Real-time observation of DNA recognition and rejection by the RNA-guided endonuclease Cas9

Nature Communications

Volumn 7, Issue , 2016, Pages

  Singh, Digvijay ^a,f,g   Sternberg, Samuel H ^b   Fei, Jingyi ^c,d,h   Doudna, Jennifer A ^b,d,e   Ha, Taekjip ^a,c,d,g  

^a UNIVERSITY OF ILLINOIS AT URBANA CHAMPAIGN   (United States)

^b UNIVERSITY OF CALIFORNIA   (United States)

^c University of Illinois at Urbana Champaign   (United States)

^d HOWARD HUGHES MEDICAL INSTITUTE   (United States)

^e LAWRENCE BERKELEY NATIONAL LABORATORY   (United States)

^f JOHNS HOPKINS UNIVERSITY SCHOOL OF MEDICINE   (United States)

^g Johns Hopkins University   (United States)

^h UNIVERSITY OF CHICAGO   (United States)

Author keywords

[No Author keywords available]

Indexed keywords

CASPASE 9; DNA; ENDONUCLEASE; RNA; BACTERIAL PROTEIN; CAS9 ENDONUCLEASE STREPTOCOCCUS PYOGENES; GUIDE RNA;

CHEMICAL BINDING; DNA; GENE; GENE EXPRESSION; GENETIC ANALYSIS; GENOMICS; REAL TIME; RNA;

ARTICLE; BOUND FRACTION; CONTROLLED STUDY; DISSOCIATION; DNA BINDING; DNA CLEAVAGE; DNA RNA HYBRIDIZATION; FLUORESCENCE RESONANCE ENERGY TRANSFER; GENE INTERACTION; HUMAN; PROTEIN EXPRESSION; PROTEIN PURIFICATION; RNA BINDING; BASE MISPAIRING; CRISPR CAS SYSTEM; ESCHERICHIA COLI; KINETICS; METABOLISM;

BACTERIAL PROTEINS; BASE PAIR MISMATCH; CRISPR-CAS SYSTEMS; DNA; ENDONUCLEASES; ESCHERICHIA COLI; KINETICS; RNA, GUIDE;

EID: 84987837936     PISSN: None     EISSN: 20411723     Source Type: Journal    
DOI: 10.1038/ncomms12778     Document Type: Article

References (68)

