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Genome editing of the HIV co-receptors CCR5 and CXCR4 by CRISPR-Cas9 protects CD4+ T cells from HIV-1 infection

Liu, Zhepeng, Chen, Shuliang, Jin, Xu, Wang, Qiankun, Yang, Kongxiang, Li, Chenlin, Xiao, Qiaoqiao, Hou, Panpan, Liu, Shuai, Wu, Shaoshuai, Hou, Wei, Xiong, Yong, Kong, Chunyan, Zhao, Xixian, Wu, Li, Li, Chunmei, Sun, Guihong, Guo, Deyin·2017·Cell & Bioscience
DOI 10.1186/s13578-017-0174-2PMC PMC5591563
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Fig. 1 Schematic diagram of sgRNA of CXCR4 and CCR5 targets and vector construction. a Schematic of the CXCR4 and CCR5 coding region in genomic DNA sequences targeted by lenti-X4R5-Cas9-#1,#2. b Structure of lenti-X4R5-Cas9-#1,#2 vectors expressing Cas9 and dual sgRNA. c gRNA sequences used in lenti-X4R5-Cas9-#1,#2 vectors
Fig. 1 Schematic diagram of sgRNA of CXCR4 and CCR5 targets and vector construction. a Schematic of the CXCR4 and CCR5 coding region in genomic DNA sequences targeted by lenti-X4R5-Cas9-#1,#2. b Structure of lenti-X4R5-Cas9-#1,#2 vectors expressing Cas9 and dual sgRNA. c gRNA sequences used in lenti-X4R5-Cas9-#1,#2 vectorsCC BY · PubMed Central
Fig. 2 Disruption of CXCR4 and CCR5 protects TZM-bl cells from HIV-1 infection. a T7E1 assay for genome level cleavage efficacy by lenti-X4R5-Cas9-#1,#2 in TZM-bl. b Expression of CXCR4 or CCR5 in TZM-bl cell line transfected with lenti-X4R5-Cas9 by lipo2000 transfection reagent were analyzed with flow cytometry. c On-target analysis of the cleavage on target sites. d lenti-X4R5-Cas9 transfected TZM-bl cell line challenged with HIV-1NL4-3 or HIV-1YU-2 (3 days post infection). The data shown were the mean ± SD of three independent experiments. *P < 0.05; **P < 0.01; ***P < 0.001; t test
Fig. 2 Disruption of CXCR4 and CCR5 protects TZM-bl cells from HIV-1 infection. a T7E1 assay for genome level cleavage efficacy by lenti-X4R5-Cas9-#1,#2 in TZM-bl. b Expression of CXCR4 or CCR5 in TZM-bl cell line transfected with lenti-X4R5-Cas9 by lipo2000 transfection reagent were analyzed with flow cytometry. c On-target analysis of the cleavage on target sites. d lenti-X4R5-Cas9 transfected TZM-bl cell line challenged with HIV-1NL4-3 or HIV-1YU-2 (3 days post infection). The data shown were the mean ± SD of three independent experiments. *P < 0.05; **P < 0.01; ***P < 0.001; t testCC BY · PubMed Central
Fig. 3 Jurkat T cell line modified by X4R5-Cas9 lentivirus antagonized HIV infection. a T7E1 assay identification of packaged X4R5-Cas9 lentivirus mediated cleavage at the genome level. b On-target analysis of each target in Jurkat T cells. c Flow cytometry analysis of CXCR4 expression on the cell surface. Jurkat T cell line was transduced with X4R5-Cas9 lentivirus at MOI = 40. CCR5 surface expression detection was excluded because of its low expression on Jurkat T cells. d Detection of protein level of CXCR4 and CCR5 after Jurkat cells were transduced with X4R5-Cas9 lentivirus. e HIV-1 titer change detected by p24 gag ELISA from day 1 to day 5 post-infection. The data shown were the mean ± SD of three independent experiments. *P < 0.05; **P < 0.01; ***P < 0.001; t test
