Modulation of Insulin Gene Expression with CRISPR/Cas9-based Transcription Factors

Authors

  • Bakhytzhan Alzhanuly Aitkhozhin Institute of Molecular Biology and Biochemistry, Almaty, Kazakhstan; Department of Molecular Biology and Genetics, Faculty of Biology and Biotechnology, Al-Farabi Kazakh National University, Almaty, Kazakhstan; Department of Research and Development, Almaty Management University, Almaty, Kazakhstan
  • Zhussipbek Y. Mukhatayev Aitkhozhin Institute of Molecular Biology and Biochemistry, Almaty, Kazakhstan; Department of Molecular Biology and Genetics, Faculty of Biology and Biotechnology, Al-Farabi Kazakh National University, Almaty, Kazakhstan
  • Dauren M. Botbayev Aitkhozhin Institute of Molecular Biology and Biochemistry, Almaty, Kazakhstan; Department of Molecular Biology and Genetics, Faculty of Biology and Biotechnology, Al-Farabi Kazakh National University, Almaty, Kazakhstan
  • Yeldar Ashirbekov Aitkhozhin Institute of Molecular Biology and Biochemistry, Almaty, Kazakhstan
  • Nurlybek D. Katkenov Zhangir Khan University, Uralsk, Kazakhstan
  • Nurlan T. Dzhaynakbaev Kazakh-Russian Medical University, Almaty, Kazakhstan
  • Kamalidin O. Sharipov Aitkhozhin Institute of Molecular Biology and Biochemistry, Almaty, Kazakhstan

DOI:

https://doi.org/10.3889/oamjms.2021.6980

Keywords:

CRISPR/dCas9 system, Diabetes type I, Embryonic stem cells, HEK293 cells

Abstract

Background: The discovery and use of CRISPR/Cas9 technology have enabled researchers throughout the globe to continuously edit genomes for the benefit of science and medicine. Diabetes type I is one field of medicine where CRISPR/Cas9 has a strong potential for cell therapy development. The long-lasting paucity of healthy cells for clinical transplantation into diabetic patients has led to the search of new methods for producing β-cells from other human cell types. Embryonic stem cells are being studied worldwide as one most promising solution of this need. Aim: The aim of the study is to to check the feasibility of modulating human insulin transcription using CRISPR/Cas9-based synthetic transcription regulation factors.

Results: A new approach for creating potential therapeutic donor cells with enhanced and suppressed insulin production based on one of the latest achievements of human genome editing was developed. Both synthetic transcription activator (VP64) and transcription repressor (KRAB) proteins were shown to function adequately well as a part of the whole CRISPR/Cas9-based system. We claim that our results have a lot to offer and can bring light to many studies where numerous labs are struggling on to treat this disease.

Downloads

Download data is not yet available.

Metrics

Metrics Loading ...

Plum Analytics Artifact Widget Block

References

Hussey BJ, McMillen DR. Programmable T7-based synthetic transcription factors. Nucleic Acids Res. 2018;46(18):9842-54. http://doi.org/10.1093/nar/gky785 PMid:30169636 DOI: https://doi.org/10.1093/nar/gky785

Beltran AS, Russo A, Lara H, Fan C, Lizardi PM, Blancafort P. Suppression of breast tumor growth and metastasis by an engineered transcription factor. PLoS One. 2011;6(9):e24595. http://doi.org/10.1371/journal.pone.0024595 PMid:21931769 DOI: https://doi.org/10.1371/journal.pone.0024595

Wada T, Wallerich S, Becskei A. Synthetic transcription factors switch from local to long-range control during cell differentiation. ACS Synth Biol. 2019;8(2):223-31. https://doi.org/10.1021/acssynbio.8b00369 DOI: https://doi.org/10.1021/acssynbio.8b00369

Rebar EJ, Huang Y, Hickey R, Nath AK, Meoli D, Nath S, et al. Induction of angiogenesis in a mouse model using engineered transcription factors. Nat Med. 2002;8(12):1427-32. https://doi.org/10.1038/nm1202-795 PMid:12415262 DOI: https://doi.org/10.1038/nm1202-795

Zhang F, Cong L, Lodato S, Kosuri S, Church GM, Arlotta P. Efficient construction of sequence-specific TAL effectors for modulating mammalian transcription. Nat Biotechnol. 2011;29(2):149-53. DOI: https://doi.org/10.1038/nbt.1775

Beerli RR, Barbas CF 3rd. Engineering polydactyl zinc-finger transcription factors. Nat Biotechnol. 2002;20(2):135-41. https://doi.org/10.1038/nbt0202-135 PMid:11821858 DOI: https://doi.org/10.1038/nbt0202-135

