Orthogonal inducible control of Cas13 circuits enables programmable RNA regulation in mammalian cells
Abstract RNA plays an indispensable role in mammalian cell functions. Cas13, a class of RNA-guided ribonuclease, is a flexible tool for modifying and regulating coding and non-coding RNAs, with enormous potential for creating new cell functions. However, the lack of control over Cas13 activity has l...
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
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Nature Portfolio
2024-02-01
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| Series: | Nature Communications |
| Online Access: | https://doi.org/10.1038/s41467-024-45795-x |
| _version_ | 1797273864164605952 |
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| author | Yage Ding Cristina Tous Jaehoon Choi Jingyao Chen Wilson W. Wong |
| author_facet | Yage Ding Cristina Tous Jaehoon Choi Jingyao Chen Wilson W. Wong |
| author_sort | Yage Ding |
| collection | DOAJ |
| description | Abstract RNA plays an indispensable role in mammalian cell functions. Cas13, a class of RNA-guided ribonuclease, is a flexible tool for modifying and regulating coding and non-coding RNAs, with enormous potential for creating new cell functions. However, the lack of control over Cas13 activity has limited its cell engineering capability. Here, we present the CRISTAL (Control of RNA with Inducible SpliT CAs13 Orthologs and Exogenous Ligands) platform. CRISTAL is powered by a collection (10 total) of orthogonal split inducible Cas13 effectors that can be turned ON or OFF via small molecules in multiple cell types, providing precise temporal control. Also, we engineer Cas13 logic circuits that can respond to endogenous signaling and exogenous small molecule inputs. Furthermore, the orthogonality, low leakiness, and high dynamic range of our inducible Cas13d and Cas13b enable the design and construction of a robust incoherent feedforward loop, leading to near-perfect and tunable adaptation response. Finally, using our inducible Cas13 effectors, we achieve simultaneous multiplexed control of multiple genes in vitro and in mice. Together, our CRISTAL design represents a powerful platform for precisely regulating RNA dynamics to advance cell engineering and elucidate RNA biology. |
| first_indexed | 2024-03-07T14:50:14Z |
| format | Article |
| id | doaj.art-0fcd5fbb0a114c568dc3773dbd052712 |
| institution | Directory Open Access Journal |
| issn | 2041-1723 |
| language | English |
| last_indexed | 2024-03-07T14:50:14Z |
| publishDate | 2024-02-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Nature Communications |
| spelling | doaj.art-0fcd5fbb0a114c568dc3773dbd0527122024-03-05T19:43:00ZengNature PortfolioNature Communications2041-17232024-02-0115111610.1038/s41467-024-45795-xOrthogonal inducible control of Cas13 circuits enables programmable RNA regulation in mammalian cellsYage Ding0Cristina Tous1Jaehoon Choi2Jingyao Chen3Wilson W. Wong4Department of Biomedical Engineering, Biological Design Center, Boston UniversityDepartment of Biomedical Engineering, Biological Design Center, Boston UniversityDepartment of Biomedical Engineering, Biological Design Center, Boston UniversityDepartment of Biomedical Engineering, Biological Design Center, Boston UniversityDepartment of Biomedical Engineering, Biological Design Center, Boston UniversityAbstract RNA plays an indispensable role in mammalian cell functions. Cas13, a class of RNA-guided ribonuclease, is a flexible tool for modifying and regulating coding and non-coding RNAs, with enormous potential for creating new cell functions. However, the lack of control over Cas13 activity has limited its cell engineering capability. Here, we present the CRISTAL (Control of RNA with Inducible SpliT CAs13 Orthologs and Exogenous Ligands) platform. CRISTAL is powered by a collection (10 total) of orthogonal split inducible Cas13 effectors that can be turned ON or OFF via small molecules in multiple cell types, providing precise temporal control. Also, we engineer Cas13 logic circuits that can respond to endogenous signaling and exogenous small molecule inputs. Furthermore, the orthogonality, low leakiness, and high dynamic range of our inducible Cas13d and Cas13b enable the design and construction of a robust incoherent feedforward loop, leading to near-perfect and tunable adaptation response. Finally, using our inducible Cas13 effectors, we achieve simultaneous multiplexed control of multiple genes in vitro and in mice. Together, our CRISTAL design represents a powerful platform for precisely regulating RNA dynamics to advance cell engineering and elucidate RNA biology.https://doi.org/10.1038/s41467-024-45795-x |
| spellingShingle | Yage Ding Cristina Tous Jaehoon Choi Jingyao Chen Wilson W. Wong Orthogonal inducible control of Cas13 circuits enables programmable RNA regulation in mammalian cells Nature Communications |
| title | Orthogonal inducible control of Cas13 circuits enables programmable RNA regulation in mammalian cells |
| title_full | Orthogonal inducible control of Cas13 circuits enables programmable RNA regulation in mammalian cells |
| title_fullStr | Orthogonal inducible control of Cas13 circuits enables programmable RNA regulation in mammalian cells |
| title_full_unstemmed | Orthogonal inducible control of Cas13 circuits enables programmable RNA regulation in mammalian cells |
| title_short | Orthogonal inducible control of Cas13 circuits enables programmable RNA regulation in mammalian cells |
| title_sort | orthogonal inducible control of cas13 circuits enables programmable rna regulation in mammalian cells |
| url | https://doi.org/10.1038/s41467-024-45795-x |
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