The CRISPR effector Cam1 mediates membrane depolarization for phage defence

Baca, Christian F.; Yu, You; Rostøl, Jakob T.; Majumder, Puja; Patel, Dinshaw J.; Marraffini, Luciano A.

doi:10.1038/s41586-023-06902-y

Title: The CRISPR effector Cam1 mediates membrane depolarization for phage defence

Journal Article · 10 January 2024 · Nature (London)

DOI: https://doi.org/10.1038/s41586-023-06902-y · OSTI ID:2470118

^[1];

^[2]; Rostøl, Jakob T. ^[3];

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^[5]

Imperial College, London (United Kingdom); Rockefeller Univ., New York, NY (United States); Memorial Sloan Kettering Cancer Center, New York, NY (United States)
Imperial College, London (United Kingdom); Memorial Sloan Kettering Cancer Center, New York, NY (United States)
Imperial College, London (United Kingdom)
Memorial Sloan Kettering Cancer Center, New York, NY (United States)
Rockefeller Univ., New York, NY (United States)

Prokaryotic type III CRISPR–Cas systems provide immunity against viruses and plasmids using CRISPR-associated Rossman fold (CARF) protein effectors. Recognition of transcripts of these invaders with sequences that are complementary to CRISPR RNA guides leads to the production of cyclic oligoadenylate second messengers, which bind CARF domains and trigger the activity of an effector domain. Whereas most effectors degrade host and invader nucleic acids, some are predicted to contain transmembrane helices without an enzymatic function. Whether and how these CARF–transmembrane helix fusion proteins facilitate the type III CRISPR–Cas immune response remains unknown. Here we investigate the role of cyclic oligoadenylate-activated membrane protein 1 (Cam1) during type III CRISPR immunity. Structural and biochemical analyses reveal that the CARF domains of a Cam1 dimer bind cyclic tetra-adenylate second messengers. In vivo, Cam1 localizes to the membrane, is predicted to form a tetrameric transmembrane pore, and provides defence against viral infection through the induction of membrane depolarization and growth arrest. These results reveal that CRISPR immunity does not always operate through the degradation of nucleic acids, but is instead mediated via a wider range of cellular responses.

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Research Organization:: Argonne National Laboratory (ANL), Argonne, IL (United States). Advanced Photon Source (APS)

Sponsoring Organization:: USDOE Office of Science (SC), Basic Energy Sciences (BES). Scientific User Facilities (SUF)

Grant/Contract Number:: AC02-06CH11357

OSTI ID:: 2470118

Journal Information:: Nature (London), Journal Name: Nature (London) Journal Issue: 7996 Vol. 625; ISSN 0028-0836

Publisher:: Nature Publishing GroupCopyright Statement

Country of Publication:: United States

Language:: English

References (49)

