scholarly journals Competition between mobile genetic elements drives optimization of a phage-encoded CRISPR-Cas system: insights from a natural arms race

2019 ◽  
Vol 374 (1772) ◽  
pp. 20180089 ◽  
Author(s):  
Amelia C. McKitterick ◽  
Kristen N. LeGault ◽  
Angus Angermeyer ◽  
Munirul Alam ◽  
Kimberley D. Seed
Keyword(s):  
High Throughput Sequencing ◽  
Mobile Genetic Element ◽  
Arms Race ◽  
Type I ◽  
Helicase Activity ◽  
Wild Populations ◽  
Immune Systems ◽  
Adaptive Immune ◽  
Adaptive Immune Systems

CRISPR-Cas systems function as adaptive immune systems by acquiring nucleotide sequences called spacers that mediate sequence-specific defence against competitors. Uniquely, the phage ICP1 encodes a Type I-F CRISPR-Cas system that is deployed to target and overcome PLE, a mobile genetic element with anti-phage activity in Vibrio cholerae . Here, we exploit the arms race between ICP1 and PLE to examine spacer acquisition and interference under laboratory conditions to reconcile findings from wild populations. Natural ICP1 isolates encode multiple spacers directed against PLE, but we find that single spacers do not interfere equally with PLE mobilization. High-throughput sequencing to assay spacer acquisition reveals that ICP1 can also acquire spacers that target the V. cholerae chromosome. We find that targeting the V. cholerae chromosome proximal to PLE is sufficient to block PLE and is dependent on Cas2-3 helicase activity. We propose a model in which indirect chromosomal spacers are able to circumvent PLE by Cas2-3-mediated processive degradation of the V. cholerae chromosome before PLE mobilization. Generally, laboratory-acquired spacers are much more diverse than the subset of spacers maintained by ICP1 in nature, showing how evolutionary pressures can constrain CRISPR-Cas targeting in ways that are often not appreciated through in vitro analyses. This article is part of a discussion meeting issue ‘The ecology and evolution of prokaryotic CRISPR-Cas adaptive immune systems’.

scholarly journals Competition between mobile genetic elements drives optimization of a phage-encoded CRISPR-Cas system: Insights from a natural arms-race

2018 ◽  
Author(s):  
Amelia C. McKitterick ◽  
Kristen N. LeGault ◽  
Angus Angermeyer ◽  
Muniral Alam ◽  
Kimberley D. Seed
Keyword(s):  
High Throughput Sequencing ◽  
Mobile Genetic Element ◽  
Arms Race ◽  
Type I ◽  
Genetic Element ◽  
Wild Populations ◽  
Adaptive Immune ◽  
Adaptive Immune Systems ◽  
Phage Activity

AbstractCRISPR-Cas systems function as adaptive immune systems by acquiring nucleotide sequences called spacers that mediate sequence-specific defense against competitors. Uniquely, the phage ICP1 encodes a Type I-F CRISPR-Cas system that is deployed to target and overcome PLE, a mobile genetic element with anti-phage activity in Vibrio cholerae. Here, we exploit the arms race between ICP1 and PLE to examine spacer acquisition and interference under laboratory conditions to reconcile findings from wild populations. Natural ICP1 isolates encode multiple spacers directed against PLE, but we find that single spacers do not equally interfere with PLE mobilization. High-throughput sequencing to assay spacer acquisition reveals that ICP1 can also acquire spacers that target the V. cholerae chromosome. We find that targeting the V. cholerae chromosome proximal to PLE is sufficient to block PLE and propose a model in which indirect chromosomal spacers are able to circumvent PLE by Cas2-3-mediated processive degradation of the V. cholerae chromosome before PLE mobilization. Generally, laboratory acquired spacers are much more diverse than the subset of spacers maintained by ICP1 in nature, showing how evolutionary pressures can constrain CRISPR-Cas targeting in ways that are often not appreciated through in vitro analyses.

scholarly journals CRISPR-Cas immunity repressed by a biofilm-activating pathway inPseudomonas aeruginosa

2019 ◽  
Author(s):  
Adair L. Borges ◽  
Bardo Castro ◽  
Sutharsan Govindarajan ◽  
Tina Solvik ◽  
Veronica Escalante ◽  
...  
Keyword(s):  
Genetic Screen ◽  
Type I ◽  
Two Component System ◽  
Component System ◽  
Forward Genetic Screen ◽  
Immune Systems ◽  
Adaptive Immune ◽  
Biofilm Production ◽  
Two Component ◽  
Adaptive Immune Systems

