Genetic Approaches in Pluripotent Stem Cell Studies and Practical Applications

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Abstract

The discovery of pluripotent stem cells (PSCs) had a tremendous impact in various areas of biology, while the application of genetic approaches to these cells further expanded the repertoire of both fundamental and applied tasks addressed with the help of PSCs. For example, mouse embryonic stem cells (ESCs) have served as an indispensable tool in functional genetics, which allowed to define biological role of thousands of mouse genes using gene knockout tecniques. Subsequent discovery of human ESCs opened bond perspectives in tissue replacement therapy, while the discovery of induced pluripotent stem cells (iPSCs), which eliminated several technical, immunological, and ethical issues associated with the derivation and clinical use of ESCs, has made the perspectives even closer. The manuscript overviews some of genetic approaches that, in combination with PSC technologies, can provide a new level of solution to a wide range of biomedical tasks.

About the authors

A. A Kuzmin

Institute of Cytology of the Russian Academy of Sciences

St. Petersburg, Russia

S. A Sinenko

Institute of Cytology of the Russian Academy of Sciences

St. Petersburg, Russia

E. I Bakhmet

Institute of Cytology of the Russian Academy of Sciences

St. Petersburg, Russia

A. S Fotina

Institute of Cytology of the Russian Academy of Sciences

St. Petersburg, ussia

V. V Ermakova

Institute of Cytology of the Russian Academy of Sciences

St. Petersburg, Russia

A. N Tomilin

Institute of Cytology of the Russian Academy of Sciences

Email: a.tomilin@incras.ru
St. Petersburg, Russia

References

  1. Evans M.J., Kaufman M.H. Establishment in culture of pluripotential cells from mouse embryos // Nature. 1981. V. 292. № 5819. P. 154–156. https://doi.org/10.1038/292154a0
  2. Martin G.R. Isolation of a pluripotent cell line from early mouse embryos cultured in medium conditioned by teratocarcinoma stem cells // PNAS USA. 1981. V. 78. № 12. P. 7634–7638. https://doi.org/10.1073/pnas.78.12.7634
  3. Gordeev M.N., Bakhmet E.I., Tomilin A.N. Pluripotency dynamics during embryogenesis and in cell culture // Russ. J. Dev. Biol. 2021. V. 52. № 6. P. 379–389. https://doi.org/10.1134/s1062360421060059
  4. Thomas K.R., Capecchi M.R. Site-directed mutagenesis by gene targeting in mouse embryo-derived stem cells // Cell. 1987. V. 51. № 3. P. 503–512. https://doi.org/10.1016/0092-8674(87)90646-5
  5. Knott G.J., Doudna J.A. CRISPR-Cas guides the future of genetic engineering // Sci. 2018. V. 361. № 6405. P. 866–869. https://doi.org/10.1126/science.aat5011
  6. Thomson J.A., Itskovitz-Eldor J., Shapiro S.S. et al. Embryonic stem cell lines derived from human blastocysts // Science. 1998. V. 282. № 5391. P. 1145–1147. https://doi.org/10.1126/science.282.5391.1145
  7. Takahashi K., Yamanaka S. Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors // Cell. 2006. V. 126. № 4. P. 663–676. https://doi.org/10.1016/j.cell.2006.07.024
  8. Takahashi K., Tanabe K., Ohnuki M. et al. Induction of pluripotent stem cells from adult human fibroblasts by defined factors // Cell. 2007. V. 131. № 5. P. 861–872. https://doi.org/10.1016/J.CELL.2007.11.019
  9. Cyranoski D. “Reprogrammed” stem cells approved to mend human hearts for the first time news /631/532 // Nature. 2018. V. 557. № 7707. P. 619–620. https://doi.org/10.1038/d41586-018-05278-8
  10. Young C.S., Hicks M.R., Ermolova N.V. et al. A single CRISPR-Cas9 deletion strategy that targets the majority of DMD patients restores dystrophin function in hiPSC-derived muscle cells // Cell Stem Cell. 2016. V. 18. № 4. P. 533–540. https://doi.org/10.1016/j.stem.2016.01.021
  11. Hoang D.M., Pham P.T., Bach T.Q. et al. Stem cellbased therapy for human diseases // Signal Transduct. Target Ther. 2022. V. 7. № 1. P. 272. https://doi.org/10.1038/s41392-022-01134-4
  12. Ortuño-Costela M.D.C., Cerrada V., García-López M., Gallardo M.E. The challenge of bringing iPSCs to the patient // Int. J. Mol. Sci. 2019. V. 20. № 24. https://doi.org/10.3390/ijms20246305
  13. Pfeifer A., Ikawa M., Dayn Y., Verma I.M. Transgenesis by lentiviral vectors: Lack of gene silencing in mammalian embryonic stem cells and preimplantation embryos // PNAS USA. 2002. V. 99. № 4. P. 2140–2145. https://doi.org/10.1073/pnas.251682798
  14. Hamaguchi I., Woods N.B., Panagopoulos I. et al. Lentivirus vector gene expression during ES cell-derived hematopoietic development in vitro // J. Virol. 2000. V. 74. № 22. https://doi.org/10.1128/jvi.74.22.10778-10784.2000
  15. Asano T., Hanazono Y., Ueda Y. et al. Highly efficient gene transfer into primate embryonic stem cells with a simian with a lentivirus vector // Mol. Therapy. 2002. V. 6. № 2. P. 162–168. https://doi.org/10.1006/mthe.2002.0655
  16. Ivics Z., Hackett P.B., Plasterk R.H., Izsvák Z. Molecular reconstruction of sleeping beauty, a Tc1-like transposon from fish, and its transposition in human cells // Cell. 1997. V. 91. № 4. P. 501–510. https://doi.org/10.1016/S0092-8674(00)80436-5
