The CRISPR-Cas immune system: biology, mechanisms and applications.

Viruses are a common threat to cellular life, not the least to bacteria and archaea who constitute the majority of life on Earth. Consequently, a variety of mechanisms to resist virus infection has evolved. A recent discovery is the adaptive immune system in prokaryotes, a type of system previously thought to be present only in vertebrates. The system, called CRISPR-Cas, provide sequence-specific adaptive immunity and fundamentally affect our understanding of virus-host interaction. CRISPR-based immunity acts by integrating short virus sequences in the cell's CRISPR locus, allowing the cell to remember, recognize and clear infections. There has been rapid advancement in our understanding of this immune system and its applications, but there are many aspects that await elucidation making the field an exciting area of research. This review provides an overview of the field and highlights unresolved issues.

[1]  Jacques Nicolas,et al.  CRISPI: a CRISPR interactive database , 2009, Bioinform..

[2]  George A. O'Toole,et al.  The CRISPR/Cas Adaptive Immune System of Pseudomonas aeruginosa Mediates Resistance to Naturally Occurring and Engineered Phages , 2012, Journal of bacteriology.

[3]  R. Terns,et al.  CRISPR-based technologies: prokaryotic defense weapons repurposed. , 2014, Trends in genetics : TIG.

[4]  J. Doudna,et al.  A Programmable Dual-RNA–Guided DNA Endonuclease in Adaptive Bacterial Immunity , 2012, Science.

[5]  N. Grishin,et al.  A putative RNA-interference-based immune system in prokaryotes: computational analysis of the predicted enzymatic machinery, functional analogies with eukaryotic RNAi, and hypothetical mechanisms of action , 2006, Biology Direct.

[6]  Konstantin Severinov,et al.  CRISPR immunity relies on the consecutive binding and degradation of negatively supercoiled invader DNA by Cascade and Cas3. , 2012, Molecular cell.

[7]  Christine L. Sun,et al.  Strong bias in the bacterial CRISPR elements that confer immunity to phage , 2013, Nature Communications.

[8]  Nick V Grishin,et al.  A DNA repair system specific for thermophilic Archaea and bacteria predicted by genomic context analysis. , 2002, Nucleic acids research.

[9]  R. Wagner,et al.  Detection and characterization of spacer integration intermediates in type I-E CRISPR–Cas system , 2014, Nucleic acids research.

[10]  Matthias Mann,et al.  Structural model of a CRISPR RNA-silencing complex reveals the RNA-target cleavage activity in Cmr4. , 2014, Molecular cell.

[11]  Joshua R. Elmore,et al.  Essential features and rational design of CRISPR RNAs that function with the Cas RAMP module complex to cleave RNAs. , 2012, Molecular cell.

[12]  Stan J. J. Brouns,et al.  Crystal structure of the CRISPR RNA–guided surveillance complex from Escherichia coli , 2014, Science.

[13]  Stan J. J. Brouns,et al.  Evolution and classification of the CRISPR–Cas systems , 2011, Nature Reviews Microbiology.

[14]  Shirley Graham,et al.  Structure of the CRISPR Interference Complex CSM Reveals Key Similarities with Cascade , 2013, Molecular cell.

[15]  Alexander Bolotin,et al.  Clustered regularly interspaced short palindrome repeats (CRISPRs) have spacers of extrachromosomal origin. , 2005, Microbiology.

[16]  S. Ehrlich,et al.  Clustered regularly interspaced short palindrome repeats (CRISPRs) have spacers of extrachromosomal origin. , 2005, Microbiology.

[17]  Dipali G. Sashital,et al.  Mechanism of foreign DNA selection in a bacterial adaptive immune system. , 2012, Molecular cell.

[18]  David S. Weiss,et al.  A CRISPR-CAS System Mediates Bacterial Innate Immune Evasion and Virulence , 2013, Nature.

