Non-canonical inhibition strategies and structural basis of anti-CRISPR proteins targeting type I CRISPR-Cas systems

Yin, Peng; Zhang, Yi; Yang, Lingguang; Feng, Yue

doi:10.1016/j.jmb.2023.167996

Cited by 14 publications

(6 citation statements)

References 89 publications

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“…Phage immune evasion proteins are categorized and exhibited as colored dots colored according to the key below. Notable evasion proteins are indicated with text labels [23][24][25][26][27]40,[47][48][49][50] . , Schematic model of GajAB-Gad1 co-complex formation and domain organization of phage Phi3T Gad1.…”

Section: Extended Referencesmentioning

confidence: 99%

Structural basis of Gabija anti-phage defense and viral immune evasion

Antine

Johnson

Mooney

et al. 2023

Preprint

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Bacteria encode hundreds of diverse defense systems that protect from viral infection and inhibit phage propagation. Gabija is one of the most prevalent anti-phage defense systems, occurring in >15% of all sequenced bacterial and archaeal genomes, but the molecular basis of how Gabija defends cells from viral infection remains poorly understood. Here we use X-ray crystallography and cryo-EM to define how Gabija proteins assemble into an ~500 kDa supramolecular complex that degrades phage DNA. Gabija protein A (GajA) is a DNA endonuclease that tetramerizes to form the core of the anti-phage defense complex. Two sets of Gabija protein B (GajB) dimers dock at opposite sides of the complex and create a 4:4 GajAB assembly that is essential for phage resistance in vivo. We show that a phage-encoded protein Gabija anti-defense 1 (Gad1) directly binds the Gabija GajAB complex and inactivates defense. A cryo-EM structure of the virally inhibited state reveals that Gad1 forms an octameric web that encases the GajAB complex and inhibits DNA recognition and cleavage. Our results reveal the structural basis of assembly of the Gabija anti-phage defense complex and define a unique mechanism of viral immune evasion.

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Section: Extended Referencesmentioning

confidence: 99%

Structural basis of Gabija anti-phage defense and viral immune evasion

Antine

Johnson

Mooney

et al. 2023

Preprint

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“…5b). Interestingly, while most AcrIF proteins disrupt the specific binding of the Csy complex to target DNA in various ways 12,13 , AcrIF3 and AcrIF5 were found to inhibit the recruitment of Cas3 helicase-nuclease to the Csy-DNA compelx 10,26 . It is worth noting that AcrIF4 expression still caused a slight decrease in transformation efficiency (∼50%; P = 0.0059) and resulted in smaller colonies on selective plates (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…(summarized in Ref. 13). Interestingly, a most recent study reported that bacteriophages can also utilize RNA anti-CRISPRs (Racrs) to suppress CRISPR immunity 14 .…”

Section: Introductionmentioning

confidence: 99%

CRISPR-repressed toxin-antitoxin provides population-level immunity against diverse anti-CRISPR elements

Shu,

Wang,

et al. 2024

Preprint

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Prokaryotic CRISPR-Cas systems are highly vulnerable to phage-encoded anti-CRISPR (Acr) factors. How CRISPR-Cas systems protect themselves remains unclear. Here, we uncovered a broad-spectrum anti-anti-CRISPR strategy involving a phage-derived toxic protein. Transcription of this toxin is normally reppressed by the CRISPR-Cas effector, but is activated to halt cell division when the effector is inhibited by any anti-CRISPR proteins or RNAs. We showed that this abortive infection-like effect efficiently expels Acr elements from bacterial population. Furthermore, we exploited this anti-anti-CRISPR mechanism to develop a screening method for specific Acr candidates for a CRISPR-Cas system, and successfully identified two distinct Acr proteins that enhance the binding of CRISPR effector to non-target DNA. Our data highlight the broad-spectrum role of CRISPR-repressed toxins in counteracting various types of Acr factors, which illuminates that the regulatory function of CRISPR-Cas confers host cells herd immunity against Acr-encoding genetic invaders, no matter they are CRISPR-targeted or not.

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“…Phage immune-evasion proteins are categorized and exhibited as coloured dots coloured according to the key below. Notable evasion proteins are indicated with text labels 24 – 28 , 42 , 50 – 53 . b , Top, size-exclusion chromatography analysis (16/600 S200) of SUMO2-tagged Bc GajAB with or without phage Phi3T Gad1 used for cryo-EM structural studies.…”

Section: Extended Data Figures and Tablesmentioning

confidence: 99%

Structural basis of Gabija anti-phage defence and viral immune evasion

Antine,

Johnson,

Mooney

et al. 2023

Nature

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Bacteria encode hundreds of diverse defence systems that protect them from viral infection and inhibit phage propagation1–5. Gabija is one of the most prevalent anti-phage defence systems, occurring in more than 15% of all sequenced bacterial and archaeal genomes1,6,7, but the molecular basis of how Gabija defends cells from viral infection remains poorly understood. Here we use X-ray crystallography and cryo-electron microscopy (cryo-EM) to define how Gabija proteins assemble into a supramolecular complex of around 500 kDa that degrades phage DNA. Gabija protein A (GajA) is a DNA endonuclease that tetramerizes to form the core of the anti-phage defence complex. Two sets of Gabija protein B (GajB) dimers dock at opposite sides of the complex and create a 4:4 GajA–GajB assembly (hereafter, GajAB) that is essential for phage resistance in vivo. We show that a phage-encoded protein, Gabija anti-defence 1 (Gad1), directly binds to the Gabija GajAB complex and inactivates defence. A cryo-EM structure of the virally inhibited state shows that Gad1 forms an octameric web that encases the GajAB complex and inhibits DNA recognition and cleavage. Our results reveal the structural basis of assembly of the Gabija anti-phage defence complex and define a unique mechanism of viral immune evasion.

show abstract

Non-canonical inhibition strategies and structural basis of anti-CRISPR proteins targeting type I CRISPR-Cas systems

Cited by 14 publications

References 89 publications

Structural basis of Gabija anti-phage defense and viral immune evasion

Structural basis of Gabija anti-phage defense and viral immune evasion

CRISPR-repressed toxin-antitoxin provides population-level immunity against diverse anti-CRISPR elements

Structural basis of Gabija anti-phage defence and viral immune evasion

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