  17. Ivics Z., Li M.A., Mátés L. et al. Transposon-mediated genome manipulation in vertebrates // Nat. Methods. 2009. V. 6. № 6. P. 415–422. https://doi.org/10.1038/nmeth.1332
  18. Woltjen K., Michael I.P., Mohseni P. et al. PiggyBac transposition reprograms fibroblasts to induced pluripotent stem cells // Nature. 2009. V. 458. № 7239. P. 766–770. https://doi.org/10.1038/nature07863
  19. Yusa K., Rad R., Takeda J., Bradley A. Generation of transgene-free induced pluripotent mouse stem cells by the piggyBac transposon // Nat. Methods. 2009. V. 6. № 5. P. 363–369. https://doi.org/10.1038/nmeth.1323
  20. Rostovskaya M., Fu J., Obst M. et al. Transposon-mediated BAC transgenesis in human ES cells // Nucl. Ac. Res. 2012. V. 40. № 19. e150. https://doi.org/10.1093/nar/gks643
  21. Bhaya D., Davison M., Barrangou R. CRISPR-cas systems in bacteria and archaea: Versatile small RNAs for adaptive defense and regulation // Annu. Rev. Genet. 2011. V. 45. P. 273–297. https://doi.org/10.1146/annurev-genet-110410-132430
  22. Jinek M., Chylinski K., Fonfara I. et al. A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity // Science. 2012. V. 337. № 6096. P. 816–821. https://doi.org/10.1126/science.1225829
  23. Zetsche B., Gootenberg J.S., Abudayyeh O.O. et al. Cpf1 is a single RNA-guided endonuclease of a class 2 CRISPR-Cas system // Cell. 2015. V. 163. № 3. P. 759–771. https://doi.org/10.1016/j.cell.2015.09.038
  24. Mao Z., Bozzella M., Seluanov A., Gorbunova V. Comparison of nonhomologous end joining and homologous recombination in human cells // DNA Repair (Amst.). 2008. V. 7. № 10. P. 1765–1771. https://doi.org/10.1016/j.dnarep.2008.06.018
  25. Sfeir A., Symington L.S. Microhomology-mediated end joining: A back-up survival mechanism or dedicated pathway? // Trends Biochem. Sci. 2015. V. 40. № 11. P. 701–714. https://doi.org/10.1016/j.tibs.2015.08.006
  26. Ran F.A., Hsu P.D., Wright J. et al. Genome engineering using the CRISPR-Cas9 system // Nat. Protoc. 2013. V. 8. № 11. P. 2281–2308. https://doi.org/10.1038/nprot.2013.143
  27. Saleh-Gohari N., Helleday T. Conservative homologous recombination preferentially repairs DNA double-strand breaks in the S phase of the cell cycle un human cells // Nucl. Ac. Res. 2004. V. 32. № 12. P. 3683–3688. https://doi.org/10.1093/nar/gkh703
  28. Pardo B., Gómez-González B., Aguilera A. DNA repair in mammalian cells: DNA double-strand break repair: how to fix a broken relationship // Cell Mol. Life Sci. 2009. V. 66. № 6. P. 1039–1056. https://doi.org/10.1007/s00018-009-8740-3
  29. Yang H., Ren S., Yu S. et al. Methods favoring homology-directed repair choice in response to CRISPR/ Cas9 induced-double strand breaks // Int. J. Mol. Sci. 2020. V. 21. № 18. https://doi.org/10.3390/ijms21186461
  30. Miyaoka Y., Mayerl S.J., Chan A.H., Conklin B.R. Detection and quantification of HDR and NHEJ induced by genome editing at endogenous gene loci u s i n g d r o p l e t d i g i t a l P C R / / Methods Mol. Biol. 2018. V. 1768. P. 349–362. https://doi.org/10.1007/978-1-4939-7778-9_20
  31. Vitale I., Manic G., De Maria R. et al. DNA damage in stem cells // Mol. Cell. 2017. V. 66. № 3. P. 306–319. https://doi.org/10.1016/j.molcel.2017.04.006
  32. Reuven N., Shaul Y. Selecting for CRISPR-edited knock-in cells // Int. J. Mol. Sci. 2022. V. 23. № 19. https://doi.org/10.3390/ijms231911919
  33. Selvaraj S., Feist W.N., Viel S. et al. High-efficiency transgene integration by homology-directed repair in human primary cells using DNA-PKcs inhibition // Nat. Biotechnol. 2024. V. 42. № 5. P. 731–744. https://doi.org/10.1038/s41587-023-01888-4
  34. Singh A., Smedley G.D., Rose J.G. et al. A high efficiency precision genome editing method with CRISPR in iPSCs // Sci. Rep. 2024. V. 14. № 1. P. 9933. https://doi.org/10.1038/s41598-024-60766-4
  35. Yu C., Liu Y., Ma T. et al. Small molecules enhance crispr genome editing in pluripotent stem cells // Cell Stem Cell. 2015. V. 16. № 2. https://doi.org/10.1016/j.stem.2015.01.003
  36. Guo Q., Mintier G., Ma-Edmonds M. et al. “Cold shock” increases the frequency of homology directed repair gene editing in induced pluripotent stem cells // Sci. Rep. 2018. V. 8. № 1. P. 2080. https://doi.org/10.1038/s41598-018-20358-5
  37. Richardson C.D., Ray G.J., DeWitt M.A. et al. Enhancing homology-directed genome editing by catalytically active and inactive CRISPR-Cas9 using asymmetric donor DNA // Nat. Biotechnol. 2016. V. 34. № 3. P. 339–344. https://doi.org/10.1038/nbt.3481
  38. Liang X., Potter J., Kumar S. et al. Enhanced CRISPR/ Cas9-mediated precise genome editing by improved design and delivery of gRNA, Cas9 nuclease, and donor DNA // J. Biotechnol. 2017. V. 241. P. 136–146. https://doi.org/10.1016/j.jbiotec.2016.11.011
  39. Anzalone A.V., Randolph P.B., Davis J.R. et al. Searchand-replace genome editing without double-strand breaks or donor DNA // Nature. 2019. V. 576. № 7785. P. 149–157. https://doi.org/10.1038/s41586-019-1711-4
  40. Li H., Busquets O., Verma Y. et al. Highly efficient generation of isogenic pluripotent stem cell models using prime editing // Elife. 2022. V. 11. https://doi.org/10.7554/eLife.79208