Accurate flow cytometric membrane potential measurement in bacteria using diethyloxacarbocyanine and a ratiometric technique Novo, David; Perlmutter, Nancy G.; Hunt, Richard H. Cytometry, Vol. 35, Issue 1 https://doi.org/10.1002/(SICI)1097-0320(19990101)35:1<55::AID-CYTO8>3.0.CO;2-2	journal	January 1999
Predicting transmembrane protein topology with a hidden markov model: application to complete genomes11Edited by F. Cohen Krogh, Anders; Larsson, Björn; von Heijne, Gunnar Journal of Molecular Biology, Vol. 305, Issue 3 https://doi.org/10.1006/jmbi.2000.4315	journal	January 2001
Stoichiometry of the Large Conductance Bacterial Mechanosensitive Channel of E. coli. A Biochemical Study Sukharev, S. I.; Schroeder, M. J.; McCaslin, D. R. Journal of Membrane Biology, Vol. 171, Issue 3 https://doi.org/10.1007/s002329900570	journal	October 1999
Intervening Sequences of Regularly Spaced Prokaryotic Repeats Derive from Foreign Genetic Elements Mojica, Francisco J. M.; D�ez-Villase�or, Chc)sar; Garc�a-Mart�nez, Jes�s Journal of Molecular Evolution, Vol. 60, Issue 2 https://doi.org/10.1007/s00239-004-0046-3	journal	February 2005
The Repetitive DNA Elements Called CRISPRs and Their Associated Genes: Evidence of Horizontal Transfer Among Prokaryotes Godde, James S.; Bickerton, Amanda Journal of Molecular Evolution, Vol. 62, Issue 6 https://doi.org/10.1007/s00239-005-0223-z	journal	April 2006
RNA-Guided RNA Cleavage by a CRISPR RNA-Cas Protein Complex Hale, Caryn R.; Zhao, Peng; Olson, Sara Cell, Vol. 139, Issue 5 https://doi.org/10.1016/j.cell.2009.07.040	journal	November 2009
Co-transcriptional DNA and RNA Cleavage during Type III CRISPR-Cas Immunity Samai, Poulami; Pyenson, Nora; Jiang, Wenyan Cell, Vol. 161, Issue 5 https://doi.org/10.1016/j.cell.2015.04.027	journal	May 2015
Degradation of Phage Transcripts by CRISPR-Associated RNases Enables Type III CRISPR-Cas Immunity Jiang, Wenyan; Samai, Poulami; Marraffini, Luciano A. Cell, Vol. 164, Issue 4 https://doi.org/10.1016/j.cell.2015.12.053	journal	February 2016
Spatiotemporal Control of Type III-A CRISPR-Cas Immunity: Coupling DNA Degradation with the Target RNA Recognition Kazlauskiene, Migle; Tamulaitis, Gintautas; Kostiuk, Georgij Molecular Cell, Vol. 62, Issue 2 https://doi.org/10.1016/j.molcel.2016.03.024	journal	April 2016
CRISPR-Cas III-A Csm6 CARF Domain Is a Ring Nuclease Triggering Stepwise cA4 Cleavage with ApA>p Formation Terminating RNase Activity Jia, Ning; Jones, Roger; Yang, Guangli Molecular Cell, Vol. 75, Issue 5 https://doi.org/10.1016/j.molcel.2019.06.014	journal	September 2019
Static light scattering to characterize membrane proteins in detergent solution Slotboom, Dirk Jan; Duurkens, Ria H.; Olieman, Kees Methods, Vol. 46, Issue 2 https://doi.org/10.1016/j.ymeth.2008.06.012	journal	October 2008
Comparative network clustering of direct repeats (DRs) and cas genes confirms the possibility of the horizontal transfer of CRISPR locus among bacteria Chakraborty, Sajib; Snijders, Ambrosius P.; Chakravorty, Rajib Molecular Phylogenetics and Evolution, Vol. 56, Issue 3 https://doi.org/10.1016/j.ympev.2010.05.020	journal	September 2010
The toxic shock syndrome exotoxin structural gene is not detectably transmitted by a prophage Kreiswirth, Barry N.; Löfdahl, Sven; Betley, Marsha J. Nature, Vol. 305, Issue 5936 https://doi.org/10.1038/305709a0	journal	October 1983
The CRISPR/Cas bacterial immune system cleaves bacteriophage and plasmid DNA Garneau, Josiane E.; Dupuis, Marie-Ève; Villion, Manuela Nature, Vol. 468, Issue 7320 https://doi.org/10.1038/nature09523	journal	November 2010
CRISPR RNA maturation by trans-encoded small RNA and host factor RNase III Deltcheva, Elitza; Chylinski, Krzysztof; Sharma, Cynthia M. Nature, Vol. 471, Issue 7340 https://doi.org/10.1038/nature09886	journal	March 2011
Conditional tolerance of temperate phages via transcription-dependent CRISPR-Cas targeting Goldberg, Gregory W.; Jiang, Wenyan; Bikard, David Nature, Vol. 514, Issue 7524 https://doi.org/10.1038/nature13637	journal	August 2014
