The effect of crRNA–target mismatches on cOA-mediated interference by a type III-A CRISPR-Cas system
CRISPR systems elicit interference when a foreign nucleic acid is detected by its ability to base-pair to crRNA. Understanding what degree of complementarity between a foreign nucleic acid and crRNA is required for interference is a central question in the study of CRISPR systems. A clear descriptio...
| Main Authors: | , , |
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| Format: | Article |
| Language: | English |
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Taylor & Francis Group
2022-12-01
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| Series: | RNA Biology |
| Subjects: | |
| Online Access: | http://dx.doi.org/10.1080/15476286.2022.2150812 |
| _version_ | 1797403949244874752 |
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| author | Mohamed Nasef Sarah A. Khweis Jack A. Dunkle |
| author_facet | Mohamed Nasef Sarah A. Khweis Jack A. Dunkle |
| author_sort | Mohamed Nasef |
| collection | DOAJ |
| description | CRISPR systems elicit interference when a foreign nucleic acid is detected by its ability to base-pair to crRNA. Understanding what degree of complementarity between a foreign nucleic acid and crRNA is required for interference is a central question in the study of CRISPR systems. A clear description of which target–crRNA mismatches abrogate interference in type III, Cas10-containing, CRISPR systems has proved elusive due to the complexity of the system which utilizes three distinct interference activities. We characterized the effect of target–crRNA mismatches on in vitro cyclic oligoadenylate (cOA) synthesis and in vivo in an interference assay that depends on cOA synthesis. We found that sequence context affected whether a mismatched target was recognized by crRNA both in vitro and in vivo. We also investigated how the position of a mismatch within the target–crRNA duplex affected recognition by crRNA. Our data provide support for the hypothesis that a Cas10-activating region exists in the crRNA–target duplex, that the Cas10-proximal region of the duplex is the most critical in regulating cOA synthesis. Understanding the rules governing target recognition by type III CRISPR systems is critical: as one of the most prevalent CRISPR systems in nature, it plays an important role in the survival of many genera of bacteria. Recently, type III systems were re-purposed as a sensitive and accurate molecular diagnostic tool. Understanding the rules of target recognition in this system will be critical as it is engineered for biotechnology purposes. |
| first_indexed | 2024-03-09T02:46:59Z |
| format | Article |
| id | doaj.art-1683acffc2d94ad49e7ca8d97cf3f9c2 |
| institution | Directory Open Access Journal |
| issn | 1547-6286 1555-8584 |
| language | English |
| last_indexed | 2024-03-09T02:46:59Z |
| publishDate | 2022-12-01 |
| publisher | Taylor & Francis Group |
| record_format | Article |
| series | RNA Biology |
| spelling | doaj.art-1683acffc2d94ad49e7ca8d97cf3f9c22023-12-05T16:09:51ZengTaylor & Francis GroupRNA Biology1547-62861555-85842022-12-011911293130410.1080/15476286.2022.21508122150812The effect of crRNA–target mismatches on cOA-mediated interference by a type III-A CRISPR-Cas systemMohamed Nasef0Sarah A. Khweis1Jack A. Dunkle2University of AlabamaUniversity of AlabamaUniversity of AlabamaCRISPR systems elicit interference when a foreign nucleic acid is detected by its ability to base-pair to crRNA. Understanding what degree of complementarity between a foreign nucleic acid and crRNA is required for interference is a central question in the study of CRISPR systems. A clear description of which target–crRNA mismatches abrogate interference in type III, Cas10-containing, CRISPR systems has proved elusive due to the complexity of the system which utilizes three distinct interference activities. We characterized the effect of target–crRNA mismatches on in vitro cyclic oligoadenylate (cOA) synthesis and in vivo in an interference assay that depends on cOA synthesis. We found that sequence context affected whether a mismatched target was recognized by crRNA both in vitro and in vivo. We also investigated how the position of a mismatch within the target–crRNA duplex affected recognition by crRNA. Our data provide support for the hypothesis that a Cas10-activating region exists in the crRNA–target duplex, that the Cas10-proximal region of the duplex is the most critical in regulating cOA synthesis. Understanding the rules governing target recognition by type III CRISPR systems is critical: as one of the most prevalent CRISPR systems in nature, it plays an important role in the survival of many genera of bacteria. Recently, type III systems were re-purposed as a sensitive and accurate molecular diagnostic tool. Understanding the rules of target recognition in this system will be critical as it is engineered for biotechnology purposes.http://dx.doi.org/10.1080/15476286.2022.2150812crispr-cascrrnacas10-csmcyclic oligoadenylatesbacterial immunity |
| spellingShingle | Mohamed Nasef Sarah A. Khweis Jack A. Dunkle The effect of crRNA–target mismatches on cOA-mediated interference by a type III-A CRISPR-Cas system RNA Biology crispr-cas crrna cas10-csm cyclic oligoadenylates bacterial immunity |
| title | The effect of crRNA–target mismatches on cOA-mediated interference by a type III-A CRISPR-Cas system |
| title_full | The effect of crRNA–target mismatches on cOA-mediated interference by a type III-A CRISPR-Cas system |
| title_fullStr | The effect of crRNA–target mismatches on cOA-mediated interference by a type III-A CRISPR-Cas system |
| title_full_unstemmed | The effect of crRNA–target mismatches on cOA-mediated interference by a type III-A CRISPR-Cas system |
| title_short | The effect of crRNA–target mismatches on cOA-mediated interference by a type III-A CRISPR-Cas system |
| title_sort | effect of crrna target mismatches on coa mediated interference by a type iii a crispr cas system |
| topic | crispr-cas crrna cas10-csm cyclic oligoadenylates bacterial immunity |
| url | http://dx.doi.org/10.1080/15476286.2022.2150812 |
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