Fig. 3 Jurkat T cell line modified by X4R5-Cas9 lentivirus antagonized HIV infection. a T7E1 assay identification of packaged X4R5-Cas9 lentivirus mediated cleavage at the genome level. b On-target analysis of each target in Jurkat T cells. c Flow cytometry analysis of CXCR4 expression on the cell surface. Jurkat T cell line was transduced with X4R5-Cas9 lentivirus at MOI = 40. CCR5 surface expression detection was excluded because of its low expression on Jurkat T cells. d Detection of protein level of CXCR4 and CCR5 after Jurkat cells were transduced with X4R5-Cas9 lentivirus. e HIV-1 titer change detected by p24 gag ELISA from day 1 to day 5 post-infection. The data shown were the mean ± SD of three independent experiments. *P < 0.05; **P < 0.01; ***P < 0.001; t testCC BY · PubMed Central
Fig. 4 X4R5-Cas9 lentivirus modified Jurkat T cells were enriched after CXCR4-tropic (NL4-3) and CCR5-tropic (YU-2) HIV-1 challenge. a HIV replication in X4R5-Cas9 lentivirus modified as well as mock and control Jurkat T cells infected with X4-tropic and R5-tropic HIV-1 concurrently. Values represent the mean of duplicate infections. b cleavage analysis of CXCR4 and CCR5 by T7 endonuclease 1 in mock, control, lenti-X4R5-Cas9-#1 and lenti-X4R5-Cas9-#2 group at 0, 9 and 18 days after HIV-1 challenge. The lower migrating bands (indicated by arrows) in each lane indicate the disrupted CXCR4 and CCR5 alleles. DPI days post infection
Fig. 4 X4R5-Cas9 lentivirus modified Jurkat T cells were enriched after CXCR4-tropic (NL4-3) and CCR5-tropic (YU-2) HIV-1 challenge. a HIV replication in X4R5-Cas9 lentivirus modified as well as mock and control Jurkat T cells infected with X4-tropic and R5-tropic HIV-1 concurrently. Values represent the mean of duplicate infections. b cleavage analysis of CXCR4 and CCR5 by T7 endonuclease 1 in mock, control, lenti-X4R5-Cas9-#1 and lenti-X4R5-Cas9-#2 group at 0, 9 and 18 days after HIV-1 challenge. The lower migrating bands (indicated by arrows) in each lane indicate the disrupted CXCR4 and CCR5 alleles. DPI days post infectionCC BY · PubMed Central
Fig. 5 lenti-X4R5-Cas9 modified primary CD4+ T cell resists HIV challenge. a T7E1 analysis of CXCR4 and CCR5 disruption. b Deep sequencing analysis of typical NHEJ (indels) of related targets. c lenti-X4R5-Cas9 modified CD4+ T cell challenged with HIV-1NL4-3 or HIV-1YU-2. d lenti-X4R5-Cas9 modified CD4+ T cell exposed to dual-tropic HIV-1 variants (NL4-3 & YU-2, 1:1). The CCR5, CXCR4-1, CXCR4-2 represent single disruption of CCR5 or CXCR4, which use the same corresponding gRNAs used in lenti-X4R5-Cas9-#1 or #2. The data shown were the mean ± SD of three independent experiments. *P < 0.05; **P < 0.01; ***P < 0.001; t test
Fig. 5 lenti-X4R5-Cas9 modified primary CD4+ T cell resists HIV challenge. a T7E1 analysis of CXCR4 and CCR5 disruption. b Deep sequencing analysis of typical NHEJ (indels) of related targets. c lenti-X4R5-Cas9 modified CD4+ T cell challenged with HIV-1NL4-3 or HIV-1YU-2. d lenti-X4R5-Cas9 modified CD4+ T cell exposed to dual-tropic HIV-1 variants (NL4-3 & YU-2, 1:1). The CCR5, CXCR4-1, CXCR4-2 represent single disruption of CCR5 or CXCR4, which use the same corresponding gRNAs used in lenti-X4R5-Cas9-#1 or #2. The data shown were the mean ± SD of three independent experiments. *P < 0.05; **P < 0.01; ***P < 0.001; t testCC BY · PubMed Central
Fig. 6 off-target analysis of CCR5. a Mutation frequency analysis at predicted off-target sites of CCR5. The off-target sites were predicted and aligned with the human genome. The sites were amplified and cloned into T-vector and subjected to sequencing. b T7E1 analysis of all predicted off-target sites