[19]  K. Makino,et al.  Nucleotide sequence of the iap gene, responsible for alkaline phosphatase isozyme conversion in Escherichia coli, and identification of the gene product , 1987, Journal of bacteriology.

[20]  Asaf Levy,et al.  CRISPR adaptation biases explain preference for acquisition of foreign DNA , 2015, Nature.

[21]  L. Marraffini,et al.  CRISPR Interference Limits Horizontal Gene Transfer in Staphylococci by Targeting DNA , 2008, Science.

[22]  Peter C. Fineran,et al.  Cytotoxic Chromosomal Targeting by CRISPR/Cas Systems Can Reshape Bacterial Genomes and Expel or Remodel Pathogenicity Islands , 2013, PLoS genetics.

[23]  Peter C. Fineran,et al.  Memory of viral infections by CRISPR-Cas adaptive immune systems: acquisition of new information. , 2012, Virology.

[24]  Jennifer A. Doudna,et al.  Structures of the RNA-guided surveillance complex from a bacterial immune system , 2011, Nature.

[25]  Elo Leung,et al.  Targeted Genome Editing Across Species Using ZFNs and TALENs , 2011, Science.

[26]  Konstantin Severinov,et al.  Interference by clustered regularly interspaced short palindromic repeat (CRISPR) RNA is governed by a seed sequence , 2011, Proceedings of the National Academy of Sciences.

[27]  R. Barrangou,et al.  CRISPR Provides Acquired Resistance Against Viruses in Prokaryotes , 2007, Science.

[28]  Rolf Wagner,et al.  Identification and characterization of E. coli CRISPR‐cas promoters and their silencing by H‐NS , 2010, Molecular microbiology.

[29]  E. Koonin,et al.  A Novel Family of Sequence-specific Endoribonucleases Associated with the Clustered Regularly Interspaced Short Palindromic Repeats* , 2008, Journal of Biological Chemistry.

[30]  Michael S. Spilman,et al.  Structure of an RNA silencing complex of the CRISPR-Cas immune system. , 2013, Molecular cell.

[31]  Alan R. Davidson,et al.  Bacteriophage genes that inactivate the CRISPR/Cas bacterial immune system , 2012, Nature.

[32]  Albert J R Heck,et al.  Structural basis for CRISPR RNA-guided DNA recognition by Cascade , 2011, Nature Structural &Molecular Biology.

[33]  Jennifer A. Doudna,et al.  Integrase-mediated spacer acquisition during CRISPR–Cas adaptive immunity , 2015, Nature.

[34]  H. Xiang,et al.  Adaptation of the Haloarcula hispanica CRISPR-Cas system to a purified virus strictly requires a priming process , 2013, Nucleic acids research.

[35]  Luciano A. Marraffini,et al.  Conditional tolerance of temperate phages via transcription-dependent CRISPR-Cas targeting , 2014, Nature.

[36]  Jennifer A. Doudna,et al.  A prudent path forward for genomic engineering and germline gene modification , 2015, Science.

[37]  Rodolphe Barrangou,et al.  Novel Virulence Gene and Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR) Multilocus Sequence Typing Scheme for Subtyping of the Major Serovars of Salmonella enterica subsp. enterica , 2011, Applied and Environmental Microbiology.

[38]  J. García-Martínez,et al.  Intervening Sequences of Regularly Spaced Prokaryotic Repeats Derive from Foreign Genetic Elements , 2005, Journal of Molecular Evolution.

[39]  R. Carlson.  Planetary science: A new recipe for Earth formation , 2015, Nature.

[40]  Gang Bao,et al.  Seamless modification of wild-type induced pluripotent stem cells to the natural CCR5Δ32 mutation confers resistance to HIV infection , 2014, Proceedings of the National Academy of Sciences.

[41]  Emmanuelle Charpentier,et al.  The tracrRNA and Cas9 families of type II CRISPR-Cas immunity systems , 2013, RNA biology.