  41. Chen P.J., Hussmann J.A., Yan J. et al. Enhanced prime editing systems by manipulating cellular determinants of editing outcomes // Cell. 2021. V. 184. № 22. P. 5635–5652.e29. https://doi.org/10.1016/j.cell.2021.09.018
  42. Wu Y., Zhong A., Sidharta M. et al. Robust and inducible genome editing via an all-in-one prime editor in human pluripotent stem cells // Nat. Commun. 2024. V. 15. № 1. P. 10824. https://doi.org/10.1038/s41467-024-55104-1
  43. Eggenschwiler R., Gschwendtberger T., Felski C. et al. A selectable all-in-one CRISPR prime editing piggyBac transposon allows for highly efficient gene editing in human cell lines // Sci. Rep. 2021. V. 11. № 1. P. 22154. https://doi.org/10.1038/s41598-021-01689-2
  44. Zheng C., Liu B., Dong X. et al. Template-jumping prime editing enables large insertion and exon rewriting in vivo // Nat. Commun. 2023. V. 14. № 1. P. 3369. https://doi.org/10.1038/s41467-023-39137-6
  45. Klompe S.E., Vo P.L.H., Halpin-Healy T.S., Sternberg S.H. Transposon-encoded CRISPR–Cas systems direct RNA-guided DNA integration // Nature. 2019. V. 571. № 7764. P. 219–225. https://doi.org/10.1038/s41586-019-1323-z
  46. Lampe G.D., King R.T., Halpin-Healy T.S. et al. Targeted DNA integration in human cells without double-strand breaks using CRISPR-associated transposases // Nat. Biotechnol. 2024. V. 42. № 1. P. 87–98. https://doi.org/10.1038/s41587-023-01748-1
  47. Ye L., Chang J.C., Lin C. et al. Generation of induced pluripotent stem cells using site-specific integration with phage integrase // PNAS USA. 2010. V. 107. № 45. P. 19467–19472. https://doi.org/10.1073/pnas.1012677107
  48. Duportet X., Wroblewska L., Guye P. et al. A platform for rapid prototyping of synthetic gene networks in mammalian cells // Nucl. Ac. Res. 2014. V. 42. № 21. P. 13440–13451. https://doi.org/10.1093/nar/gku1082
  49. Rath P., Kramer P., Biggs D. et al. Optimizing approaches for targeted integration of transgenic cassettes by integrase-mediated cassette exchange in mouse and human stem cells // Stem Cells. 2025. V. 43. № 1. https://doi.org/10.1093/STMCLS/SXAE092
  50. Blanch-Asensio A., Grandela C., Brandão K.O. et al. STRAIGHT-IN enables high-throughput targeting of large DNA payloads in human pluripotent stem cells // Cell Rep. Meth. 2022. V. 2. № 10. https://doi.org/10.1016/j.crmeth.2022.100300
  51. Pandey S., Gao X.D., Krasnow N.A. et al. Efficient site-specific integration of large genes in mammalian cells via continuously evolved recombinases and prime editing // Nat. Biomed. Eng. 2024. V. 9. № 1. P. 22–39. https://doi.org/10.1038/s41551-024-01227-1
  52. Yarnall M.T.N., Ioannidi E.I., Schmitt-Ulms C. et al. Drag-and-drop genome insertion of large sequences without double-strand DNA cleavage using CRISPR-directed integrases // Nat. Biotechnol. 2023. V. 41. № 4. P. 500–512. https://doi.org/10.1038/s41587-022-01527-4
  53. Durrant M.G., Fanton A., Tycko J. et al. Systematic discovery of recombinases for efficient integration of large DNA sequences into the human genome // Nat. Biotechnol. 2023. V. 41. № 4. P. 488–499. https://doi.org/10.1038/s41587-022-01494-w
  54. Qi L.S., Larson M.H., Gilbert L.A. et al. Repurposing CRISPR as an RNA-γuided platform for sequence-specific control of gene expression // Cell. 2013. V. 152. № 5. P. 1173–1183. https://doi.org/10.1016/j.cell.2013.02.022
  55. O’Geen H., Bates S.L., Carter S.S. et al. Ezh2-dCas9 and KRAB-dCas9 enable engineering of epigenetic memory in a context-dependent manner // Epigen. Chromat. 2019. V. 12. № 1. P. 26. https://doi.org/10.1186/s13072-019-0275-8
  56. Ziller M.J., Ortega J.A., Quinlan K.A. et al. Dissecting the Functional Consequences of De Novo DNA Methylation dynamics in human motor neuron differentiation and physiology // Cell Stem Cell. 2018. V. 22. № 4. P. 559–574.e9. https://doi.org/10.1016/j.stem.2018.02.012
  57. Perez-Pinera P., Kocak D.D., Vockley C.M. et al. RNA-guided gene activation by CRISPR-Cas9-based transcription factors // Nat. Methods. 2013. V. 10. № 10. P. 973–976. https://doi.org/10.1038/nmeth.2600
  58. Chavez A., Scheiman J., Vora S. et al. Highly efficient Cas9-mediated transcriptional programming // Nat. Methods. 2015. V. 12. № 4. P. 326–328. https://doi.org/10.1038/nmeth.3312
  59. Tanenbaum M.E., Gilbert L.A., Qi L.S. et al. A protein-tagging system for signal amplification in gene expression and fluorescence imaging // Cell. 2014. V. 159. № 3. P. 635–646. https://doi.org/10.1016/j.cell.2014.09.039
  60. Konermann S., Brigham M.D., Trevino A.E. et al. Genome-scale transcriptional activation by an engineered CRISPR-Cas9 complex // Nature. 2015. V. 517. № 7536. P. 583–588. https://doi.org/10.1038/nature14136
  61. Mandegar M.A., Huebsch N., Frolov E.B. et al. CRISPR interference efficiently induces specific and reversible gene silencing in human iPSCs // Cell Stem Cell. 2016. V. 18. № 4. P. 541–553. https://doi.org/10.1016/j.stem.2016.01.022
  62. Liu P., Chen M., Liu Y. et al. CRISPR-Based chromatin remodeling of the endogenous Oct4 or Sox2 locus enables reprogramming to pluripotency // Cell Stem Cell. 2018. V. 22. № 2. P. 252–261.e4. https://doi.org/10.1016/j.stem.2017.12.001