1. Wright, A. V., Nunez, J. K., Doudna, J. A. Biology and applications of CRISPR systems: harnessing nature's toolbox for genome engineering. Cell 164, 29-44 (2016).
2. Jinek, M. et al. A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity. Science 337, 816-821 (2012).
3. Gasiunas, G., Barrangou, R., Horvath, P., Siksnys, V. Cas9-crRNA ribonucleoprotein complex mediates specific DNA cleavage for adaptive immunity in bacteria. Proc. Natl Acad. Sci. USA 109, E2579-E2586 (2012).
4. Sternberg, S. H., Redding, S., Jinek, M., Greene, E. C., Doudna, J. A. DNA interrogation by the CRISPR RNA-guided endonuclease Cas9. Nature 507, 62-67 (2014).
5. Doudna, J. A., Charpentier, E. Genome editing. The new frontier of genome engineering with CRISPR-Cas9. Science 346, 1258096 (2014).
6. Wu, X., Kriz, A. J., Sharp, P. A. Target specificity of the CRISPR-Cas9 system. Quant. Biol. 2, 59-70 (2014).
7. O'Geen, H., Yu, A. S., Segal, D. J. How specific is CRISPR/Cas9 really? Curr. Opin. Chem. Biol. 29, 72-78 (2015).
8. Sapranauskas, R. et al. The Streptococcus thermophilus CRISPR/Cas system provides immunity in Escherichia coli. Nucleic Acids Res. 39, 9275-9282 (2011).
9. Semenova, E. et al. Interference by clustered regularly interspaced short palindromic repeat (CRISPR) RNA is governed by a seed sequence. Proc. Natl Acad. Sci. USA 108, 10098-10103 (2011).
10. Cong, L. et al. Multiplex genome engineering using CRISPR/Cas systems. Science 339, 819-823 (2013).
11. Fu, Y. et al. High-frequency off-target mutagenesis induced by CRISPR-Cas nucleases in human cells. Nat. Biotechnol. 31, 822-826 (2013).
12. Hsu, P. D. et al. DNA targeting specificity of RNA-guided Cas9 nucleases. Nat. Biotechnol. 31, 827-832 (2013).
13. Jiang, W., Bikard, D., Cox, D., Zhang, F., Marraffini, L. A. RNA-guided editing of bacterial genomes using CRISPR-Cas systems. Nat. Biotechnol. 31, 233-239 (2013).
14. Jinek, M. et al. RNA-programmed genome editing in human cells. Elife 2, e00471 (2013).
15. Mali, P. et al. CAS9 transcriptional activators for target specificity screening and paired nickases for cooperative genome engineering. Nat. Biotechnol. 31, 833-838 (2013).
16. Pattanayak, V. et al. High-throughput profiling of off-target DNA cleavage reveals RNA-programmed Cas9 nuclease specificity. Nat. Biotechnol. 31, 839-843 (2013).
17. Cho, S. W. et al. Analysis of off-target effects of CRISPR/Cas-derived RNAguided endonucleases and nickases. Genome Res. 24, 132-141 (2014).
18. Doench, J. G. et al. Rational design of highly active sgRNAs for CRISPR-Cas9-mediated gene inactivation. Nat. Biotechnol. 32, 1262-1267 (2014).
19. Jinek, M. et al. Structures of Cas9 endonucleases reveal RNA-mediated conformational activation. Science 343, 1247997 (2014).
20. Gagnon, J. A. et al. Efficient mutagenesis by Cas9 protein-mediated oligonucleotide insertion and large-scale assessment of single-guide RNAs. PLoS ONE 9, e98186 (2014).
21. Smith, C. et al. Whole-genome sequencing analysis reveals high specificity of CRISPR/Cas9 and TALEN-based genome editing in human iPSCs. Cell Stem Cell 15, 12-13 (2014).
22. Shen, B. et al. Efficient genome modification by CRISPR-Cas9 nickase with minimal off-target effects. Nat. Methods 11, 399-402 (2014).
23. Zhang, Y. et al. Comparison of non-canonical PAMs for CRISPR/Cas9-mediated DNA cleavage in human cells. Sci. Rep. 4, 5405 (2014).
24. Frock, R. L. et al. Genome-wide detection of DNA double-stranded breaks induced by engineered nucleases. Nat. Biotechnol. 33, 179-186 (2015).
25. Iyer, V. et al. Off-target mutations are rare in Cas9-modified mice. Nat. Methods 12, 479 (2015).
26. Kim, D. et al. Digenome-seq: genome-wide profiling of CRISPR-Cas9 off-target effects in human cells. Nat. Methods 12, 237-243 (2015).
27. Paulis, M. et al. A pre-screening FISH-based method to detect CRISPR/Cas9 off-targets in mouse embryonic stem cells. Sci. Rep. 5, 12327 (2015).
28. Singh, R., Kuscu, C., Quinlan, A., Qi, Y., Adli, M. Cas9-chromatin binding information enables more accurate CRISPR off-target prediction. Nucleic Acids Res. 43, e118 (2015).
29. Tan, E.-P., Li, Y., Velasco-Herrera, M. D. C., Yusa, K., Bradley, A. Off-target assessment of CRISPR-Cas9 guiding RNAs in human iPS and mouse ES cells. Genesis 53, 225-236 (2015).
30. Tsai, S. Q. et al. GUIDE-seq enables genome-wide profiling of off-target cleavage by CRISPR-Cas nucleases. Nat. Biotechnol. 33, 187-197 (2015).
31. Wang, X. et al. Unbiased detection of off-target cleavage by CRISPR-Cas9 and TALENs using integrase-defective lentiviral vectors. Nat. Biotechnol. 33, 175-178 (2015).
32. Xu, H. et al. Sequence determinants of improved CRISPR sgRNA design. Genome Res. 25, 1147-1157 (2015).
33. Doench, J. G. et al. Optimized sgRNA design to maximize activity and minimize off-target effects of CRISPR-Cas9. Nat. Biotechnol. 34, 184-191 (2016).
34. Cencic, R. et al. Protospacer adjacent motif (PAM)-distal sequences engage CRISPR Cas9 DNA target cleavage. PLoS ONE 9, e109213 (2014).
35. Duan, J. et al. Genome-wide identification of CRISPR/Cas9 off-targets in human genome. Cell Res. 24, 1009-1012 (2014).