Fig. 6 off-target analysis of CCR5. a Mutation frequency analysis at predicted off-target sites of CCR5. The off-target sites were predicted and aligned with the human genome. The sites were amplified and cloned into T-vector and subjected to sequencing. b T7E1 analysis of all predicted off-target sitesCC BY · PubMed Central
Fig. 7 Apoptosis analysis after lenti-X4R5-Cas9 modification in primary CD4+ T cells. a Annexin V and 7-AAD were utilized to stain modified CD4+T at 1,3 and 5 days post nucleofection with flow cytometry. Necrotic cells (Annexin V-/7AAD +), necrotic or late apoptotic cells (Annexin V +/7AAD +); early apoptotic cells (Annexin V +/7AAD-); viable cells (Annexin V-/7AAD-). b Early apoptosis ratio at day 1,3,5 post gene disruption. c relative early apoptosis ratio (Q3/total none viable cells). The total none viable cells = Q2 + Q3. The data shown were the mean ± SD of three independent experiments. *P < 0.05
Fig. 7 Apoptosis analysis after lenti-X4R5-Cas9 modification in primary CD4+ T cells. a Annexin V and 7-AAD were utilized to stain modified CD4+T at 1,3 and 5 days post nucleofection with flow cytometry. Necrotic cells (Annexin V-/7AAD +), necrotic or late apoptotic cells (Annexin V +/7AAD +); early apoptotic cells (Annexin V +/7AAD-); viable cells (Annexin V-/7AAD-). b Early apoptosis ratio at day 1,3,5 post gene disruption. c relative early apoptosis ratio (Q3/total none viable cells). The total none viable cells = Q2 + Q3. The data shown were the mean ± SD of three independent experiments. *P < 0.05CC BY · PubMed Central

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Research works printed but not linked: 42

  1. Update on acquired immune deficiency syndrome (AIDS)–United States. MMWR. 1982;31(37):507-8, 13-4. (Epub 1982/09/24).
  2. Bayer R, Levine C, Murray TH. Guidelines for confidentiality in research on AIDS. AIDS Res. 1983;1(4):275–97 (Epub 1983/01/01). · DOI
  3. Cunningham AL, Donaghy H, Harman AN, Kim M, Turville SG. Manipulation of dendritic cell function by viruses. Curr Opin Microbiol. 2010;13(4):524–9 (Epub 2010/07/06). · DOI
  4. Klasse PJ. The molecular basis of HIV entry. Cell Microbiol. 2012;14(8):1183–92 (Epub 2012/05/16). · DOI
  5. Chan DC, Kim PS. HIV entry and its inhibition. Cell. 1998;93(5):681–4 (Epub 1998/06/18). · DOI
  6. Zheng YH, Lovsin N, Peterlin BM. Newly identified host factors modulate HIV replication. Immunol Lett. 2005;97(2):225–34 (Epub 2005/03/09). · DOI
  7. Kim SY, Byrn R, Groopman J, Baltimore D. Temporal aspects of DNA and RNA synthesis during human immunodeficiency virus infection: evidence for differential gene expression. J Virol. 1989;63(9):3708–13 (Epub 1989/09/01). · DOI
  8. Pomerantz RJ, Trono D, Feinberg MB, Baltimore D. Cells nonproductively infected with HIV-1 exhibit an aberrant pattern of viral RNA expression: a molecular model for latency. Cell. 1990;61(7):1271–6 (Epub 1990/06/29). · DOI
  9. Moore RD, Chaisson RE. Natural history of HIV infection in the era of combination antiretroviral therapy. Aids. 1999;13(14):1933–42. · DOI
  10. Palella FJ Jr, Delaney KM, Moorman AC, Loveless MO, Fuhrer J, Satten GA, et al. Declining morbidity and mortality among patients with advanced human immunodeficiency virus infection. HIV Outpatient Study Investigators. N Engl J Med. 1998;338(13):853–60. · DOI
  11. Durand CM, Blankson JN, Siliciano RF. Developing strategies for HIV-1 eradication. Trends Immunol. 2012;33(11):554–62. · DOI
  12. Siliciano JD, Siliciano RF. HIV-1 eradication strategies: design and assessment. Curr Opin HIV AIDS. 2013;8(4):318–25.