CRISPR-Cas systems are adaptive immune systems that protect bacteria from bacteriophage (phage) infection. To provide immunity, RNA-guided protein surveillance complexes recognize foreign nucleic acids, triggering their destruction by Cas nucleases. While the essential requirements for immune activity are well understood, the physiological cues that regulate CRISPR-Cas expression are not. Here, a forward genetic screen identifies a two-component system (KinB/AlgB), previously characterized in regulatingPseudomonas aeruginosavirulence and biofilm establishment, as a regulator of the biogenesis and activity of the Type I-F CRISPR-Cas system. Downstream of the KinB/AlgB system, activators of biofilm production AlgU (a σEorthologue) and AlgR, act as repressors of CRISPR-Cas activity during planktonic and surface-associated growth. AmrZ, another biofilm activator, functions as a surface-specific repressor of CRISPR-Cas immunity.Pseudomonasphages and plasmids have taken advantage of this regulatory scheme, and carry hijacked homologs of AmrZ, which are functional CRISPR-Cas repressors. This suggests that while CRISPR-Cas regulation may be important to limit self-toxicity, endogenous repressive pathways represent a vulnerability for parasite manipulation.

scholarly journals Structural basis for inhibition of the type I-F CRISPR–Cas surveillance complex by AcrIF4, AcrIF7 and AcrIF14

2020 ◽  
Author(s):  
Clinton Gabel ◽  
Zhuang Li ◽  
Heng Zhang ◽  
Leifu Chang
Keyword(s):  
Genome Editing ◽  
Conformational Changes ◽  
Structural Basis ◽  
Type I ◽  
Structural Detail ◽  
Immune Systems ◽  
Helical Bundle ◽  
Adaptive Immune ◽  
Target Dna ◽  
Adaptive Immune Systems

Abstract CRISPR–Cas systems are adaptive immune systems in bacteria and archaea to defend against mobile genetic elements (MGEs) and have been repurposed as genome editing tools. Anti-CRISPR (Acr) proteins are produced by MGEs to counteract CRISPR–Cas systems and can be used to regulate genome editing by CRISPR techniques. Here, we report the cryo-EM structures of three type I-F Acr proteins, AcrIF4, AcrIF7 and AcrIF14, bound to the type I-F CRISPR–Cas surveillance complex (the Csy complex) from Pseudomonas aeruginosa. AcrIF4 binds to an unprecedented site on the C-terminal helical bundle of Cas8f subunit, precluding conformational changes required for activation of the Csy complex. AcrIF7 mimics the PAM duplex of target DNA and is bound to the N-terminal DNA vise of Cas8f. Two copies of AcrIF14 bind to the thumb domains of Cas7.4f and Cas7.6f, preventing hybridization between target DNA and the crRNA. Our results reveal structural detail of three AcrIF proteins, each binding to a different site on the Csy complex for inhibiting degradation of MGEs.

scholarly journals Why put up with immunity when there is resistance: an excursion into the population and evolutionary dynamics of restriction–modification and CRISPR-Cas

2019 ◽  
Vol 374 (1772) ◽  
pp. 20180096 ◽  
Author(s):  
James Gurney ◽  
Maroš Pleška ◽  
Bruce R. Levin
Keyword(s):  
Evolutionary Dynamics ◽  
Arms Race ◽  
Lytic Phage ◽  
Bacterial Populations ◽  
Immune Systems ◽  
Adaptive Immune ◽  
Escape Mutants ◽  
Adaptive Immune Systems ◽  
Restriction Modification ◽  
Coevolutionary Arms Race

Bacteria can readily generate mutations that prevent bacteriophage (phage) adsorption and thus make bacteria resistant to infections with these viruses. Nevertheless, the majority of bacteria carry complex innate and/or adaptive immune systems: restriction–modification (RM) and CRISPR-Cas, respectively. Both RM and CRISPR-Cas are commonly assumed to have evolved and be maintained to protect bacteria from succumbing to infections with lytic phage. Using mathematical models and computer simulations, we explore the conditions under which selection mediated by lytic phage will favour such complex innate and adaptive immune systems, as opposed to simple envelope resistance. The results of our analysis suggest that when populations of bacteria are confronted with lytic phage: (i) In the absence of immunity, resistance to even multiple bacteriophage species with independent receptors can evolve readily. (ii) RM immunity can benefit bacteria by preventing phage from invading established bacterial populations and particularly so when there are multiple bacteriophage species adsorbing to different receptors. (iii) Whether CRISPR-Cas immunity will prevail over envelope resistance depends critically on the number of steps in the coevolutionary arms race between the bacteria-acquiring spacers and the phage-generating CRISPR-escape mutants. We discuss the implications of these results in the context of the evolution and maintenance of RM and CRISPR-Cas and highlight fundamental questions that remain unanswered. This article is part of a discussion meeting issue ‘The ecology and evolution of prokaryotic CRISPR-Cas adaptive immune systems’.