36. Kuscu, C., Arslan, S., Singh, R., Thorpe, J., Adli, M. Genome-wide analysis reveals characteristics of off-target sites bound by the Cas9 endonuclease. Nat. Biotechnol. 32, 677-683 (2014).
37. Wu, X. et al. Genome-wide binding of the CRISPR endonuclease Cas9 in mammalian cells. Nat. Biotechnol. 32, 670-676 (2014).
38. O'Geen, H., Henry, I. M., Bhakta, M. S., Meckler, J. F., Segal, D. J. A genomewide analysis of Cas9 binding specificity using ChIP-seq and targeted sequence capture. Nucleic Acids Res. 43, 3389-3404 (2015).
39. Polstein, L. R. et al. Genome-wide specificity of DNA binding, gene regulation, and chromatin remodeling by TALE-and CRISPR/Cas9-based transcriptional activators. Genome Res. 25, 1158-1169 (2015).
40. Rutkauskas, M. et al. Directional R-loop formation by the CRISPR-Cas surveillance complex cascade provides efficient off-target site rejection. Cell Rep. S2211-1247, 00135-00137 (2015).
41. Josephs, E. A. et al. Structure and specificity of the RNA-guided endonuclease Cas9 during DNA interrogation, target binding and cleavage. Nucleic Acids Res. 43, 8924-8941 (2015).
42. Blosser, T. R. et al. Two distinct DNA binding modes guide dual roles of a CRISPR-Cas protein complex. Mol. Cell 58, 60-70 (2015).
43. Knight, S. C. et al. Dynamics of CRISPR-Cas9 genome interrogation in living cells. Science 350, 823-826 (2015).
44. Joo, C., Balci, H., Ishitsuka, Y., Buranachai, C., Ha, T. Advances in singlemolecule fluorescence methods for molecular biology. Annu. Rev. Biochem. 77, 51-76 (2008).
45. Ragunathan, K., Joo, C., Ha, T. Real-time observation of strand exchange reaction with high spatiotemporal resolution. Structure 19, 1064-1073 (2011).
46. Lee, J. Y. et al. DNA recombination. Base triplet stepping by the Rad51/RecA family of recombinases. Science 349, 977-981 (2015).
47. Ragunathan, K., Liu, C., Ha, T. RecA filament sliding on DNA facilitates homology search. Elife 1, e00067 (2012).
48. Szczelkun, M. D. et al. Direct observation of R-loop formation by single RNAguided Cas9 and Cascade effector complexes. Proc. Natl Acad. Sci. USA 111, 9798-9803 (2014).
49. Redding, S. et al. Surveillance and processing of foreign DNA by the Escherichia coli CRISPR-Cas system. Cell 163, 854-865 (2015).
50. Ha, T. et al. Probing the interaction between two single molecules: fluorescence resonance energy transfer between a single donor and a single acceptor. Proc. Natl Acad. Sci. USA 93, 6264-6268 (1996).
51. Roy, R., Hohng, S., Ha, T. A practical guide to single-molecule FRET. Nat. Methods 5, 507-516 (2008).
52. Anders, C., Niewoehner, O., Duerst, A., Jinek, M. Structural basis of PAM-dependent target DNA recognition by the Cas9 endonuclease. Nature 513, 569-573 (2014).
53. McKinney, S. A., Joo, C., Ha, T. Analysis of single-molecule FRET trajectories using hidden Markov modeling. Biophys. J. 91, 1941-1951 (2006).
54. Slaymaker, I. M. et al. Rationally engineered Cas9 nucleases with improved specificity. Science 351, 84-88 (2016).
55. Kleinstiver, B. P. et al. High-fidelity CRISPR-Cas9 nucleases with no detectable genome-wide off-target effects. Nature 529, 490-495 (2016).
56. Nishimasu, H. et al. Crystal structure of Cas9 in complex with guide RNA and target DNA. Cell 156, 935-949 (2014).
57. Hsu, P. D. et al. DNA targeting specificity of RNA-guided Cas9 nucleases. Nat. Biotechnol. 31, 827-832 (2013).
58. Bae, S., Park, J., Kim, J.-S. Cas-OFFinder: a fast and versatile algorithm that searches for potential off-target sites of Cas9 RNA-guided endonucleases. Bioinformatics 30, 1473-1475 (2014).
59. Heigwer, F., Kerr, G., Boutros, M. E-CRISP: fast CRISPR target site identification. Nat. Methods 11, 122-123 (2014).
60. Ren, X. et al. Enhanced specificity and efficiency of the CRISPR/Cas9 system with optimized sgRNA parameters in Drosophila. Cell Rep. 9, 1151-1162 (2014).
61. Wang, T., Wei, J. J., Sabatini, D. M., Lander, E. S. Genetic screens in human cells using the CRISPR-Cas9 system. Science 343, 80-84 (2014).
62. Chari, R., Mali, P., Moosburner, M., Church, G. M. Unraveling CRISPR-Cas9 genome engineering parameters via a library-on-library approach. Nat. Methods 12, 823-826 (2015).
63. Farboud, B., Meyer, B. J. Dramatic enhancement of genome editing by CRISPR/Cas9 through improved guide RNA design. Genetics 199, 959-971 (2015).
64. Moreno-Mateos, M. A. et al. CRISPRscan: designing highly efficient sgRNAs for CRISPR-Cas9 targeting in vivo. Nat. Methods 12, 982-988 (2015).
65. Stemmer, M., Thumberger, T., Del Sol Keyer, M., Wittbrodt, J., Mateo, J. L. CCTop: an intuitive, flexible and reliable CRISPR/Cas9 target prediction tool. PLoS ONE 10, e0124633 (2015).
66. Tsai, S. Q. et al. GUIDE-seq enables genome-wide profiling of off-target cleavage by CRISPR-Cas nucleases. Nat. Biotechnol. 33, 187-197 (2015).
67. Wong, N., Liu, W., Wang, X. WU-CRISPR: characteristics of functional guide RNAs for the CRISPR/Cas9 system. Genome Biol. 16, 218 (2015).
68. Labun, K. et al. CHOPCHOP v2: a web tool for the next generation of CRISPR genome engineering. Nucleic Acids Res. 44, W272-W276 (2016).