[42]  Anders F. Andersson,et al.  Virus Population Dynamics and Acquired Virus Resistance in Natural Microbial Communities , 2008, Science.

[43]  Chase L. Beisel,et al.  Repurposing endogenous type I CRISPR-Cas systems for programmable gene repression , 2014, Nucleic acids research.

[44]  G. O’Toole,et al.  Interaction between Bacteriophage DMS3 and Host CRISPR Region Inhibits Group Behaviors of Pseudomonas aeruginosa , 2008, Journal of bacteriology.

[45]  Albert J R Heck,et al.  Structure and activity of the RNA-targeting Type III-B CRISPR-Cas complex of Thermus thermophilus. , 2013, Molecular cell.

[46]  Jos Boekhorst,et al.  Degenerate target sites mediate rapid primed CRISPR adaptation , 2014, Proceedings of the National Academy of Sciences.

[47]  D van Soolingen,et al.  Simultaneous detection and strain differentiation of Mycobacterium tuberculosis for diagnosis and epidemiology , 1997, Journal of clinical microbiology.

[48]  Eugene V Koonin,et al.  CRISPR-Cas , 2013, RNA biology.

[49]  Hao Yin,et al.  Genome editing with Cas9 in adult mice corrects a disease mutation and phenotype , 2014, Nature Biotechnology.

[50]  James E. DiCarlo,et al.  RNA-Guided Human Genome Engineering via Cas9 , 2013, Science.

[51]  C. Suttle.  Viruses in the sea , 2005, Nature.

[52]  J. Fu,et al.  Harnessing the clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated Cas9 system to disrupt the hepatitis B virus , 2015, Gene Therapy.

[53]  Joshua R. Elmore,et al.  Essential structural and functional roles of the Cmr4 subunit in RNA cleavage by the Cmr CRISPR-Cas complex. , 2014, Cell reports.

[54]  L. Schouls,et al.  Identification of a novel family of sequence repeats among prokaryotes. , 2002, Omics : a journal of integrative biology.

[55]  Erik J. Sontheimer,et al.  Self vs. non-self discrimination during CRISPR RNA-directed immunity , 2009, Nature.

[56]  Chad W. Euler,et al.  Exploiting CRISPR-Cas nucleases to produce sequence-specific antimicrobials , 2014, Nature Biotechnology.

[57]  R. Barrangou,et al.  Subtyping Salmonella enterica Serovar Enteritidis Isolates from Different Sources by Using Sequence Typing Based on Virulence Genes and Clustered Regularly Interspaced Short Palindromic Repeats (CRISPRs) , 2011, Applied and Environmental Microbiology.

[58]  E. Olson,et al.  Prevention of muscular dystrophy in mice by CRISPR/Cas9–mediated editing of germline DNA , 2014, Science.

[59]  Peter C. Fineran,et al.  Priming in the Type I-F CRISPR-Cas system triggers strand-independent spacer acquisition, bi-directionally from the primed protospacer , 2014, Nucleic acids research.

[60]  J. Banfield,et al.  Rapidly evolving CRISPRs implicated in acquired resistance of microorganisms to viruses. , 2007, Environmental microbiology.

[61]  Jennifer A. Doudna,et al.  DNA interrogation by the CRISPR RNA-guided endonuclease Cas9 , 2014, Nature.

[62]  Philippe Horvath,et al.  Phage Response to CRISPR-Encoded Resistance in Streptococcus thermophilus , 2007, Journal of bacteriology.

[63]  Rolf Backofen,et al.  Essential requirements for the detection and degradation of invaders by the Haloferax volcanii CRISPR/Cas system I-B , 2013, RNA biology.

[64]  J. García-Martínez,et al.  Short motif sequences determine the targets of the prokaryotic CRISPR defence system. , 2009, Microbiology.

[65]  Samuel H Sternberg,et al.  CasA mediates Cas3-catalyzed target degradation during CRISPR RNA-guided interference , 2014, Proceedings of the National Academy of Sciences.