scholarly journals Mesenchymal Stem Cells Regulate the Innate and Adaptive Immune Responses Dampening Arthritis Progression

2016 ◽  
Vol 2016 ◽  
pp. 1-10 ◽  
Author(s):  
R. A. Contreras ◽  
F. E. Figueroa ◽  
F. Djouad ◽  
P. Luz-Crawford
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Therapeutic Potential ◽  
Multipotent Stem Cells ◽  
Immune Systems ◽  
Adaptive Immune ◽  
Adaptive Immune Systems

Mesenchymal stem cells (MSCs) are multipotent stem cells that are able to immunomodulate cells from both the innate and the adaptive immune systems promoting an anti-inflammatory environment. During the last decade, MSCs have been intensively studiedin vitroandin vivoin experimental animal model of autoimmune and inflammatory disorders. Based on these studies, MSCs are currently widely used for the treatment of autoimmune diseases such as rheumatoid arthritis (RA) characterized by complex deregulation of the immune systems. However, the therapeutic properties of MSCs in arthritis are still controverted. These controversies might be due to the diversity of MSC sources and isolation protocols used, the time, the route and dose of MSC administration, the variety of the mechanisms involved in the MSCs suppressive effects, and the complexity of arthritis pathogenesis. In this review, we discuss the role of the interactions between MSCs and the different immune cells associated with arthritis pathogenesis and the possible means described in the literature that could enhance MSCs therapeutic potential counteracting arthritis development and progression.

scholarly journals Distinct subcellular localization of a Type I CRISPR complex and the Cas3 nuclease in bacteria

2020 ◽  
Author(s):  
Sutharsan Govindarajan ◽  
Adair Borges ◽  
Joseph Bondy-Denomy
Keyword(s):  
Fusion Proteins ◽  
Cell Biology ◽  
Type I ◽  
Immune Systems ◽  
Adaptive Immune ◽  
Spatiotemporal Regulation ◽  
Cas9 Nuclease ◽  
Adaptive Immune Systems ◽  
Single Focus

AbstractCRISPR-Cas systems are prokaryotic adaptive immune systems that have been well characterized biochemically, but in vivo spatiotemporal regulation and cell biology remains largely unaddressed. Here, we used fluorescent fusion proteins to study the localization of the Type I-F CRISPR-Cas system native to Pseudomonas aeruginosa. When targeted to an integrated prophage, the crRNA-guided (Csy) complex and a majority of Cas3 molecules in the cell are recruited to a single focus. When lacking a target in the cell, however, the Csy complex is broadly nucleoid bound, while Cas3 is diffuse in the cytoplasm. Nucleoid association for the Csy proteins is crRNA-dependent, and inhibited by expression of anti-CRISPR AcrIF2, which blocks PAM binding. The Cas9 nuclease is also nucleoid localized, only when gRNA-bound, which is abolished by PAM mimic, AcrIIA4. Our findings reveal PAM-dependent nucleoid surveillance and spatiotemporal regulation in Type I CRISPR-Cas that separates the nuclease-helicase Cas3 from the crRNA-guided surveillance complex.

scholarly journals CRISPRs for strain tracking and its application to microbiota transplantation data analysis

2018 ◽  
Author(s):  
Tony J. Lam ◽  
Yuzhen Ye
Keyword(s):  
Data Analysis ◽  
General Purpose ◽  
Arms Race ◽  
Donor Strain ◽  
Fecal Transplantation ◽  
Immune Systems ◽  
Adaptive Immune ◽  
Microbiome Data ◽  
Adaptive Immune Systems ◽  
Potential Use

AbstractCRISPR-Cas systems are adaptive immune systems naturally found in bacteria and archaea. Bacteria and archaea use these systems to defend against invaders, including phages, plasmids and other mobile genetic elements. Relying on integration of invader sequences (protospacers) into CRISPR loci (forming spacers flanked by repeats), CRISPR-Cas systems store genetic memory of past invasions. While CRISPR-Cas systems have evolved in response to invading mobile elements, invaders have also developed mechanisms to avoid detection. As a result of arms-race between CRISPR-Cas systems and their targets, the CRISPR arrays typically undergo rapid turnover of the spacers with removal of old spacers and acquisition of new ones. Additionally, different individuals rarely share spacers amongst their microbiome. In this paper, we developed a pipeline (called CRISPRtrack) for strain tracking based on CRISPR spacer content, and applied it to fecal transplantation microbiome data to study the retention of donor strains in recipients. Our results demonstrate the potential use of CRISPRs as a simple yet effective tool for donor strain tracking in fecal transplantation, and also as a general purpose tool for quantifying microbiome similarity.