  63. Balboa D., Weltner J., Eurola S. et al. Conditionally stabilized dCas9 activator for controlling gene expression in human cell reprogramming and differentiation // Stem Cell Reports. 2015. V. 5. № 3. P. 448–459. https://doi.org/10.1016/j.stemcr.2015.08.001
  64. Wei S., Zou Q., Lai S. et al. Conversion of embryonic stem cells into extraembryonic lineages by CRISPR-mediated activators // Sci. Rep. 2016. V. 6. https://doi.org/10.1038/srep19648
  65. Chakraborty S., Ji H., Kabadi A.M. et al. A CRISPR/ Cas9-based system for reprogramming cell lineage specification // Stem Cell Reports. 2014. V. 3. № 6. P. 940–947. https://doi.org/10.1016/j.stemcr.2014.09.013
  66. Black J.B., Adler A.F., Wang H.G. et al. Targeted epigenetic remodeling of endogenous loci by CRISPR/ Cas9-Based transcriptional activators directly converts fibroblasts to neuronal cells // Cell Stem Cell. 2016. V. 19. № 3. P. 406–414. https://doi.org/10.1016/j.stem.2016.07.001
  67. Khoo T.S., Jamal R., Abdul Ghani N.A. et al. Retention of somatic memory associated with cell identity, age and metabolism in induced pluripotent stem (iPS) cells reprogramming // Stem Cell Rev. Rep. 2020. V. 16. № 2. P. 251–261. https://doi.org/10.1007/s12015-020-09956-x
  68. Blanco E., González-Ramírez M., Alcaine-Colet A. et al. The bivalent genome: Characterization, structure, and regulation // Trends Genet. 2020. V. 36. № 2. P. 118–131. https://doi.org/10.1016/j.tig.2019.11.004
  69. Nakamura M., Gao Y., Dominguez A.A., Qi L.S. CRISPR technologies for precise epigenome editing // Nat. Cell Biol. 2021. V. 23. № 1. P. 11–22. https://doi.org/10.1038/s41556-020-00620-7
  70. Gao X., Tsang J.C.H., Gaba F. et al. Comparison of TALE designer transcription factors and the CRISPR/ dCas9 in regulation of gene expression by targeting enhancers // Nucl. Ac. Res. 2014. V. 42. № 20. e155. https://doi.org/10.1093/nar/gku836
  71. Schoger E., Argyriou L., Zimmermann W.H. et al. Generation of homozygous CRISPRa human induced pluripotent stem cell (hiPSC) lines for sustained endogenous gene activation // Stem Cell Res. 2020. V. 48. https://doi.org/10.1016/j.scr.2020.101944
  72. Schoger E., Zimmermann W.H., Cyganek L., Zelarayán L.C. Establishment of two homozygous CRISPR interference (CRISPRi) knock-in human induced pluripotent stem cell (hiPSC) lines for titratable endogenous gene repression // Stem Cell Res. 2021. V. 55. https://doi.org/10.1016/j.scr.2021.102473
  73. Schuster A., Erasimus H., Fritah S. et al. RNAi/CRISPR screens: from a pool to a valid hit // Trends Biotechnol. 2019. V. 37. № 1. https://doi.org/10.1016/j.tibtech.2018.08.002
  74. Bock C., Datlinger P., Chardon F. et al. High-content CRISPR screening // Nat. Rev. Methods Primers. 2022. V. 2. № 1. P. 9. https://doi.org/10.1038/s43586-022-00098-7
  75. Li S., Zhang A., Xue H. et al. One-step piggyBac transposon-based CRISPR/Cas9 activation of multiple genes // Mol. Ther. Nucl. Ac. 2017. V. 8. P. 64–76. https://doi.org/10.1016/j.omtn.2017.06.007
  76. Tian R., Gachechiladze M.A., Ludwig C.H. et al. CRISPR interference-based platform for multimodal genetic screens in human iPSC-derived neurons // Neuron. 2019. V. 104. № 2. P. 239–255.e12. https://doi.org/10.1016/j.neuron.2019.07.014
  77. Friedman C.E., Nguyen Q., Lukowski S.W. et al. Single-cell transcriptomic analysis of cardiac differentiation from human PSCs reveals HOPX-dependent cardiomyocyte maturation // Cell Stem Cell. 2018. V. 23. № 4. P. 586–598.e8. https://doi.org/10.1016/j.stem.2018.09.009
  78. Eskildsen T.V., Ayoubi S., Thomassen M. et al. MESP1 knock-down in human iPSC attenuates early vascular progenitor cell differentiation after completed primitive streak specification // Dev. Biol. 2019. V. 445. № 1. P. 1–7. https://doi.org/10.1016/j.ydbio.2018.10.020
  79. Usluer S., Hallast P., Crepaldi L. et al. Optimized whole-genome CRISPR interference screens identify ARID1A-dependent growth regulators in human induced pluripotent stem cells // Stem Cell Reports. 2023. V. 18. № 5. P. 1061–1074. https://doi.org/10.1016/j.stemcr.2023.03.008
  80. Black J.B., McCutcheon S.R., Dube S. et al. Master regulators and cofactors of human neuronal cell fate specification identified by CRISPR gene activation screens // Cell Rep. 2020. V. 33. № 9. https://doi.org/10.1016/j.celrep.2020.108460
  81. Liu Y., Yu C., Daley T.P. et al. CRISPR activation screens systematically identify factors that drive neuronal fate and reprogramming // Cell Stem Cell. 2018. V. 23. № 5. P. 758–771.e8. https://doi.org/10.1016/j.stem.2018.09.003
  82. Yang J., Rajan S.S., Friedrich M.J. et al. Genome-scale CRISPRa screen identifies novel factors for cellular reprogramming // Stem Cell Reports. 2019. V. 12. № 4. P. 757–771. https://doi.org/10.1016/j.stemcr.2019.02.010
  83. Kakeda M., Nagata K., Osawa K. et al. A new chromosome 14-based human artificial chromosome (HAC) vector system for efficient transgene expression in human primary cells // Biochem. Biophys. Res. Commun. 2011. V. 415. № 3. P. 439–444. https://doi.org/10.1016/j.bbrc.2011.10.088
  84. Farr C.J., Stevanovic M., Thomson E.J. et al. Telomere-associated chromosome fragmentation: Applications in genome manipulation and analysis // Nat. Genet. 1992. V. 2. № 4. P. 275–282. https://doi.org/10.1038/ng1292-275
