CRISPR-Cas Controls Cryptic Prophages
The bacterial archetypal adaptive immune system, CRISPR-Cas, is thought to be repressed in the best-studied bacterium, <i>Escherichia coli</i> K-12. We show here that the <i>E. coli</i> CRISPR-Cas system is active and serves to inhibit its nine defective (i.e., cryptic) proph...
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| Format: | Article |
| Language: | English |
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MDPI AG
2022-12-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/23/24/16195 |
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| author | Sooyeon Song Ekaterina Semenova Konstantin Severinov Laura Fernández-García Michael J. Benedik Toshinari Maeda Thomas K. Wood |
| author_facet | Sooyeon Song Ekaterina Semenova Konstantin Severinov Laura Fernández-García Michael J. Benedik Toshinari Maeda Thomas K. Wood |
| author_sort | Sooyeon Song |
| collection | DOAJ |
| description | The bacterial archetypal adaptive immune system, CRISPR-Cas, is thought to be repressed in the best-studied bacterium, <i>Escherichia coli</i> K-12. We show here that the <i>E. coli</i> CRISPR-Cas system is active and serves to inhibit its nine defective (i.e., cryptic) prophages. Specifically, compared to the wild-type strain, reducing the amounts of specific interfering RNAs (crRNA) decreases growth by 40%, increases cell death by 700%, and prevents persister cell resuscitation. Similar results were obtained by inactivating CRISPR-Cas by deleting the entire 13 spacer region (CRISPR array); hence, CRISPR-Cas serves to inhibit the remaining deleterious effects of these cryptic prophages, most likely through CRISPR array-derived crRNA binding to cryptic prophage mRNA rather than through cleavage of cryptic prophage DNA, i.e., self-targeting. Consistently, four of the 13 <i>E. coli</i> spacers contain complementary regions to the mRNA sequences of seven cryptic prophages, and inactivation of CRISPR-Cas increases the level of mRNA for lysis protein YdfD of cryptic prophage Qin and lysis protein RzoD of cryptic prophage DLP-12. In addition, lysis is clearly seen via transmission electron microscopy when the whole CRISPR-Cas array is deleted, and eliminating spacer #12, which encodes crRNA with complementary regions for DLP-12 (including <i>rzoD</i>), Rac, Qin (including <i>ydfD</i>), and CP4-57 cryptic prophages, also results in growth inhibition and cell lysis. Therefore, we report the novel results that (i) CRISPR-Cas is active in <i>E. coli</i> and (ii) CRISPR-Cas is used to tame cryptic prophages, likely through RNAi, i.e., unlike with active lysogens, active CRISPR-Cas and cryptic prophages may stably co-exist. |
| first_indexed | 2024-03-09T16:19:19Z |
| format | Article |
| id | doaj.art-fd07c43a9dd44212b39af1db4decd5ca |
| institution | Directory Open Access Journal |
| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-09T16:19:19Z |
| publishDate | 2022-12-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | International Journal of Molecular Sciences |
| spelling | doaj.art-fd07c43a9dd44212b39af1db4decd5ca2023-11-24T15:34:38ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672022-12-0123241619510.3390/ijms232416195CRISPR-Cas Controls Cryptic ProphagesSooyeon Song0Ekaterina Semenova1Konstantin Severinov2Laura Fernández-García3Michael J. Benedik4Toshinari Maeda5Thomas K. Wood6Department of Chemical Engineering, Pennsylvania State University, University Park, PA 16802, USAWaksman Institute of Microbiology, Rutgers, The State University of New Jersey, Piscataway, NJ 08854, USAWaksman Institute of Microbiology, Rutgers, The State University of New Jersey, Piscataway, NJ 08854, USADepartment of Chemical Engineering, Pennsylvania State University, University Park, PA 16802, USAOffice of the Provost, Hamad bin Khalifa University, Education City, Doha P.O. Box 34110, QatarDepartment of Biological Functions Engineering, Kyushu Institute of Technology, Kitakyushu 808-0196, JapanDepartment of Chemical Engineering, Pennsylvania State University, University Park, PA 16802, USAThe bacterial archetypal adaptive immune system, CRISPR-Cas, is thought to be repressed in the best-studied bacterium, <i>Escherichia coli</i> K-12. We show here that the <i>E. coli</i> CRISPR-Cas system is active and serves to inhibit its nine defective (i.e., cryptic) prophages. Specifically, compared to the wild-type strain, reducing the amounts of specific interfering RNAs (crRNA) decreases growth by 40%, increases cell death by 700%, and prevents persister cell resuscitation. Similar results were obtained by inactivating CRISPR-Cas by deleting the entire 13 spacer region (CRISPR array); hence, CRISPR-Cas serves to inhibit the remaining deleterious effects of these cryptic prophages, most likely through CRISPR array-derived crRNA binding to cryptic prophage mRNA rather than through cleavage of cryptic prophage DNA, i.e., self-targeting. Consistently, four of the 13 <i>E. coli</i> spacers contain complementary regions to the mRNA sequences of seven cryptic prophages, and inactivation of CRISPR-Cas increases the level of mRNA for lysis protein YdfD of cryptic prophage Qin and lysis protein RzoD of cryptic prophage DLP-12. In addition, lysis is clearly seen via transmission electron microscopy when the whole CRISPR-Cas array is deleted, and eliminating spacer #12, which encodes crRNA with complementary regions for DLP-12 (including <i>rzoD</i>), Rac, Qin (including <i>ydfD</i>), and CP4-57 cryptic prophages, also results in growth inhibition and cell lysis. Therefore, we report the novel results that (i) CRISPR-Cas is active in <i>E. coli</i> and (ii) CRISPR-Cas is used to tame cryptic prophages, likely through RNAi, i.e., unlike with active lysogens, active CRISPR-Cas and cryptic prophages may stably co-exist.https://www.mdpi.com/1422-0067/23/24/16195CRISPR-Caspersisterscryptic prophage |
| spellingShingle | Sooyeon Song Ekaterina Semenova Konstantin Severinov Laura Fernández-García Michael J. Benedik Toshinari Maeda Thomas K. Wood CRISPR-Cas Controls Cryptic Prophages International Journal of Molecular Sciences CRISPR-Cas persisters cryptic prophage |
| title | CRISPR-Cas Controls Cryptic Prophages |
| title_full | CRISPR-Cas Controls Cryptic Prophages |
| title_fullStr | CRISPR-Cas Controls Cryptic Prophages |
| title_full_unstemmed | CRISPR-Cas Controls Cryptic Prophages |
| title_short | CRISPR-Cas Controls Cryptic Prophages |
| title_sort | crispr cas controls cryptic prophages |
| topic | CRISPR-Cas persisters cryptic prophage |
| url | https://www.mdpi.com/1422-0067/23/24/16195 |
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