  13. Blas-Garcia A, Apostolova N, Esplugues JV. Oxidative stress and mitochondrial impairment after treatment with anti-HIV drugs: clinical implications. Curr Pharm Des. 2011;17(36):4076–86 (Epub 2011/12/23). · DOI
  14. Liu R, Paxton WA, Choe S, Ceradini D, Martin SR, Horuk R, et al. Homozygous defect in HIV-1 coreceptor accounts for resistance of some multiply-exposed individuals to HIV-1 infection. Cell. 1996;86(3):367–77 (Epub 1996/08/09). · DOI · Paper page
  15. Dean M, Carrington M, Winkler C, Huttley GA, Smith MW, Allikmets R, et al. Genetic restriction of HIV-1 infection and progression to AIDS by a deletion allele of the CKR5 structural gene. Hemophilia growth and Development Study, Multicenter AIDS Cohort Study, Multicenter Hemophilia Cohort Study, San Francisco City Cohort, ALIVE Study. Science. 1996;273(5283):1856–62 (Epub 1996/09/27). · DOI
  16. Broder CC, Collman RG. Chemokine receptors and HIV. J Leukoc Biol. 1997;62(1):20–9 (Epub 1997/07/01). · DOI
  17. Hutter G, Nowak D, Mossner M, Ganepola S, Mussig A, Allers K, et al. Long-term control of HIV by CCR5 Delta32/Delta32 stem-cell transplantation. New Engl J Med. 2009;360(7):692–8 (Epub 2009/02/14). · DOI
  18. Allers K, Hutter G, Hofmann J, Loddenkemper C, Rieger K, Thiel E, et al. Evidence for the cure of HIV infection by CCR5Delta32/Delta32 stem cell transplantation. Blood. 2011;117(10):2791–9 (Epub 2010/12/15). · DOI · Paper page
  19. Yuan J, Wang J, Crain K, Fearns C, Kim KA, Hua KL, et al. Zinc-finger nuclease editing of human cxcr4 promotes HIV-1 CD4+ T cell resistance and enrichment. Mol Ther. 2012;20(4):849–59 (Epub 2012/01/26). · DOI · Paper page
  20. Didigu CA, Wilen CB, Wang J, Duong J, Secreto AJ, Danet-Desnoyers GA, et al. Simultaneous zinc-finger nuclease editing of the HIV coreceptors ccr5 and cxcr4 protects CD4+ T cells from HIV-1 infection. Blood. 2014;123(1):61–9 (Epub 2013/10/29). · DOI
  21. Tebas P, Stein D, Tang WW, Frank I, Wang SQ, Lee G, et al. Gene editing of CCR5 in autologous CD4 T cells of persons infected with HIV. New Engl J Med. 2014;370(10):901–10 (Epub 2014/03/07). · DOI · Paper page
  22. Cong L, Ran FA, Cox D, Lin S, Barretto R, Habib N, et al. Multiplex genome engineering using CRISPR/Cas systems. Science. 2013;339(6121):819–23. · DOI · Paper page
  23. Hsu PD, Lander ES, Zhang F. Development and applications of CRISPR-Cas9 for genome engineering. Cell. 2014;157(6):1262–78 (Epub 2014/06/07). · DOI · Paper page
  24. Makarova KS, Haft DH, Barrangou R, Brouns SJ, Charpentier E, Horvath P, et al. Evolution and classification of the CRISPR-Cas systems. Nat Rev Microbiol. 2011;9(6):467–77. · DOI · Paper page
  25. 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 (Epub 2012/06/30). · DOI · Paper page
  26. Wang W, Ye C, Liu J, Zhang D, Kimata JT, Zhou P. CCR5 gene disruption via lentiviral vectors expressing Cas9 and single guided RNA renders cells resistant to HIV-1 infection. PloS one. 2014;9(12):e115987 (Epub 2014/12/30). · DOI
  27. Hutter G, Bodor J, Ledger S, Boyd M, Millington M, Tsie M, et al. CCR5 targeted cell therapy for HIV and prevention of viral escape. Viruses. 2015;7(8):4186–203 (Epub 2015/08/01). · DOI
  28. Poveda E. HIV tropism shift: new paradigm on cell therapy strategies for HIV cure. AIDS Rev. 2015;17(1):65 (Epub 2015/01/23).