  85. Mills W., Critcher R., Lee C., Farr C.J. Generation of an ~2.4 Mb human X centromere-based minichromosome by targeted telomere-associated chromosome fragmentation in DT40 // Hum. Mol. Genet. 1999. V. 8. № 5. P. 751–761. https://doi.org/10.1093/HMG/8.5.751
  86. Kuroiwa Y., Tomizuka K., Shinohara T. et al. Manipulation of human minichromosomes to carry greater than megabase-sized chromosome inserts // Nat. Biotechnol. 2000. V. 18. № 10. P. 1086–1090. https://doi.org/10.1038/80287
  87. Shen M.H., Mee P.J., Nichols J. et al. A structurally defined mini-chromosome vector for the mouse germ line // Curr. Biol. 2000. V. 10. № 1. P. 31–34. https://doi.org/10.1016/S0960-9822(99)00261-4
  88. Kazuki Y., Oshimura M. Human artificial chromosomes for gene delivery and the development of animal models // Mol. Ther. 2011. V. 19. № 9. P. 1591–1601. https://doi.org/10.1038/mt.2011.136
  89. Kazuki Y., Hoshiya H., Takiguchi M. et al. Refined human artificial chromosome vectors for gene therapy and animal transgenesis // Gene Ther. 2011. V. 18. № 4. P. 384–393. https://doi.org/10.1038/gt.2010.147
  90. Katoh M., Ayabe F., Norikane S. et al. Construction of a novel human artificial chromosome vector for gene delivery // Biochem. Biophys. Res. Commun. 2004. V. 321. № 2. P. 280–290. https://doi.org/10.1016/j.bbrc.2004.06.145
  91. Hoshiya H., Kazuki Y., Abe S. et al. A highly stable and nonintegrated human artificial chromosome (HAC) containing the 2.4 Mb entire human dystrophin gene // Mol. Ther. 2009. V. 17. № 2. P. 309–317. https://doi.org/10.1038/mt.2008.253
  92. Yamaguchi S., Kazuki Y., Nakayama Y. et al. A method for producing transgenic cells using a multi-integrase system on a human artificial chromosome vector // PLoS One. 2011. V. 6. № 2. https://doi.org/10.1371/journal.pone.0017267
  93. Suda T., Katoh M., Hiratsuka M. et al. Heat-regulated production and secretion of insulin from a human artificial chromosome vector // Biochem. Biophys. Res. Commun. 2006. V. 340. № 4. P. 1053–1061. https://doi.org/10.1016/j.bbrc.2005.12.106
  94. Kakeda M., Hiratsuka M., Nagata K. et al. Human artificial chromosome (HAC) vector provides long-term therapeutic transgene expression in normal human primary fibroblasts // Gene Ther. 2005. V. 12. № 10. P. 852–856. https://doi.org/10.1038/sj.gt.3302483
  95. Yamada H., Kunisato A., Kawahara M. et al. Exogenous gene expression and growth regulation of hematopoietic cells via a novel human artificial chromosome // J. Hum. Genet. 2006. V. 51. № 2. P. 147–150. https://doi.org/10.1007/s10038-005-0334-9
  96. Kazuki Y., Hiratsuka M., Takiguchi M. et al. Complete genetic correction of iPS cells from Duchenne muscular dystrophy // Mol. Ther. 2010. V. 18. № 2. P. 386–393. https://doi.org/10.1038/mt.2009.274
  97. Sinenko S.A., Ponomartsev S.V., Tomilin A.N. Human artificial chromosomes for pluripotent stem cell-based tissue replacement therapy // Exp. Cell Res. 2020. V. 389. № 1. https://doi.org/10.1016/j.yexcr.2020.111882
  98. Benedetti S., Uno N., Hoshiya H. et al. Reversible immortalisation enables genetic correction of human muscle progenitors and engineering of next‐generation human artificial chromosomes for Duchenne muscular dystrophy // EMBO Mol. Med. 2018. V. 10. № 2. P. 254–275. https://doi.org/10.15252/emmm.201607284
  99. Kurosaki H., Hiratsuka M., Imaoka N. et al. Integration-free and stable expression of FVIII using a human artificial chromosome // J. Hum. Genet. 2011. V. 56. № 10. P. 727–733. https://doi.org/10.1038/jhg.2011.88.
  100. Wang Y., Kazuki K., Hichiwa G. et al. Human artificial chromosome carrying R-spondin1 and IL-22 expression cassettes in rejuvenated MSCs enhances therapeutic efficacy in ulcerative colitis model mice // Biomed. Pharmacother. 2025. V. 182. https://doi.org/10.1016/J.BIOPHA.2024.117751
  101. Hiramuki Y., Hosokawa M., Osawa K. et al. Titin fragment is a sensitive biomarker in Duchenne muscular dystrophy model mice carrying full-length human dystrophin gene on human artificial chromosome // Sci. Rep. 2025. V. 15. № 1. P. 1–9. https://doi.org/10.1038/s41598-025-85369-5
  102. Watanabe M., Miyamoto H., Okamoto K. et al. Phenotypic features of dystrophin gene knockout pigs harboring a human artificial chromosome containing the entire dystrophin gene // Mol. Ther. Nucl. A. 2023. V. 33. P. 444–453. https://doi.org/10.1016/j.omtn.2023.07.021
  103. Harrington J.J., Van Bokkelen G., Mays R.W. et al. Formation of de novo centromeres and construction of first-generation human artificial microchromosomes // Nat. Genet. 1997. V. 15. № 4. P. 345–355. https://doi.org/10.1038/ng0497-345
  104. Ikeno M., Grimes B., Okazaki T. et al. Construction of YAC-based mammalian artificial chromosomes // Nat. Biotechnol. 1998. V. 16. № 5. P. 431–439. https://doi.org/10.1038/nbt0598-431
  105. Guiducci C., Ascenzioni F., Auriche C. et al. Use of a human minichromosome as a cloning and expression vector for mammalian cells // Hum. Mol. Genet. 1999. V. 8. № 8. https://doi.org/10.1093/hmg/8.8.1417
  106. Ebersole T.A., Ross A., Clark E. et al. Mammalian artificial chromosome formation from circular alphoid input DNA does not require telomere repeats // Hum. Mol. Genet. 2000. V. 9. № 11. https://doi.org/10.1093/hmg/9.11.1623