CRISPR–Cas systems exploit viral DNA injection to establish and maintain adaptive immunity Modell, Joshua W.; Jiang, Wenyan; Marraffini, Luciano A. Nature, Vol. 544, Issue 7648 https://doi.org/10.1038/nature21719	journal	March 2017
Type III CRISPR–Cas systems produce cyclic oligoadenylate second messengers Niewoehner, Ole; Garcia-Doval, Carmela; Rostøl, Jakob T. Nature, Vol. 548, Issue 7669 https://doi.org/10.1038/nature23467	journal	July 2017
Structural basis for CRISPR RNA-guided DNA recognition by Cascade Jore, Matthijs M.; Lundgren, Magnus; van Duijn, Esther Nature Structural & Molecular Biology, Vol. 18, Issue 5 https://doi.org/10.1038/nsmb.2019	journal	April 2011
Structure of Csx1-cOA4 complex reveals the basis of RNA decay in Type III-B CRISPR-Cas Molina, Rafael; Stella, Stefano; Feng, Mingxia Nature Communications, Vol. 10, Issue 1 https://doi.org/10.1038/s41467-019-12244-z	journal	September 2019
Structure and mechanism of a Type III CRISPR defence DNA nuclease activated by cyclic oligoadenylate McMahon, Stephen A.; Zhu, Wenlong; Graham, Shirley Nature Communications, Vol. 11, Issue 1 https://doi.org/10.1038/s41467-019-14222-x	journal	January 2020
Non-specific degradation of transcripts promotes plasmid clearance during type III-A CRISPR–Cas immunity Rostøl, Jakob T.; Marraffini, Luciano A. Nature Microbiology, Vol. 4, Issue 4 https://doi.org/10.1038/s41564-018-0353-x	journal	January 2019
Evolutionary classification of CRISPR–Cas systems: a burst of class 2 and derived variants Makarova, Kira S.; Wolf, Yuri I.; Iranzo, Jaime Nature Reviews Microbiology, Vol. 18, Issue 2 https://doi.org/10.1038/s41579-019-0299-x	journal	December 2019
The Card1 nuclease provides defence during type III CRISPR immunity Rostøl, Jakob T.; Xie, Wei; Kuryavyi, Vitaly Nature, Vol. 590, Issue 7847 https://doi.org/10.1038/s41586-021-03206-x	journal	January 2021
Highly accurate protein structure prediction with AlphaFold Jumper, John; Evans, Richard; Pritzel, Alexander Nature https://doi.org/10.1038/s41586-021-03819-2	journal	July 2021
Systematic prediction of genes functionally linked to CRISPR-Cas systems by gene neighborhood analysis Shmakov, Sergey A.; Makarova, Kira S.; Wolf, Yuri I. Proceedings of the National Academy of Sciences, Vol. 115, Issue 23 https://doi.org/10.1073/pnas.1803440115	journal	May 2018
Evidence for the Assembly of a Bacterial Tripartite Multidrug Pump with a Stoichiometry of 3:6:3 Janganan, Thamarai K.; Bavro, Vassiliy N.; Zhang, Li Journal of Biological Chemistry, Vol. 286, Issue 30 https://doi.org/10.1074/jbc.M111.246595	journal	July 2011
Transmembrane Topology of AgrB, the Protein Involved in the Post-translational Modification of AgrD in Staphylococcus aureus Zhang, Linsheng; Gray, Lillian; Novick, Richard P. Journal of Biological Chemistry, Vol. 277, Issue 38 https://doi.org/10.1074/jbc.M205367200	journal	September 2002
Comprehensive search for accessory proteins encoded with archaeal and bacterial type III CRISPR- cas gene cassettes reveals 39 new cas gene families Shah, Shiraz A.; Alkhnbashi, Omer S.; Behler, Juliane RNA Biology, Vol. 16, Issue 4 https://doi.org/10.1080/15476286.2018.1483685	journal	June 2018
The ribonuclease activity of Csm6 is required for anti-plasmid immunity by Type III-A CRISPR-Cas systems Foster, Kawanda; Kalter, Joshua; Woodside, Walter RNA Biology, Vol. 16, Issue 4 https://doi.org/10.1080/15476286.2018.1493334	journal	June 2018
The CRISPR ancillary effector Can2 is a dual-specificity nuclease potentiating type III CRISPR defence Zhu, Wenlong; McQuarrie, Stuart; Grüschow, Sabine Nucleic Acids Research, Vol. 49, Issue 5 https://doi.org/10.1093/nar/gkab073	journal	February 2021
AlphaFold Protein Structure Database: massively expanding the structural coverage of protein-sequence space with high-accuracy models Varadi, Mihaly; Anyango, Stephen; Deshpande, Mandar Nucleic Acids Research, Vol. 50, Issue D1 https://doi.org/10.1093/nar/gkab1061	journal	November 2021