[66]  Philippe Horvath,et al.  The CRISPR/Cas bacterial immune system cleaves bacteriophage and plasmid DNA , 2010, Nature.

[67]  Marko Djordjevic,et al.  Transcription, processing and function of CRISPR cassettes in Escherichia coli , 2010, Molecular microbiology.

[68]  Albert J R Heck,et al.  RNA targeting by the type III-A CRISPR-Cas Csm complex of Thermus thermophilus. , 2014, Molecular cell.

[69]  Andrew Camilli,et al.  A bacteriophage encodes its own CRISPR/Cas adaptive response to evade host innate immunity , 2013, Nature.

[70]  Shiraz A. Shah,et al.  CRISPR families of the crenarchaeal genus Sulfolobus: bidirectional transcription and dynamic properties , 2009, Molecular microbiology.

[71]  Hongwei Wang,et al.  Cas5d protein processes pre-crRNA and assembles into a cascade-like interference complex in subtype I-C/Dvulg CRISPR-Cas system. , 2012, Structure.

[72]  Magnus Lundgren,et al.  Efficient programmable gene silencing by Cascade , 2014, Nucleic acids research.

[73]  Xu Peng,et al.  A novel interference mechanism by a type IIIB CRISPR‐Cmr module in Sulfolobus , 2013, Molecular microbiology.

[74]  Henning Urlaub,et al.  In vitro assembly and activity of an archaeal CRISPR-Cas type I-A Cascade interference complex , 2014, Nucleic acids research.

[75]  Yanli Wang,et al.  Crystal structure of the RNA-guided immune surveillance Cascade complex in Escherichia coli , 2014, Nature.

[76]  N. Cianciotto,et al.  The CRISPR-Associated Gene cas2 of Legionella pneumophila Is Required for Intracellular Infection of Amoebae , 2013, mBio.

[77]  P. Groenen,et al.  Nature of DNA polymorphism in the direct repeat cluster of Mycobacterium tuberculosis; application for strain differentiation by a novel typing method , 1993, Molecular microbiology.

[78]  Q. She,et al.  An archaeal CRISPR type III-B system exhibiting distinctive RNA targeting features and mediating dual RNA and DNA interference , 2014, Nucleic acids research.

[79]  Timothy K Lu,et al.  Sequence-specific antimicrobials using efficiently delivered RNA-guided nucleases , 2014, Nature Biotechnology.

[80]  K. Datsenko,et al.  CRISPR RNA binding and DNA target recognition by purified Cascade complexes from Escherichia coli , 2014, Nucleic acids research.

[81]  J. Vogel,et al.  CRISPR RNA maturation by trans-encoded small RNA and host factor RNase III , 2011, Nature.

[82]  David A. Scott,et al.  In vivo genome editing using Staphylococcus aureus Cas9 , 2015, Nature.

[83]  Hiroshi Nishimasu.  Crystal Structure of Cas9 , 2015 .

[84]  U. Qimron,et al.  Proteins and DNA elements essential for the CRISPR adaptation process in Escherichia coli , 2012, Nucleic acids research.

[85]  S. Mulepati,et al.  Crystal structure of a CRISPR RNA–guided surveillance complex bound to a ssDNA target , 2014, Science.

[86]  Luciano A. Marraffini,et al.  Cas9 specifies functional viral targets during CRISPR-Cas adaptation , 2015, Nature.

[87]  Le Cong,et al.  Multiplex Genome Engineering Using CRISPR/Cas Systems , 2013, Science.

[88]  G Vergnaud,et al.  CRISPR elements in Yersinia pestis acquire new repeats by preferential uptake of bacteriophage DNA, and provide additional tools for evolutionary studies. , 2005, Microbiology.

[89]  Gang Bao,et al.  CRISPR/Cas9 systems have off-target activity with insertions or deletions between target DNA and guide RNA sequences , 2014, Nucleic acids research.