  107. Mejía J.E., Alazami A., Willmott A. et al. Efficiency of de novo centromere formation in human artificial chromosomes // Genomics. 2002. V. 79. № 3. P. 297–304. https://doi.org/10.1006/geno.2002.6704
  108. Kouprina N., Ebersole T., Pak E. et al. Cloning of human centromeres by transformation-associated recombination in yeast and generation of functional human artificial chromosomes // Nucl. Ac. Res. 2003. V. 31. № 3. P. 922–934. https://doi.org/10.1093/nar/gkg182
  109. Basu J., Stromberg G., Compitello G. et al. Rapid creation of BAC-based human artificial chromosome vectors by transposition with synthetic alpha-satellite arrays // Nucl. Ac. Res. 2005. V. 33. № 2. P. 587–596. https://doi.org/10.1093/nar/gki207
  110. Moralli D., Simpson K.M., Wade-Martins R., Monaco Z.L. A novel human artificial chromosome gene expression system using herpes simplex virus type 1 vectors // EMBO Rep. 2006. V. 7. № 9. P. 911–918. https://doi.org/10.1038/sj.embor.7400768
  111. Ohzeki J.I., Bergmann J.H., Kouprina N. et al. Breaking the HAC barrier: Histone H3K9 acetyl/methyl balance regulates CENP-A assembly // EMBO J. 2012. V. 31. № 10. P. 2391–2402. https://doi.org/10.1038/emboj.2012.82
  112. Iida Y., Kim J.H., Kazuki Y. et al. Human artificial chromosome with a conditional centromere for gene delivery and gene expression // DNA Res. 2010. V. 17. № 5. P. 293–301. https://doi.org/10.1093/dnares/dsq020
  113. Liskovykh M., Lee N.C., Larionov V., Kouprina N. Moving toward a higher efficiency of microcell-mediated chromosome transfer // Mol. Ther. Methods Clin. Dev. 2016. V. 3. https://doi.org/10.1038/mtm.2016.43
  114. Ponomartsev S.V., Sinenko S.A., Tomilin A.N. Human artificial chromosomes and their transfer to target cells // Acta Naturae. 2022. V. 14. № 3. P. 35–45. https://doi.org/10.32607/actanaturae.11670
  115. Uno N., Miyamoto H., Yamazaki K. et al. Microcell-mediated chromosome transfer between non-identical human iPSCs // Mol. Ther. Nucl. Ac. 2024. V. 35. № 4. https://doi.org/10.1016/j.omtn.2024.102382
  116. Suzuki T., Kazuki Y., Hara T., Oshimura M. Current advances in microcell-mediated chromosome transfer technology and its applications // Exp. Cell Res. 2020. V. 390. № 1. https://doi.org/10.1016/j.yexcr.2020.111915
  117. Nakano M., Cardinale S., Noskov V.N. et al. Inactivation of a Human Kinetochore by Specific Targeting of Chromatin Modifiers // Dev. Cell. 2008. V. 14. № 4. P. 507–522. https://doi.org/10.1016/j.devcel.2008.02.001
  118. Kouprina N., Samoshkin A., Erliandri I. et al. Organization of synthetic alphoid DNA array in human artificial chromosome (HAC) with a conditional centromere // ACS Synth. Biol. 2012. V. 1. № 12. P. 590–601. https://doi.org/10.1021/sb3000436
  119. Ebersole T., Okamoto Y., Noskov V.N. et al. Rapid generation of long synthetic tandem repeats and its application for analysis in human artificial chromosome formation // Nucl. Ac. Res. 2005. V. 33. № 15. e130. https://doi.org/10.1093/nar/gni129
  120. Breman A.M., Steiner C.M., Slee R.B., Grimes B.R. Input DNA ratio determines copy number of the 33 kb factor IX gene on de novo human artificial chromosomes // Mol. Therapy. 2008. V. 16. № 2. P. 315–323. https://doi.org/10.1038/sj.mt.6300361
  121. Alazami A.M., Mejía J.E., Monaco Z.L. Human artificial chromosomes containing chromosome 17 alphoid DNA maintain an active centromere in murine cells but are not stable // Genomics. 2004. V. 83. № 5. P. 844–851. https://doi.org/10.1016/j.ygeno.2003.11.011
  122. Moralli D., Chan D.Y.L., Jefferson A. et al. HAC stability in murine cells is influenced by nuclear localization and chromatin organization // BMC Cell Biol. 2009. V. 10. https://doi.org/10.1186/1471-2121-10-18
  123. Kim J.H., Kononenko A., Erliandri I. et al. Human artificial chromosome (HAC) vector with a conditional centromere for correction of genetic deficiencies in human cells // PNAS USA. 2011. V. 108. № 50. P. 20048–20053. https://doi.org/10.1073/pnas.1114483108
  124. Ponomartsev S.V., Sinenko S.A., Skvortsova E.V. et al. Human AlphoidtetO artificial chromosome as a gene therapy vector for the developing hemophilia a model in mice // Cells. 2020. V. 9. № 4. https://doi.org/10.3390/cells9040879
  125. Liskovykh M., Ponomartsev S., Popova E. et al. Stable maintenance of de novo assembled human artificial chromosomes in embryonic stem cells and their differentiated progeny in mice // Cell Cycle. 2015. V. 14. № 8. P. 1268–1273. https://doi.org/10.1080/15384101.2015.1014151
  126. Sinenko S.A., Skvortsova E.V., Liskovykh M.A. et al. Transfer of synthetic human chromosome into human induced pluripotent stem cells for biomedical applications // Cells. 2018. V. 7. № 12. https://doi.org/10.3390/cells7120261
  127. Ikeno M., Hasegawa Y. Applications of bottom-up human artificial chromosomes in cell research and cell engineering // Exp. Cell Res. 2020. V. 390. № 1. https://doi.org/10.1016/j.yexcr.2019.111793
  128. Sinenko S.A., Ponomartsev S.V., Tomilin A.N. Pluripotent stem cell-based gene therapy approach: human de novo synthesized chromosomes // Cell. Mol. Life Sci. 2021. V. 78. № 4. P. 1207–1220. https://doi.org/10.1007/s00018-020-03653-1