The EMBL-EBI search and sequence analysis tools APIs in 2019 Madeira, Fábio; Park, Young mi; Lee, Joon Nucleic Acids Research, Vol. 47, Issue W1 https://doi.org/10.1093/nar/gkz268	journal	April 2019
Cas6 is an endoribonuclease that generates guide RNAs for invader defense in prokaryotes Carte, J.; Wang, R.; Li, H. Genes & Development, Vol. 22, Issue 24 https://doi.org/10.1101/gad.1742908	journal	December 2008
XDS Kabsch, Wolfgang Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 2 https://doi.org/10.1107/S0907444909047337	journal	January 2010
Features and development of Coot Emsley, P.; Lohkamp, B.; Scott, W. G. Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 4 https://doi.org/10.1107/S0907444910007493	journal	March 2010
Towards automated crystallographic structure refinement with phenix.refine Afonine, Pavel V.; Grosse-Kunstleve, Ralf W.; Echols, Nathaniel Acta Crystallographica Section D Biological Crystallography, Vol. 68, Issue 4 https://doi.org/10.1107/S0907444912001308	journal	March 2012
CRISPR Provides Acquired Resistance Against Viruses in Prokaryotes Barrangou, R.; Fremaux, C.; Deveau, H. Science, Vol. 315, Issue 5819 https://doi.org/10.1126/science.1138140	journal	March 2007
Small CRISPR RNAs Guide Antiviral Defense in Prokaryotes Brouns, S. J. J.; Jore, M. M.; Lundgren, M. Science, Vol. 321, Issue 5891, p. 960-964 https://doi.org/10.1126/science.1159689	journal	August 2008
CRISPR Interference Limits Horizontal Gene Transfer in Staphylococci by Targeting DNA Marraffini, Luciano A.; Sontheimer, Erik J. Science, Vol. 322, Issue 5909 https://doi.org/10.1126/science.1165771	journal	December 2008
A cyclic oligonucleotide signaling pathway in type III CRISPR-Cas systems Kazlauskiene, Migle; Kostiuk, Georgij; Venclovas, Česlovas Science, Vol. 357, Issue 6351 https://doi.org/10.1126/science.aao0100	journal	June 2017
The gRAMP CRISPR-Cas effector is an RNA endonuclease complexed with a caspase-like peptidase van Beljouw, Sam P. B.; Haagsma, Anna C.; Rodríguez-Molina, Alicia Science, Vol. 373, Issue 6561 https://doi.org/10.1126/science.abk2718	journal	September 2021
Csx28 is a membrane pore that enhances CRISPR-Cas13b–dependent antiphage defense VanderWal, Arica R.; Park, Jung-Un; Polevoda, Bogdan Science, Vol. 380, Issue 6643 https://doi.org/10.1126/science.abm1184	journal	April 2023
Daptomycin Pore Formation and Stoichiometry Depend on Membrane Potential of Target Membrane Seydlová, Gabriela; Sokol, Albert; Lišková, Petra Antimicrobial Agents and Chemotherapy, Vol. 63, Issue 1 https://doi.org/10.1128/AAC.01589-18	journal	January 2019
Correlation of Daptomycin Bactericidal Activity and Membrane Depolarization in Staphylococcus aureus Silverman, Jared A.; Perlmutter, Nancy G.; Shapiro, Howard M. Antimicrobial Agents and Chemotherapy, Vol. 47, Issue 8 https://doi.org/10.1128/AAC.47.8.2538-2544.2003	journal	August 2003
The Pinholin of Lambdoid Phage 21: Control of Lysis by Membrane Depolarization Park, Taehyun; Struck, Douglas K.; Dankenbring, Chelsey A. Journal of Bacteriology, Vol. 189, Issue 24 https://doi.org/10.1128/JB.00847-07	journal	December 2007
Nucleotide sequence and functional map of pC194, a plasmid that specifies inducible chloramphenicol resistance Horinouchi, S.; Weisblum, B. Journal of Bacteriology, Vol. 150, Issue 2 https://doi.org/10.1128/jb.150.2.815-825.1982	journal	May 1982
Abortive Infection: Bacterial Suicide as an Antiviral Immune Strategy Lopatina, Anna; Tal, Nitzan; Sorek, Rotem Annual Review of Virology, Vol. 7, Issue 1 https://doi.org/10.1146/annurev-virology-011620-040628	journal	September 2020
MUSCLE: a multiple sequence alignment method with reduced time and space complexity Edgar, Robert C. BMC Bioinformatics, Vol. 5, Issue 1, p. 113 https://doi.org/10.1186/1471-2105-5-113	journal	August 2004

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