[90]  K. Zhou,et al.  Structural basis for DNase activity of a conserved protein implicated in CRISPR-mediated genome defense. , 2009, Structure.

[91]  Quincy Teng,et al.  Structural Biology , 2013, Springer US.

[92]  Jennifer A. Doudna,et al.  Cas1–Cas2 complex formation mediates spacer acquisition during CRISPR–Cas adaptive immunity , 2014, Nature Structural &Molecular Biology.

[93]  E. Lander,et al.  Genetic Screens in Human Cells Using the CRISPR-Cas9 System , 2013, Science.

[94]  Stan J. J. Brouns,et al.  Small CRISPR RNAs Guide Antiviral Defense in Prokaryotes , 2008, Science.

[95]  Fan Yang,et al.  RNA-directed gene editing specifically eradicates latent and prevents new HIV-1 infection , 2014, Proceedings of the National Academy of Sciences.

[96]  Jing Zhang,et al.  Structure and mechanism of the CMR complex for CRISPR-mediated antiviral immunity. , 2012, Molecular cell.

[97]  Ibtissem Grissa,et al.  CRISPRFinder: a web tool to identify clustered regularly interspaced short palindromic repeats , 2007, Nucleic Acids Res..

[98]  Luke A. Gilbert,et al.  Repurposing CRISPR as an RNA-Guided Platform for Sequence-Specific Control of Gene Expression , 2013, Cell.

[99]  Stan J. J. Brouns,et al.  CRISPR Interference Directs Strand Specific Spacer Acquisition , 2012, PloS one.

[100]  E. Koonin,et al.  Evolution of adaptive immunity from transposable elements combined with innate immune systems , 2014, Nature Reviews Genetics.

[101]  Andrey P. Anisimov,et al.  Insight into Microevolution of Yersinia pestis by Clustered Regularly Interspaced Short Palindromic Repeats , 2008, PloS one.

[102]  R. Terns,et al.  Sequences spanning the leader-repeat junction mediate CRISPR adaptation to phage in Streptococcus thermophilus , 2015, Nucleic acids research.

[103]  Q. She,et al.  Transcriptional regulator-mediated activation of adaptation genes triggers CRISPR de novo spacer acquisition , 2015, Nucleic acids research.

[104]  M. DeLisa,et al.  Envelope stress is a trigger of CRISPR RNA‐mediated DNA silencing in Escherichia coli , 2011, Molecular Microbiology.

[105]  Hans Clevers,et al.  Functional repair of CFTR by CRISPR/Cas9 in intestinal stem cell organoids of cystic fibrosis patients. , 2013, Cell stem cell.

[106]  Feng Zhang,et al.  Programmable repression and activation of bacterial gene expression using an engineered CRISPR-Cas system , 2013, Nucleic acids research.

[107]  L. Marraffini,et al.  Mature clustered, regularly interspaced, short palindromic repeats RNA (crRNA) length is measured by a ruler mechanism anchored at the precursor processing site , 2011, Proceedings of the National Academy of Sciences.

[108]  Rodolphe Barrangou,et al.  The Population and Evolutionary Dynamics of Phage and Bacteria with CRISPR–Mediated Immunity , 2013, PLoS genetics.

[109]  B. Graveley,et al.  RNA-Guided RNA Cleavage by a CRISPR RNA-Cas Protein Complex , 2009, Cell.

[110]  Luke A. Gilbert,et al.  CRISPR-Mediated Modular RNA-Guided Regulation of Transcription in Eukaryotes , 2013, Cell.

[111]  Konstantin Severinov,et al.  Molecular memory of prior infections activates the CRISPR/Cas adaptive bacterial immunity system , 2012, Nature Communications.

[112]  Jing Zhang,et al.  CRISPR-mediated targeted mRNA degradation in the archaeon Sulfolobus solfataricus , 2014, Nucleic acids research.

