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| Content Provider | World Health Organization (WHO)-Global Index Medicus |
|---|---|
| Author | Sharma, Kundan Randau, Lennart Lenz, Christof Allers, Thorsten Urlaub, Henning Maier, Lisa-katharina Nickel, Lisa Backofen, Rolf Lange, Sita J. Schmitz, Ruth A. Stachler, Aris-edda Brendel, Jutta Marchfelder, Anita Stoll, Britta Richter, Hagen |
| Description | Author Affiliation: Brendel J ( From the Department of Biology II, Ulm University, 89069 Ulm, Germany.) |
| Abstract | The clustered regularly interspaced short palindromic repeats/CRISPR-associated (CRISPR-Cas) system is a prokaryotic defense mechanism against foreign genetic elements. A plethora of CRISPR-Cas versions exist, with more than 40 different Cas protein families and several different molecular approaches to fight the invading DNA. One of the key players in the system is the CRISPR-derived RNA (crRNA), which directs the invader-degrading Cas protein complex to the invader. The CRISPR-Cas types I and III use the Cas6 protein to generate mature crRNAs. Here, we show that the Cas6 protein is necessary for crRNA production but that additional Cas proteins that form a CRISPR-associated complex for antiviral defense (Cascade)-like complex are needed for crRNA stability in the CRISPR-Cas type I-B system in Haloferax volcanii in vivo. Deletion of the cas6 gene results in the loss of mature crRNAs and interference. However, cells that have the complete cas gene cluster (cas1–8b) removed and are transformed with the cas6 gene are not able to produce and stably maintain mature crRNAs. crRNA production and stability is rescued only if cas5, -6, and -7 are present. Mutational analysis of the cas6 gene reveals three amino acids (His-41, Gly-256, and Gly-258) that are essential for pre-crRNA cleavage, whereas the mutation of two amino acids (Ser-115 and Ser-224) leads to an increase of crRNA amounts. This is the first systematic in vivo analysis of Cas6 protein variants. In addition, we show that the H. volcanii I-B system contains a Cascade-like complex with a Cas7, Cas5, and Cas6 core that protects the crRNA. |
| ISSN | 00219258 |
| e-ISSN | 1083351X |
| Journal | Journal of Biological Chemistry |
| Issue Number | 10 |
| Volume Number | 289 |
| Language | English |
| Publisher | American Society for Biochemistry and Molecular Biology (United States) |
| Publisher Date | 2014-03-07 |
| Publisher Place | United States |
| Access Restriction | Open |
| Subject Keyword | Archaeal Proteins Metabolism Clustered Regularly Interspaced Short Palindromic Repeats Genetics Haloferax Volcanii RNA Stability RNA, Archaeal Chemistry Amino Acid Sequence Molecular Sequence Data Research Support, Non-U.S. Gov't Biochemistry Molecular Biology |
| Content Type | Text |
| Resource Type | Article |
| Subject | Cell Biology Biochemistry Molecular Biology |
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