scholarly journals Why Put-Up with Immunity when there is Resistance: An Excursion into the Population and Evolutionary Dynamics of Restriction-Modification and CRISPR-Cas

2018 ◽  
Author(s):  
James R Gurney ◽  
Maros Pleska ◽  
Bruce R Levin
Keyword(s):  
Computer Simulations ◽  
Evolutionary Dynamics ◽  
Arms Race ◽  
Lytic Phage ◽  
Bacterial Populations ◽  
Immune Systems ◽  
Adaptive Immune ◽  
Escape Mutants ◽  
Adaptive Immune Systems ◽  
Restriction Modification

Bacteria can readily generate mutations that prevent bacteriophage (phage) adsorption and thus make bacteria resistant to infections with these viruses. Nevertheless, the majority of bacteria carry complex innate and/or adaptive immune systems: restriction-modification (RM) and CRISPR-Cas, respectively. Both RM and CRISPR-Cas are commonly assumed to have evolved and be maintained to protect bacteria from succumbing to infections with lytic phage. Using mathematical models and computer simulations, we explore the conditions, under which selection mediated by lytic phage will favor such complex innate and adaptive immune systems, as opposed to simple envelope resistance. The results of our analysis suggest that when populations of bacteria are confronted with lytic phage: (i) In the absence of immunity, resistance to even multiple bacteriophage species with independent receptors can evolve readily. (ii) RM immunity can benefit bacteria by preventing phage from invading established bacterial populations and particularly so when there are multiple bacteriophage species adsorbing to different receptors. (iii) Whether CRISPR-Cas immunity will prevail over envelope resistance depends critically on the length of the co-evolutionary arms race between the bacteria acquiring spacers and the phage generating CRISPR-escape mutants. We discuss the implications of these results in the context of the evolution and maintenance of RM and CRISPR-Cas and highlight fundamental questions that remain unanswered.

Phage-Encoded Anti-CRISPR Defenses

2018 ◽  
Vol 52 (1) ◽  
pp. 445-464 ◽  
Author(s):  
Sabrina Y. Stanley ◽  
Karen L. Maxwell
Keyword(s):  
Mechanisms Of Action ◽  
Arms Race ◽  
Phage Infection ◽  
Bacterial Evolution ◽  
Immune Systems ◽  
Adaptive Immune ◽  
Evolutionary Origins ◽  
Potential Impact ◽  
The Impact ◽  
Adaptive Immune Systems

The battle for survival between bacteria and bacteriophages (phages) is an arms race where bacteria develop defenses to protect themselves from phages and phages evolve counterstrategies to bypass these defenses. CRISPR-Cas adaptive immune systems represent a widespread mechanism by which bacteria protect themselves from phage infection. In response to CRISPR-Cas, phages have evolved protein inhibitors known as anti-CRISPRs. Here, we describe the discovery and mechanisms of action of anti-CRISPR proteins. We discuss the potential impact of anti-CRISPRs on bacterial evolution, speculate on their evolutionary origins, and contemplate the possible next steps in the CRISPR-Cas evolutionary arms race. We also touch on the impact of anti-CRISPRs on the development of CRISPR-Cas-based biotechnological tools.

scholarly journals Structure reveals mechanism of CRISPR RNA-guided nuclease recruitment and anti-CRISPR viral mimicry

2018 ◽  
Author(s):  
MaryClare F. Rollins ◽  
Saikat Chowdhury ◽  
Joshua Carter ◽  
Sarah M. Golden ◽  
Heini M. Miettinen ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Type I ◽  
Structural Homology ◽  
Immune Systems ◽  
Helical Bundle ◽  
Adaptive Immune ◽  
Double Stranded Dna ◽  
Viral Mimicry ◽  
Cas Genes ◽  
Adaptive Immune Systems

AbstractBacteria and archaea have evolved sophisticated adaptive immune systems that rely on CRISPR RNA (crRNA)-guided detection and nuclease-mediated elimination of invading nucleic acids. Here we present the cryo-EM structure of the type I-F CRISPR RNA-guided surveillance complex (Csy complex) from Pseudomonas aeruginosa bound to a double-stranded DNA target. Comparison of this structure to previously determined structures of this complex reveals a Ȉ180-degree rotation of the C-terminal helical bundle on the “large” Cas8f subunit. We show that the dsDNA-induced conformational change in Cas8f exposes a Cas2/3 “nuclease recruitment helix” that is structurally homologous to a virally encoded anti-CRISPR protein (AcrIF3). Structural homology between Cas8f and AcrIF3 suggests that AcrIF3 is a mimic of the Cas8f “nuclease recruitment helix”, implying that cas genes may sometimes serve as genetic fodder for the evolution of anti-CRISPRs.

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