[113]  Eugene V Koonin,et al.  Unification of Cas protein families and a simple scenario for the origin and evolution of CRISPR-Cas systems , 2011, Biology Direct.

[114]  Andrew Emili,et al.  A dual function of the CRISPR–Cas system in bacterial antivirus immunity and DNA repair , 2011, Molecular microbiology.

[115]  R. Terns,et al.  Cas9 function and host genome sampling in Type II-A CRISPR–Cas adaptation , 2015, Genes & development.

[116]  Martin J. Aryee,et al.  GUIDE-Seq enables genome-wide profiling of off-target cleavage by CRISPR-Cas nucleases , 2014, Nature Biotechnology.

[117]  Philippe Horvath,et al.  Cas3 is a single‐stranded DNA nuclease and ATP‐dependent helicase in the CRISPR/Cas immune system , 2011, The EMBO journal.

[118]  S. Moineau,et al.  CRISPR-Cas and restriction–modification systems are compatible and increase phage resistance , 2013, Nature Communications.

[119]  H. Endtz,et al.  A novel link between Campylobacter jejuni bacteriophage defence, virulence and Guillain–Barré syndrome , 2012, European Journal of Clinical Microbiology & Infectious Diseases.

[120]  R. Garrett,et al.  Selective and hyperactive uptake of foreign DNA by adaptive immune systems of an archaeon via two distinct mechanisms , 2012, Molecular microbiology.

[121]  I. Mokrousov,et al.  Efficient Discrimination within a Corynebacterium diphtheriae Epidemic Clonal Group by a Novel Macroarray-Based Method , 2005, Journal of Clinical Microbiology.

[122]  Ceslovas Venclovas,et al.  Programmable RNA shredding by the type III-A CRISPR-Cas system of Streptococcus thermophilus. , 2014, Molecular cell.

[123]  Yarden Katz,et al.  Multiplexed activation of endogenous genes by CRISPR-on, an RNA-guided transcriptional activator system , 2013, Cell Research.

[124]  Pascale Cossart,et al.  Identification of new noncoding RNAs in Listeria monocytogenes and prediction of mRNA targets , 2007, Nucleic acids research.

[125]  Alan R. Davidson,et al.  A New Group of Phage Anti-CRISPR Genes Inhibits the Type I-E CRISPR-Cas System of Pseudomonas aeruginosa , 2014, mBio.

[126]  T. Lu,et al.  Tunable and Multifunctional Eukaryotic Transcription Factors Based on CRISPR/Cas , 2013, ACS synthetic biology.

[127]  Alexandro E. Trevino,et al.  Genome-scale transcriptional activation by an engineered CRISPR-Cas9 complex , 2014, Nature.

[128]  R. Barrangou,et al.  The three major types of CRISPR‐Cas systems function independently in CRISPR RNA biogenesis in Streptococcus thermophilus , 2014, Molecular microbiology.

[129]  Wei Tang,et al.  Correction of a genetic disease in mouse via use of CRISPR-Cas9. , 2013, Cell stem cell.

[130]  Albert J R Heck,et al.  RNA-guided complex from a bacterial immune system enhances target recognition through seed sequence interactions , 2011, Proceedings of the National Academy of Sciences.

[131]  Benjamin L. Oakes,et al.  Programmable RNA recognition and cleavage by CRISPR/Cas9 , 2014, Nature.

[132]  Lei Wang,et al.  Generation of Gene-Modified Cynomolgus Monkey via Cas9/RNA-Mediated Gene Targeting in One-Cell Embryos , 2014, Cell.

[133]  Jennifer A. Doudna,et al.  Structures of Cas9 Endonucleases Reveal RNA-Mediated Conformational Activation , 2014, Science.

[134]  K. Ye,et al.  Cmr4 is the slicer in the RNA-targeting Cmr CRISPR complex , 2014, Nucleic acids research.