes a distinctive primordial dwarfism syndrome Genes Dev. 2020 34 1520 1533 10.1101/gad.340190.120 33060134 PMC7608753 · DOI
  58. [62]Rodriguez CI High-efficiency deleter mice show that FLPe is an alternative to Cre-loxP Nat. Genet. 2000 25 139 140 10.1038/75973 10835623 · DOI
  59. [63]Lakso M Efficient in vivo manipulation of mouse genomic sequences at the zygote stage Proc. Natl. Acad. Sci. USA 1996 93 5860 5865 10.1073/pnas.93.12.5860 8650183 PMC39152 · DOI · Paper page
  60. [64]Zhu JY Abate M Rice PW Cole CN The ability of simian virus 40 large T antigen to immortalize primary mouse embryo fibroblasts cosegregates with its ability to bind to p53 J. Virol. 1991 65 6872 10.1128/jvi.65.12.6872-6880.1991 1658380 PMC250785 · DOI
  61. [65]Silver DP Livingston DM Self-excising retroviral vectors encoding the cre recombinase overcome cre-mediated cellular toxicity Mol. Cell 2001 8 233 243 10.1016/S1097-2765(01)00295-7 11511376 · DOI
  62. [66]Ran FA Genome engineering using the CRISPR-Cas9 system Nat. Protocols 2013 8 2281 2308 10.1038/nprot.2013.143 24157548 PMC3969860 · DOI · Paper page
  63. [67]Lamesch P hORFeome v3.1: a resource of human open reading frames representing over 10,000 human genes Genomics 2007 89 307 315 10.1016/j.ygeno.2006.11.012 17207965 PMC4647941 · DOI
  64. [68]Agrotis A Pengo N Burden JJ Ketteler R Redundancy of human ATG4 protease isoforms in autophagy and LC3/GABARAP processing revealed in cells Autophagy 2019 15 976 997 10.1080/15548627.2019.1569925 30661429 PMC6526816 · DOI
  65. [69]Schlabach MR Cancer proliferation gene discovery through functional genomics Science (New York, N.Y.) 2008 319 620 624 10.1126/science.1149200 18239126 PMC2981870 · DOI
  66. [70]Cong L Multiplex genome engineering using CRISPR/Cas systems Science (New York, N.Y.) 2013 339 819 823 10.1126/science.1231143 23287718 PMC3795411 · DOI · Paper page
  67. [71]Mendez J Stillman B Chromatin association of human origin recognition complex, cdc6, and minichromosome maintenance proteins during the cell cycle: assembly of prereplication complexes in late mitosis Mol. Cell Biol. 2000 20 8602 8612 10.1128/MCB.20.22.8602-8612.2000 11046155 PMC102165 · DOI
  68. [72]Rappsilber J Mann M Ishihama Y Protocol for micro-purification, enrichment, pre-fractionation and storage of peptides for proteomics using StageTips Nat. Protoc. 2007 2 1896 1906 10.1038/nprot.2007.261 17703201 · DOI · Paper page
  69. [73]Olsen JV Parts per million mass accuracy on an Orbitrap mass spectrometer via lock mass injection into a C-trap Mol. Cell Proteomics 2005 4 2010 2021 10.1074/mcp.T500030-MCP200 16249172 · DOI
  70. [74]Bunkenborg J Garcia GE Paz MI Andersen JS Molina H The minotaur proteome: avoiding cross-species identifications deriving from bovine serum in cell culture models Proteomics 2010 10 3040 3044 10.1002/pmic.201000103 20641139 · DOI
  71. [75]Spivak M Weston J Bottou L Kall L Noble WS Improvements to the percolator algorithm for Peptide identification from shotgun proteomics data sets J. Proteome Res. 2009 8 3737 3745 10.1021/pr801109k 19385687 PMC2710313 · DOI
  72. [76]Silva JC Gorenstein MV Li GZ Vissers JP Geromanos SJ Absolute quantification of proteins by LCMSE: a virtue of parallel MS acquisition Mol. Cell Proteomics 2006 5 144 156 10.1074/mcp.M500230-MCP200 16219938 · DOI · Paper page
  73. [77]Gallo D Wang G Yip CM Brown GW Analysis of replicating yeast chromosomes by DNA combing Cold Spring Harbor Protocols 2016 2016 pdb.prot085118 10.1101/pdb.prot085118 26832684 · DOI
  74. [78]Gyori BM Venkatachalam G Thiagarajan PS Hsu D Clement MV OpenComet: an automated tool for comet assay image analysis Redox Biol. 2014 2 457 465 10.1016/j.redox.2013.12.020 24624335 PMC3949099 · DOI

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Authors
Conti, B. A., Ruiz, P. D., Broton, C., Blobel, N. J., Kottemann, M. C., Sridhar, S., Lach, F. P., Wiley, T. F., Sasi, N. K., Carroll, T., & Smogorzewska, A.
Affiliations
Rockefeller University (US)
Corresponding author
Smogorzewska, Agata
ORCID
Venue
Nature Communications
Publisher
Springer Science and Business Media LLC
Published
2024-03-02
Volume
15
Issue
1
Pages
1943-1943
ISSN
2041-1723 (ISSN-L)
DOI
10.1038/s41467-024-45947-z
PMC
PMC10908796
OpenAlex
W4392348878
Cited by
8
References
4
Citations by year
2026: 4 · 2025: 2 · 2024: 2
FWCI
1.08
Type
journal article
Language
en
Open access
Licence
Locations
  • Nature Communications (publishedVersion, cc-by)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by-nc-nd)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • PubMed Central (submittedVersion, cc-by-nc-nd)
  • RePEc: Research Papers in Economics (submittedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion)
Funders
U.S. Department of Health & Human Services | NIH | National Institute of General Medical Sciences (R01GM140400, GM066699, GM007739, GM143866)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA Repair Mechanisms
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Genomics and Chromatin Dynamics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA and Nucleic Acid Chemistry
Keywords
RNase H2, replication restart, replication fork stalling, replisome, PRIM1, DNA polymerase alpha-primase, RNA-DNA heteroduplex, mammalian development

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