Jinek M, Chylinski K, Fonfara I, Hauer M, Doudna JA, Charpentier E. A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity. Science. 2012;337(6096):816-21. https://doi.org/10.1126/science.1225829 PMid:22745249 DOI: https://doi.org/10.1126/science.1225829

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):823-6. https://doi.org/10.1126/science.1232033 PMid:23287722 DOI: https://doi.org/10.1126/science.1232033

Matjusaitis M, Wagstaff LJ, Martella A, Baranowski B, Blin C, Gogolok S, et al. Reprogramming of fibroblasts to oligodendrocyte progenitor-like cells using CRISPR/Cas9-based synthetic transcription factors. Stem Cell Rep. 2019;13(6):1053-67. https://doi.org/10.1016/j.stemcr.2019.10.010 PMid:31708478 DOI: https://doi.org/10.1016/j.stemcr.2019.10.010

Mol M, Kabra R, Singh S. Genome modularity and synthetic biology: Engineering systems. Prog Biophys Mol Biol. 2018;132:43-51. https://doi.org/10.1016/j.pbiomolbio.2017.08.002 PMid:28801037 DOI: https://doi.org/10.1016/j.pbiomolbio.2017.08.002

Pepper AR, Gala-Lopez B, Ziff O, Shapiro AJ. Current status of clinical islet transplantation. World J Transplant. 2013;3(4):48-53. https://doi.org/10.5500/wjt.v3.i4.48 PMid:24392308 DOI: https://doi.org/10.5500/wjt.v3.i4.48

Dinnyes A, Schnur A, Muenthaisong S, Bartenstein P, Burcez CT, Burton N, et al. Integration of nano-and biotechnology for beta-cell and islet transplantation in Type-1 diabetes treatment. Cell Prolif. 2020;53(5):e12785. https://doi.org/10.1111/cpr.12785 DOI: https://doi.org/10.1111/cpr.12785

Kroon E, Martinson LA, Kadoya K, Bang AG, Kelly OG, Eliazer S, et al. Pancreatic endoderm derived from human embryonic stem cells generates glucose-responsive insulin-secreting cells in vivo. Nat Biotechnol. 2008;26(4):443-52. https://doi.org/10.1038/nbt1393 PMid:18288110 DOI: https://doi.org/10.1038/nbt1393

Pagliuca FW, Millman JR, Gurtler M, Segel M, Van Dervort A, Ryu JH, et al. Generation of functional human pancreatic beta cells in vitro. Cell 2014;159(2):428-39. https://doi.org/10.1016/j.cell.2014.09.040 PMid:25303535 DOI: https://doi.org/10.1016/j.cell.2014.09.040

Nostro MC, Sarangi F, Yang C, Holland A, Elefanty AG, Stanley EG, et al. Efficient generation of NKX6-1+ pancreatic progenitors from multiple human pluripotent stem cell lines. Stem Cell Reports. 2015;4(4):591-604. https://doi.org/10.1016/j.stemcr.2015.02.017 PMid:25843049 DOI: https://doi.org/10.1016/j.stemcr.2015.02.017

Millman JR, Xie C, Van Dervort A, Gürtler M, Pagliuca FW, Melton DA. Generation of stem cell-derived beta-cells from patients with Type 1 diabetes. Nat Commun. 2016;7:11463. https://doi.org/10.1038/ncomms11463 PMid:27163171 DOI: https://doi.org/10.1038/ncomms11463

Poitout V, Hagman D, Stein R, Artner I, Robertson RP, Harmon JS. Regulation of the insulin gene by glucose and fatty acids. J Nutr. 2006;136(4):873-6. https://doi.org/10.1093/jn/136.4.873 PMid:16549443 DOI: https://doi.org/10.1093/jn/136.4.873

Kuroda A, Rauch TA, Todorov I, Ku HT, Al-Abdullah IH, Kandeel F, et al. Insulin gene expression is regulated by DNA methylation. PLoS One. 2009;4(9):e6953. https://doi.org/10.1371/journal.pone.0006953 PMid:19742322 DOI: https://doi.org/10.1371/journal.pone.0006953

Melloul D, Marshak S, Cerasi E. Regulation of insulin gene transcription. Diabetologia. 2002;45(3):309-26. https://doi.org/10.1007/s00125-001-0728-y PMid:11914736 DOI: https://doi.org/10.1007/s00125-001-0728-y

Maeder ML, Linder SJ, Reyon D, Angstman JF, Fu Y, Sander JD, et al. Robust, synergistic regulation of human gene expression using TALE activators. Nat Methods. 2013;10(3):243-5. https://doi.org/10.1038/nmeth.2366 PMid:23396285 DOI: https://doi.org/10.1038/nmeth.2366

Yin H, Song CQ, Suresh S, Kwan SY, Wu Q, Walsh S, et al. Partial DNA-guided Cas9 enables genome editing with reduced off-target activity. Nat Chem Biol. 2018;14(3):311-6. https://doi.org/10.1038/nchembio.2559 PMid:29377001 DOI: https://doi.org/10.1038/nchembio.2559

