Multiplexed activation of endogenous genes by CRISPR-on, an RNA-guided transcriptional activator system
Technologies allowing for specific regulation of endogenous genes are valuable for the study of gene functions and have great potential in therapeutics. We created the CRISPR-on system, a two-component transcriptional activator consisting of a nuclease-dead Cas9 (dCas9) protein fused with a transcri...
| Main Authors: | , , , , , , , , , |
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| Format: | Article |
| Language: | en_US |
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Nature Publishing Group
2014
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| Online Access: | http://hdl.handle.net/1721.1/84630 https://orcid.org/0000-0002-7256-3158 https://orcid.org/0000-0002-2905-0306 |
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| author | Wang, Haoyi Yang, Hui Shi, Linyu Katz, Yarden Theunissen, Thorold W Rangarajan, Sudharshan Jaenisch, Rudolf Shivalila, Chikdu Shakti Dadon, Daniel Benjamin Cheng, Albert Wu |
| author2 | Massachusetts Institute of Technology. Computational and Systems Biology Program |
| author_facet | Massachusetts Institute of Technology. Computational and Systems Biology Program Wang, Haoyi Yang, Hui Shi, Linyu Katz, Yarden Theunissen, Thorold W Rangarajan, Sudharshan Jaenisch, Rudolf Shivalila, Chikdu Shakti Dadon, Daniel Benjamin Cheng, Albert Wu |
| author_sort | Wang, Haoyi |
| collection | MIT |
| description | Technologies allowing for specific regulation of endogenous genes are valuable for the study of gene functions and have great potential in therapeutics. We created the CRISPR-on system, a two-component transcriptional activator consisting of a nuclease-dead Cas9 (dCas9) protein fused with a transcriptional activation domain and single guide RNAs (sgRNAs) with complementary sequence to gene promoters. We demonstrate that CRISPR-on can efficiently activate exogenous reporter genes in both human and mouse cells in a tunable manner. In addition, we show that robust reporter gene activation in vivo can be achieved by injecting the system components into mouse zygotes. Furthermore, we show that CRISPR-on can activate the endogenous IL1RN, SOX2, and OCT4 genes. The most efficient gene activation was achieved by clusters of 3-4 sgRNAs binding to the proximal promoters, suggesting their synergistic action in gene induction. Significantly, when sgRNAs targeting multiple genes were simultaneously introduced into cells, robust multiplexed endogenous gene activation was achieved. Genome-wide expression profiling demonstrated high specificity of the system. |
| first_indexed | 2024-09-23T10:42:03Z |
| format | Article |
| id | mit-1721.1/84630 |
| institution | Massachusetts Institute of Technology |
| language | en_US |
| last_indexed | 2024-09-23T10:42:03Z |
| publishDate | 2014 |
| publisher | Nature Publishing Group |
| record_format | dspace |
| spelling | mit-1721.1/846302022-09-27T14:21:39Z Multiplexed activation of endogenous genes by CRISPR-on, an RNA-guided transcriptional activator system Wang, Haoyi Yang, Hui Shi, Linyu Katz, Yarden Theunissen, Thorold W Rangarajan, Sudharshan Jaenisch, Rudolf Shivalila, Chikdu Shakti Dadon, Daniel Benjamin Cheng, Albert Wu Massachusetts Institute of Technology. Computational and Systems Biology Program Massachusetts Institute of Technology. Department of Biology Massachusetts Institute of Technology. Department of Brain and Cognitive Sciences Whitehead Institute for Biomedical Research Cheng, Albert W. Katz, Yarden Shivalila, Chikdu Shakti Dadon, Daniel Benjamin Jaenisch, Rudolf Technologies allowing for specific regulation of endogenous genes are valuable for the study of gene functions and have great potential in therapeutics. We created the CRISPR-on system, a two-component transcriptional activator consisting of a nuclease-dead Cas9 (dCas9) protein fused with a transcriptional activation domain and single guide RNAs (sgRNAs) with complementary sequence to gene promoters. We demonstrate that CRISPR-on can efficiently activate exogenous reporter genes in both human and mouse cells in a tunable manner. In addition, we show that robust reporter gene activation in vivo can be achieved by injecting the system components into mouse zygotes. Furthermore, we show that CRISPR-on can activate the endogenous IL1RN, SOX2, and OCT4 genes. The most efficient gene activation was achieved by clusters of 3-4 sgRNAs binding to the proximal promoters, suggesting their synergistic action in gene induction. Significantly, when sgRNAs targeting multiple genes were simultaneously introduced into cells, robust multiplexed endogenous gene activation was achieved. Genome-wide expression profiling demonstrated high specificity of the system. National Institutes of Health (U.S.) (Grant HD 045022) National Institutes of Health (U.S.) (Grant R37CA084198) 2014-01-31T19:44:23Z 2014-01-31T19:44:23Z 2013-08 2013-08 Article http://purl.org/eprint/type/JournalArticle 1001-0602 1748-7838 http://hdl.handle.net/1721.1/84630 Cheng, Albert W, Haoyi Wang, Hui Yang, Linyu Shi, Yarden Katz, Thorold W Theunissen, Sudharshan Rangarajan, Chikdu S Shivalila, Daniel B Dadon, and Rudolf Jaenisch. “Multiplexed activation of endogenous genes by CRISPR-on, an RNA-guided transcriptional activator system.” Cell Research 23, no. 10 (August 27, 2013): 1163-1171. © 2013 IBCB, SIBS, CAS https://orcid.org/0000-0002-7256-3158 https://orcid.org/0000-0002-2905-0306 en_US http://dx.doi.org/10.1038/cr.2013.122 Cell Research Creative Commons Attribution-NonCommercial-No Derivative Works 3.0 Unported License http://creativecommons.org/licenses/by-nc-nd/3.0/ application/pdf Nature Publishing Group Cell Research |
| spellingShingle | Wang, Haoyi Yang, Hui Shi, Linyu Katz, Yarden Theunissen, Thorold W Rangarajan, Sudharshan Jaenisch, Rudolf Shivalila, Chikdu Shakti Dadon, Daniel Benjamin Cheng, Albert Wu Multiplexed activation of endogenous genes by CRISPR-on, an RNA-guided transcriptional activator system |
| title | Multiplexed activation of endogenous genes by CRISPR-on, an RNA-guided transcriptional activator system |
| title_full | Multiplexed activation of endogenous genes by CRISPR-on, an RNA-guided transcriptional activator system |
| title_fullStr | Multiplexed activation of endogenous genes by CRISPR-on, an RNA-guided transcriptional activator system |
| title_full_unstemmed | Multiplexed activation of endogenous genes by CRISPR-on, an RNA-guided transcriptional activator system |
| title_short | Multiplexed activation of endogenous genes by CRISPR-on, an RNA-guided transcriptional activator system |
| title_sort | multiplexed activation of endogenous genes by crispr on an rna guided transcriptional activator system |
| url | http://hdl.handle.net/1721.1/84630 https://orcid.org/0000-0002-7256-3158 https://orcid.org/0000-0002-2905-0306 |
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