  129. Gambogi C.W., Birchak G.J., Mer E. et al. Efficient formation of single-copy human artificial chromosomes // Science. 2024. V. 383. № 6689. P. 1344–1349. https://doi.org/10.1126/science.adj3566
  130. Logsdon G.A., Gambogi C.W., Liskovykh M.A. et al. Human artificial chromosomes that bypass centromeric DNA // Cell. 2019. V. 178. № 3. P. 624–639. e19. https://doi.org/10.1016/j.cell.2019.06.006
  131. Miyamoto H., Kobayashi H., Kishima N. et al. Rapid human genomic DNA cloning into mouse artificial chromosome via direct chromosome transfer from human iPSC and CRISPR/Cas9-mediated translocation // Nucl. Ac. Res. 2024. V. 52. № 3. P. 1498–1511. https://doi.org/10.1093/nar/gkad1218
  132. Boyer L.A., Tong I.L., Cole M.F. et al. Core transcriptional regulatory circuitry in human embryonic stem cells // Cell. 2005. V. 122. № 6. P. 947–956. https://doi.org/10.1016/j.cell.2005.08.020
  133. Young R.A. Control of the embryonic stem cell state // Cell. 2011. V. 144. № 6. P. 940–954. https://doi.org/10.1016/j.cell.2011.01.032
  134. Avilion A.A., Nicolis S.K., Pevny L.H. et al. Multipotent cell lineages in early mouse development depend on SOX2 function // Gen. Dev. 2003. V. 17. № 1. P. 126–140. https://doi.org/10.1101/gad.224503
  135. Nichols J., Zevnik B., Anastassiadis K. et al. Formation of pluripotent stem cells in the mammalian embryo depends on the POU transcription factor Oct4 // Cell. 1998. V. 95. № 3. P. 379–391. https://doi.org/10.1016/S0092-8674(00)81769-9
  136. Mitsui K., Tokuzawa Y., Itoh H. et al. The homeoprotein Nanog is required for maintenance of pluripotency in mouse epiblast and ES cells // Cell. 2003. V. 113. № 5. P. 631–642. https://doi.org/10.1016/S0092-8674(03)00393-3
  137. Chambers I., Colby D., Robertson M. et al. Functional expression cloning of Nanog, a pluripotency sustaining factor in embryonic stem cells // Cell. 2003. V. 113. № 5. P. 643–655. https://doi.org/10.1016/S0092-8674(03)00392-1
  138. Chambers I., Silva J., Colby D. et al. Nanog safeguards pluripotency and mediates germline development // Nature. 2007. V. 450. № 7173. P. 1230–1234. https://doi.org/10.1038/nature06403
  139. Niwa H., Miyazaki J.-ichi, Smith A.G. Quantitative expression of Oct-3/4 defines differentiation, dedifferentiation or self-renewal of ES cells // Nat. Genet. 2000. V. 24. № 4. P. 372–376. https://doi.org/10.1038/74199
  140. Radzisheuskaya A., Le Bin Chia G., Dos Santos R.L. et al. A defined Oct4 level governs cell state transitions of pluripotency entry and differentiation into all embryonic lineages // Nat. Cell Biol. 2013. V. 15. № 6. P. 579–590. https://doi.org/10.1038/ncb2742
  141. Zeineddine D., Papadimou E., Chebli K. et al. Oct- 3/4 dose dependently regulates specification of embryonic stem cells toward a cardiac lineage and early heart development // Dev. Cell. 2006. V. 11. № 4. P. 535–546. https://doi.org/10.1016/j.devcel.2006.07.013
  142. Wang Z., Oron E., Nelson B. et al. Distinct lineage specification roles for NANOG, OCT4, and SOX2 in human embryonic stem cells // Cell Stem Cell. 2012. V. 10. № 4. P. 440–454. https://doi.org/10.1016/j.stem.2012.02.016
  143. Yu Y., Wang X., Zhang X. et al. ERK inhibition promotes neuroectodermal precursor commitment by blocking self-renewal and primitive streak formation of the epiblast // Stem Cell Res. Ther. 2018. V. 9. № 1. P. 2. https://doi.org/10.1186/s13287-017-0750-8
  144. DeVeale B., Brokhman I., Mohseni P. et al. Oct4 is required ~E7.5 for proliferation in the primitive streak // PLoS Genet. 2013. V. 9. № 11. https://doi.org/10.1371/journal.pgen.1003957
  145. Kehler J., Tolkunova E., Koschorz B. et al. Oct4 is required for primordial germ cell survival // EMBO Rep. 2004. V. 5. № 11. P. 1078–1083. https://doi.org/10.1038/sj.embor.7400279
  146. Mulas C., Chia G., Jones K.A. et al. Oct4 regulates the embryonic axis and coordinates exit from pluripotency and germ layer specification in the mouse embryo // Develop. (Cambridge). 2018. V. 145. № 12. https://doi.org/10.1242/dev.159103
  147. Masui S., Nakatake Y., Toyooka Y. et al. Pluripotency governed by Sox2 via regulation of Oct3/4 expression in mouse embryonic stem cells // Nat. Cell Biol. 2007. V. 9. № 6. P. 625–635. https://doi.org/10.1038/ncb1589
  148. Kopp J.L., Ormsbee B.D., Desler M., Rizzino A. Small increases in the level of Sox2 trigger the differentiation of mouse embryonic stem cells // Stem Cells. 2008. V. 26. № 4. P. 903–911. https://doi.org/10.1634/stemcells.2007-0951
  149. Tsai J.M., Nowak R.P., Ebert B.L., Fischer E.S. Targeted protein degradation: from mechanisms to clinic // Nat. Rev. Mol. Cell Biol. 2024. V. 25. № 9. P. 740–757. https://doi.org/10.1038/s41580-024-00729-9
  150. Nishimura K., Fukagawa T., Takisawa H. et al. An auxin-based degron system for the rapid depletion of proteins in nonplant cells // Nat. Methods. 2009. V. 6. № 12. P. 917–922. https://doi.org/10.1038/nmeth.1401
  151. Nabet B., Roberts J.M., Buckley D.L. et al. The dTAG system for immediate and target-specific protein degradation // Nat. Chem. Biol. 2018. V. 14. № 5. P. 431–441. https://doi.org/10.1038/s41589-018-0021-8
