Discovery and Characterization of Cas9 Inhibitors Disseminated across Seven Bacterial Phyla

Ruben Vazquez-Uribe, Eric van der Helm, Maria-Anna Misiakou, Sang-Woo Lee, Stefan Kol, Morten Otto Alexander Sommer*

*Corresponding author for this work

Research output: Contribution to journalJournal articleResearchpeer-review

Abstract

CRISPR-Cas systems in bacteria and archaea provide immunity against bacteriophages and plasmids. To overcome CRISPR immunity, phages have acquired anti-CRISPR genes that reduce CRISPR-Cas activity. Using a synthetic genetic circuit, we developed a high-throughput approach to discover anti-CRISPR genes from metagenomic libraries based on their functional activity rather than sequence homology or genetic context. We identified 11 DNA fragments from soil, animal, and human metagenomes that circumvent Streptococcus pyogenes Cas9 activity in our selection strain. Further in vivo and in vitro characterization of a subset of these hits validated the activity of four anti-CRISPRs. Notably, homologs of some of these anti-CRISPRs were detected in seven different phyla, namely Firmicutes, Proteobacteria, Bacteroidetes, Actinobacteria, Cyanobacteria, Spirochaetes, and Balneolaeota, and have high sequence identity suggesting recent horizontal gene transfer. Thus, anti-CRISPRs against type II-A CRISPR-Cas systems are widely distributed across bacterial phyla, suggesting a more complex ecological role than previously appreciated.
Original languageEnglish
JournalCell Host & Microbe
Volume25
Issue number2
Pages (from-to)233-241
ISSN1931-3128
DOIs
Publication statusPublished - 2019

Cite this

@article{a67c83a71ffc4234b11be46ce2697f87,
title = "Discovery and Characterization of Cas9 Inhibitors Disseminated across Seven Bacterial Phyla",
abstract = "CRISPR-Cas systems in bacteria and archaea provide immunity against bacteriophages and plasmids. To overcome CRISPR immunity, phages have acquired anti-CRISPR genes that reduce CRISPR-Cas activity. Using a synthetic genetic circuit, we developed a high-throughput approach to discover anti-CRISPR genes from metagenomic libraries based on their functional activity rather than sequence homology or genetic context. We identified 11 DNA fragments from soil, animal, and human metagenomes that circumvent Streptococcus pyogenes Cas9 activity in our selection strain. Further in vivo and in vitro characterization of a subset of these hits validated the activity of four anti-CRISPRs. Notably, homologs of some of these anti-CRISPRs were detected in seven different phyla, namely Firmicutes, Proteobacteria, Bacteroidetes, Actinobacteria, Cyanobacteria, Spirochaetes, and Balneolaeota, and have high sequence identity suggesting recent horizontal gene transfer. Thus, anti-CRISPRs against type II-A CRISPR-Cas systems are widely distributed across bacterial phyla, suggesting a more complex ecological role than previously appreciated.",
author = "Ruben Vazquez-Uribe and {van der Helm}, Eric and Maria-Anna Misiakou and Sang-Woo Lee and Stefan Kol and Sommer, {Morten Otto Alexander}",
year = "2019",
doi = "10.1016/j.chom.2019.01.003",
language = "English",
volume = "25",
pages = "233--241",
journal = "Cell Host & Microbe",
issn = "1931-3128",
publisher = "Cell Press",
number = "2",

}

Discovery and Characterization of Cas9 Inhibitors Disseminated across Seven Bacterial Phyla. / Vazquez-Uribe, Ruben; van der Helm, Eric; Misiakou, Maria-Anna; Lee, Sang-Woo; Kol, Stefan; Sommer, Morten Otto Alexander.

In: Cell Host & Microbe, Vol. 25, No. 2, 2019, p. 233-241.

Research output: Contribution to journalJournal articleResearchpeer-review

TY - JOUR

T1 - Discovery and Characterization of Cas9 Inhibitors Disseminated across Seven Bacterial Phyla

AU - Vazquez-Uribe, Ruben

AU - van der Helm, Eric

AU - Misiakou, Maria-Anna

AU - Lee, Sang-Woo

AU - Kol, Stefan

AU - Sommer, Morten Otto Alexander

PY - 2019

Y1 - 2019

N2 - CRISPR-Cas systems in bacteria and archaea provide immunity against bacteriophages and plasmids. To overcome CRISPR immunity, phages have acquired anti-CRISPR genes that reduce CRISPR-Cas activity. Using a synthetic genetic circuit, we developed a high-throughput approach to discover anti-CRISPR genes from metagenomic libraries based on their functional activity rather than sequence homology or genetic context. We identified 11 DNA fragments from soil, animal, and human metagenomes that circumvent Streptococcus pyogenes Cas9 activity in our selection strain. Further in vivo and in vitro characterization of a subset of these hits validated the activity of four anti-CRISPRs. Notably, homologs of some of these anti-CRISPRs were detected in seven different phyla, namely Firmicutes, Proteobacteria, Bacteroidetes, Actinobacteria, Cyanobacteria, Spirochaetes, and Balneolaeota, and have high sequence identity suggesting recent horizontal gene transfer. Thus, anti-CRISPRs against type II-A CRISPR-Cas systems are widely distributed across bacterial phyla, suggesting a more complex ecological role than previously appreciated.

AB - CRISPR-Cas systems in bacteria and archaea provide immunity against bacteriophages and plasmids. To overcome CRISPR immunity, phages have acquired anti-CRISPR genes that reduce CRISPR-Cas activity. Using a synthetic genetic circuit, we developed a high-throughput approach to discover anti-CRISPR genes from metagenomic libraries based on their functional activity rather than sequence homology or genetic context. We identified 11 DNA fragments from soil, animal, and human metagenomes that circumvent Streptococcus pyogenes Cas9 activity in our selection strain. Further in vivo and in vitro characterization of a subset of these hits validated the activity of four anti-CRISPRs. Notably, homologs of some of these anti-CRISPRs were detected in seven different phyla, namely Firmicutes, Proteobacteria, Bacteroidetes, Actinobacteria, Cyanobacteria, Spirochaetes, and Balneolaeota, and have high sequence identity suggesting recent horizontal gene transfer. Thus, anti-CRISPRs against type II-A CRISPR-Cas systems are widely distributed across bacterial phyla, suggesting a more complex ecological role than previously appreciated.

U2 - 10.1016/j.chom.2019.01.003

DO - 10.1016/j.chom.2019.01.003

M3 - Journal article

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VL - 25

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JO - Cell Host & Microbe

JF - Cell Host & Microbe

SN - 1931-3128

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