Mali P, Esvelt KM, Church GM. Cas9 as a versatile tool for engineering biology. Nat Methods. 2013;10(10):957-63. https://doi.org/10.1038/nmeth.2649 PMid:24076990 DOI: https://doi.org/10.1038/nmeth.2649

Perez-Pinera P, Kocak DD, Vockley CM, Adler AF, Kabadi AM, Polstein LR, et al. RNA-guided gene activation by CRISPR-Cas9- based transcription factors. Nat Methods. 2013;10(10):973-6. https://doi.org/10.1038/nmeth.2600 PMid:23892895 DOI: https://doi.org/10.1038/nmeth.2600

Doench JG, Fusi N, Sullender M, Hegde M, Vaimberg EW, Donovan KF, et al. Optimized sgRNA design to maximize activity and minimize off-target effects of CRISPR-Cas9. Nat Biotechnol. 2016;34(2):184-91. https://doi.org/10.1038/nbt.3437 PMid:26780180 DOI: https://doi.org/10.1038/nbt.3437

Guschin DY, Waite AJ, Katibah GE, Miller JC, Holmes MC, Rebar EJ. A rapid and general assay for monitoring endogenous gene modification. Methods Mol Biol. 2010;649:247-56. https://doi.org/10.1007/978-1-60761-753-2_15 PMid:20680839 DOI: https://doi.org/10.1007/978-1-60761-753-2_15

Kabadi AM, Ousterout DG, Hilton IB, Gersbach CA. Multiplex CRISPR/Cas9-based genome engineering from a single lentiviral vector. Nucleic Acids Res. 2014;42(19):e147. https://doi.org/10.1093/nar/gku749 PMid:25122746 DOI: https://doi.org/10.1093/nar/gku749

Yeo NC, Chavez A, Lance-Byrne A, Chan Y, Menn D, Milanova D, et al. An enhanced CRISPR repressor for targeted mammalian gene regulation. Nat Methods. 2018;15(8):611-6. https://doi.org/10.1038/s41592-018-0048-5 PMid:30013045 DOI: https://doi.org/10.1038/s41592-018-0048-5

Wang X, McManus M. Lentivirus production. J Vis Exp. 2009;32:1499. https://doi.org/10.3791/1499 PMid:19801965 DOI: https://doi.org/10.3791/1499

Rosenbluh J, Xu H, Harrington W, Gill S, Wang X, Vazquez F, et al. Complementary information derived from CRISPR Cas9 mediated gene deletion and suppression. Nat Commun. 2017;8:15403. https://doi.org/10.1038/ncomms15403 PMid:28534478 DOI: https://doi.org/10.1038/ncomms15403

Song B, Scheuner D, Ron D, Pennathur S, Kaufman RJ. Chop deletion reduces oxidative stress, improves beta cell function, and promotes cell survival in multiple mouse models of diabetes. J Clin Invest. 2008;118(10):3378-89. https://doi.org/10.1172/JCI34587 PMid:18776938 DOI: https://doi.org/10.1172/JCI34587

Kaufman RJ, Back SH, Song B, Han J, Hassler J. The unfolded protein response is required to maintain the integrity of the endoplasmic reticulum, prevent oxidative stress and preserve differentiation in beta-cells. Diabetes Obes Metab. 2010;12 Suppl 2:99-107. https://doi.org/10.1111/j.1463-1326.2010.01281.x PMid:21029306 DOI: https://doi.org/10.1111/j.1463-1326.2010.01281.x

Gilbert LA, Larson MH, Morsut L, Liu Z, Brar GA, Torres SE, et al. CRISPR-mediated modular RNA-guided regulation of transcription in eukaryotes. Cell. 2013;154(2):442-51. https://doi.org/10.1016/j.cell.2013.06.044 PMid:23849981 DOI: https://doi.org/10.1016/j.cell.2013.06.044

Hendel A, Fine EJ, Bao G, Porteus MH. Quantifying on- and off-target genome editing. Trends Biotechnol. 2015;33(2):132-40. https://doi.org/10.1016/j.tibtech.2014.12.001 PMid:25595557 DOI: https://doi.org/10.1016/j.tibtech.2014.12.001

Downloads

Published

2021-10-10

How to Cite

1.
Alzhanuly B, Mukhatayev ZY, Botbayev DM, Ashirbekov Y, Katkenov ND, Dzhaynakbaev NT, Sharipov KO. Modulation of Insulin Gene Expression with CRISPR/Cas9-based Transcription Factors. Open Access Maced J Med Sci [Internet]. 2021 Oct. 10 [cited 2022 May 21];9(A):876-81. Available from: https://oamjms.eu/index.php/mjms/article/view/6980