  152. Li S., Prasanna X., Salo V.T. et al. An efficient auxin-inducible degron system with low basal degradation in human cells // Nat. Methods. 2019. V. 16. № 9. P. 866–869. https://doi.org/10.1038/s41592-019-0512-x
  153. Bates L.E., Alves M.R.P., Silva J.C.R. Auxin-degron system identifies immediate mechanisms of OCT4 // Stem Cell Reports. 2021. V. 16. № 7. P. 1818–1831. https://doi.org/10.1016/j.stemcr.2021.05.016
  154. Yesbolatova A., Saito Y., Kitamoto N. et al. The auxin-inducible degron 2 technology provides sharp degradation control in yeast, mammalian cells, and mice // Nat. Commun. 2020. V. 11. № 1. P. 5701. https://doi.org/10.1038/s41467-020-19532-z
  155. Nabet B., Ferguson F.M., Seong B.K.A. et al. Rapid and direct control of target protein levels with VHL-recruiting dTAG molecules // Nat. Commun. 2020. V. 11. № 1. P. 4687. https://doi.org/10.1038/s41467-020-18377-w
  156. Liu N.Q., Maresca M., van den Brand T. et al. WAPL maintains a cohesin loading cycle to preserve cell-type-specific distal gene regulation // Nat. Genet. 2021. V. 53. № 1. P. 100–109. https://doi.org/10.1038/s41588-020-00744-4
  157. Maresca M., van den Brand T., Li H. et al. Pioneer activity distinguishes activating from non‐activating SOX2 binding sites // EMBO J. 2023. V. 42. № 20. https://doi.org/10.15252/embj.2022113150
  158. Abuhashem A., Lee A.S., Joyner A.L., Hadjantonakis A.K. Rapid and efficient degradation of endogenous proteins in vivo identifies stage-specific roles of RNA Pol II pausing in mammalian development // Dev. Cell. 2022. V. 57. № 8. P. 1068–1080.e6. https://doi.org/10.1016/j.devcel.2022.03.013
  159. Reményi A., Lins K., Nissen L.J. et al. Crystal structure of a POU/HMG/DNA ternary complex suggests differential assembly of Oct4 and Sox2 on two enhancers // Genes Dev. 2003. V. 17. № 16. P. 2048–2059. https://doi.org/10.1101/gad.269303
  160. Williams D.C., Cai M., Clore G.M. Molecular basis for synergistic transcriptional activation by Oct1 and Sox2 revealed from the solution structure of the 42- kDa Oct1.·Sox2.·Hoxb1-DNA ternary transcription factor complex // J. Biol. Chem. 2004. V. 279. № 2. P. 1449–1457. https://doi.org/10.1074/jbc.M309790200
  161. Jerabek S., Ng C.K., Wu G. et al. Changing POU dimerization preferences converts Oct6 into a pluripotency inducer // EMBO Rep. 2017. V. 18. № 2. P. 319–333. https://doi.org/10.15252/embr.201642958
  162. Malik V., Glaser L.V., Zimmer D. et al. Pluripotency reprogramming by competent and incompetent POU factors uncovers temporal dependency for Oct4 and Sox2 // Nat. Commun. 2019. V. 10. № 1. P. 3477. https://doi.org/10.1038/s41467-019-11054-7
  163. Bakhmet E.I., Tomilin A.N. Key features of the POU transcription factor Oct4 from an evolutionary perspective // Cell. Mol. Life Sci. 2021. V. 78. № 23. P. 7339–7353. https://doi.org/10.1007/s00018-021-03975-8
  164. Pan X., Cang X., Dan S. et al. Site-specific disruption of the Oct4/Sox2 protein interaction reveals coordinated mesendodermal differentiation and the epithelial-mesenchymal transition // J. Biol. Chem. 2016. V. 291. № 35. P. 18353–18369. https://doi.org/10.1074/jbc.M116.745414
  165. Esch D., Vahokoski J., Groves M.R. et al. A unique Oct4 interface is crucial for reprogramming to pluripotency // Nat. Cell Biol. 2013. V. 15. № 3. P. 295–301. https://doi.org/10.1038/ncb2680
  166. MacCarthy C.M., Wu G., Malik V. et al. Highly cooperative chimeric super-SOX induces naive pluripotency across species // Cell Stem Cell. 2024. V. 31. № 1. P. 127–147.e9. https://doi.org/10.1016/j.stem.2023.11.010
  167. Han D., Wu G., Chen R. et al. A balanced Oct4 interactome is crucial for maintaining pluripotency // Sci. Adv. 2022. V. 8. № 7. https://doi.org/10.1126/sciadv.abe4375
  168. Tan D.S., Chen Y., Gao Y. et al. Directed evolution of an enhanced POU reprogramming factor for cell fate engineering // Mol. Biol. Evol. 2021. V. 38. № 7. P. 2854–2868. https://doi.org/10.1093/molbev/msab075
  169. Jauch R., Aksoy I., Hutchins A.P. et al. Conversion of Sox17 into a pluripotency reprogramming factor by reengineering its association with Oct4 on DNA // Stem Cells. 2011. V. 29. № 6. P. 940–951. https://doi.org/10.1002/stem.639
  170. Aksoy I., Jauch R., Chen J. et al. Oct4 switches partnering from Sox2 to Sox17 to reinterpret the enhancer code and specify endoderm // EMBO J. 2013. V. 32. № 7. P. 938–953. https://doi.org/10.1038/emboj.2013.31
  171. Veerapandian V., Ackermann J.O., Srivastava Y. et al. Directed evolution of reprogramming factors by cell selection and sequencing // Stem Cell Reports. 2018. V. 11. № 2. P. 593–606. https://doi.org/10.1016/j.stemcr.2018.07.002
  172. Hu H., Ho D.H.H., Tan D.S. et al. Evaluation of the determinants for improved pluripotency induction and maintenance by engineered SOX17 // Nucl. Ac. Res. 2023. V. 51. № 17. P. 8934–8956. https://doi.org/10.1093/nar/gkad597
  173. Ho S.Y., Hu H., Ho D.H.H. et al. An acidic residue within the OCT4 dimerization interface of SOX17 is necessary and sufficient to overcome its pluripotency-inducing activity // Stem Cell Reports. 2025. V. 20. № 3. https://doi.org/10.1016/J.STEMCR.2025.102398

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