  29. Hou P, Chen S, Wang S, Yu X, Chen Y, Jiang M, et al. Genome editing of CXCR4 by CRISPR/cas9 confers cells resistant to HIV-1 infection. Sci Rep. 2015;5:15577. · DOI
  30. Yu S, Yao Y, Xiao H, Li J, Liu Q, Yang Y, et al. Simultaneous knockout of CXCR4 and CCR5 genes in CD4+ T cells via CRISPR/Cas9 confers resistance to both X4- and R5-tropic HIV-1 infection. Human gene therapy. 2017. doi: 10.1089/hum.2017.032 . · DOI
  31. Ye L, Wang J, Beyer AI, Teque F, Cradick TJ, Qi Z, et al. Seamless modification of wild-type induced pluripotent stem cells to the natural CCR5Delta32 mutation confers resistance to HIV infection. Proc Natl Acad Sci USA. 2014;111(26):9591–6 (Epub 2014/06/14). · DOI · Paper page
  32. Gao G, Wu X, Zhou J, He M, He JJ, Guo D. Inhibition of HIV-1 transcription and replication by a newly identified cyclin T1 splice variant. J Biol Chem. 2013;288(20):14297–309. · DOI
  33. Yu S, Yao Y, Xiao H, Li J, Liu Q, Yang Y, et al. Simultaneous Knockout of CXCR4 and CCR5 Genes in CD4+ T Cells via CRISPR/Cas9 Confers Resistance to Both X4- and R5-Tropic Human Immunodeficiency Virus Type 1 Infection. Human gene therapy. 2017. doi: 10.1089/hum.2017.032 · DOI
  34. Duan J, Lu G, Xie Z, Lou M, Luo J, Guo L, et al. Genome-wide identification of CRISPR/Cas9 off-targets in human genome. Cell Res. 2014;24(8):1009–12. · DOI · Paper page
  35. Kang H, Minder P, Park MA, Mesquitta WT, Torbett BE, Slukvin II. CCR5 disruption in induced pluripotent stem cells using CRISPR/Cas9 provides selective resistance of immune cells to CCR5-tropic HIV-1 virus. Mol Ther Nucleic Acids. 2015;4:e268 (Epub 2015/12/17). · DOI
  36. Shi B, Li J, Shi X, Jia W, Wen Y, Hu X, et al. TALEN-mediated knockout of CCR5 confers protection against infection of human immunodeficiency virus. J Acquir Immune Defic Syndr. 2017;74(2):229–41 (Epub 2016/10/18). · DOI
  37. Kordelas L, Verheyen J, Beelen DW, Horn PA, Heinold A, Kaiser R, et al. Shift of HIV tropism in stem-cell transplantation with CCR5 Delta32 mutation. New Engl J Med. 2014;371(9):880–2. · DOI
  38. Badia R, Riveira-Munoz E, Clotet B, Este JA, Ballana E. Gene editing using a zinc-finger nuclease mimicking the CCR5Delta32 mutation induces resistance to CCR5-using HIV-1. J Antimicrob Chemother. 2014;69(7):1755–9 (Epub 2014/03/22). · DOI
  39. Wang C, Abdel-Mohsen M, Strain MC, Lada SM, Yukl S, Cockerham LR, et al. Decreased HIV type 1 transcription in CCR5-Delta32 heterozygotes during suppressive antiretroviral therapy. J Infect Dis. 2014;210(11):1838–43 (Epub 2014/06/18). · DOI
  40. Aiuti A, Webb IJ, Bleul C, Springer T, Gutierrez-Ramos JC. The chemokine SDF-1 is a chemoattractant for human CD34+ hematopoietic progenitor cells and provides a new mechanism to explain the mobilization of CD34+ progenitors to peripheral blood. J Exp Med. 1997;185(1):111–20 (Epub 1997/01/06). · DOI
  41. Peled A, Petit I, Kollet O, Magid M, Ponomaryov T, Byk T, et al. Dependence of human stem cell engraftment and repopulation of NOD/SCID mice on CXCR4. Science. 1999;283(5403):845–8 (Epub 1999/02/05). · DOI · Paper page
  42. Chung SH, Seki K, Choi BI, Kimura KB, Ito A, Fujikado N, et al. CXC chemokine receptor 4 expressed in T cells plays an important role in the development of collagen-induced arthritis. Arthr Res Ther. 2010;12(5):R188 (Epub 2010/10/14). · DOI

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Metadata

Authors
Liu, Z., Chen, S., Jin, X., Wang, Q., Yang, K., Li, C., Xiao, Q., Hou, P., Liu, S., Wu, S., Hou, W., Xiong, Y., Kong, C., Zhao, X., Wu, L., Li, C., Sun, G., & Guo, D.
ORCID
Guo, Deyin
Venue
Cell & Bioscience
Publisher
Springer Science and Business Media LLC
Published
2017-09-09
Volume
7
Issue
1
ISSN
2045-3701
DOI
10.1186/s13578-017-0174-2
PMC
PMC5591563
Cited by
131
References
8
Language
en
Funders
  • China National Special Research Program of Major Infectious Diseases (2014ZX10001003)
  • Hubei Provincial Science & Technology Innovation Team Grant (2012FFA043)
  • Natural Science Foundation of China (81401659)
  • China Postdoctoral Science Foundation (2015T80838)
  • China Postdoctoral Science Foundation (CN) (2014M560622)

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