Guide-free Cas9 from pathogenic Campylobacter jejuni bacteria causes severe damage to DNA

Chinmoy Saha; Prarthana Mohanraju; Andrew Stubbs; Gaurav Dugar; Youri Hoogstrate; Gert Jan Kremers; Wiggert A. Van Cappellen; Deborah Horst-Kreft; Charlie Laffeber; Joyce H.G. Lebbink; Serena Bruens; Duncan Gaskin; Dior Beerens; Maarten Klunder; Rob Joosten; Jeroen A.A. Demmers; Dik Van Gent; Johan W. Mouton; Peter J. Van Der Spek; John Van Der Oost; Peter Van Baarlen; Rogier Louwen

doi:10.1126/sciadv.aaz4849

Guide-free Cas9 from pathogenic Campylobacter jejuni bacteria causes severe damage to DNA

Chinmoy Saha, Prarthana Mohanraju, Andrew Stubbs, Gaurav Dugar, Youri Hoogstrate, Gert Jan Kremers, Wiggert A. Van Cappellen, Deborah Horst-Kreft, Charlie Laffeber, Joyce H.G. Lebbink, Serena Bruens, Duncan Gaskin, Dior Beerens, Maarten Klunder, Rob Joosten, Jeroen A.A. Demmers, Dik Van Gent, Johan W. Mouton, Peter J. Van Der Spek^*, John Van Der Oost^*Peter Van Baarlen, Rogier Louwen^*

^*Corresponding author for this work

Research output: Contribution to journal › Article › Academic › peer-review

31 Citations (Scopus)

Abstract

CRISPR-Cas9 systems are enriched in human pathogenic bacteria and have been linked to cytotoxicity by an unknown mechanism. Here, we show that upon infection of human cells, Campylobacter jejuni secretes its Cas9 (CjeCas9) nuclease into their cytoplasm. Next, a native nuclear localization signal enables CjeCas9 nuclear entry, where it catalyzes metal-dependent nonspecific DNA cleavage leading to cell death. Compared to CjeCas9, native Cas9 of Streptococcus pyogenes (SpyCas9) is more suitable for guide-dependent editing. However, in human cells, native SpyCas9 may still cause some DNA damage, most likely because of its ssDNA cleavage activity. This side effect can be completely prevented by saturation of SpyCas9 with an appropriate guide RNA, which is only partially effective for CjeCas9. We conclude that CjeCas9 plays an active role in attacking human cells rather than in viral defense. Moreover, these unique catalytic features may therefore make CjeCas9 less suitable for genome editing applications.

Original language	English
Article number	eaaz4849
Journal	Science Advances
Volume	6
Issue number	25
DOIs	https://doi.org/10.1126/sciadv.aaz4849
Publication status	Published - Jun 2020

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Saha, C., Mohanraju, P., Stubbs, A., Dugar, G., Hoogstrate, Y., Kremers, G. J., Van Cappellen, W. A., Horst-Kreft, D., Laffeber, C., Lebbink, J. H. G., Bruens, S., Gaskin, D., Beerens, D., Klunder, M., Joosten, R., Demmers, J. A. A., Van Gent, D., Mouton, J. W., Van Der Spek, P. J., ... Louwen, R. (2020). Guide-free Cas9 from pathogenic Campylobacter jejuni bacteria causes severe damage to DNA. Science Advances, 6(25), Article eaaz4849. https://doi.org/10.1126/sciadv.aaz4849

@article{7a497ea4f81b4f5e996f2484de5ba023,

title = "Guide-free Cas9 from pathogenic Campylobacter jejuni bacteria causes severe damage to DNA",

abstract = "CRISPR-Cas9 systems are enriched in human pathogenic bacteria and have been linked to cytotoxicity by an unknown mechanism. Here, we show that upon infection of human cells, Campylobacter jejuni secretes its Cas9 (CjeCas9) nuclease into their cytoplasm. Next, a native nuclear localization signal enables CjeCas9 nuclear entry, where it catalyzes metal-dependent nonspecific DNA cleavage leading to cell death. Compared to CjeCas9, native Cas9 of Streptococcus pyogenes (SpyCas9) is more suitable for guide-dependent editing. However, in human cells, native SpyCas9 may still cause some DNA damage, most likely because of its ssDNA cleavage activity. This side effect can be completely prevented by saturation of SpyCas9 with an appropriate guide RNA, which is only partially effective for CjeCas9. We conclude that CjeCas9 plays an active role in attacking human cells rather than in viral defense. Moreover, these unique catalytic features may therefore make CjeCas9 less suitable for genome editing applications.",

author = "Chinmoy Saha and Prarthana Mohanraju and Andrew Stubbs and Gaurav Dugar and Youri Hoogstrate and Kremers, {Gert Jan} and {Van Cappellen}, {Wiggert A.} and Deborah Horst-Kreft and Charlie Laffeber and Lebbink, {Joyce H.G.} and Serena Bruens and Duncan Gaskin and Dior Beerens and Maarten Klunder and Rob Joosten and Demmers, {Jeroen A.A.} and {Van Gent}, Dik and Mouton, {Johan W.} and {Van Der Spek}, {Peter J.} and {Van Der Oost}, John and {Van Baarlen}, Peter and Rogier Louwen",

year = "2020",

month = jun,

doi = "10.1126/sciadv.aaz4849",

language = "English",

volume = "6",

journal = "Science Advances",

issn = "2375-2548",

publisher = "American Association for the Advancement of Science",

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Saha, C, Mohanraju, P, Stubbs, A, Dugar, G, Hoogstrate, Y, Kremers, GJ, Van Cappellen, WA, Horst-Kreft, D, Laffeber, C, Lebbink, JHG, Bruens, S, Gaskin, D, Beerens, D, Klunder, M, Joosten, R, Demmers, JAA, Van Gent, D, Mouton, JW, Van Der Spek, PJ, Van Der Oost, J , Van Baarlen, P & Louwen, R 2020, 'Guide-free Cas9 from pathogenic Campylobacter jejuni bacteria causes severe damage to DNA', Science Advances, vol. 6, no. 25, eaaz4849. https://doi.org/10.1126/sciadv.aaz4849

TY - JOUR

T1 - Guide-free Cas9 from pathogenic Campylobacter jejuni bacteria causes severe damage to DNA

AU - Saha, Chinmoy

AU - Mohanraju, Prarthana

AU - Stubbs, Andrew

AU - Dugar, Gaurav

AU - Hoogstrate, Youri

AU - Kremers, Gert Jan

AU - Van Cappellen, Wiggert A.

AU - Horst-Kreft, Deborah

AU - Laffeber, Charlie

AU - Lebbink, Joyce H.G.

AU - Bruens, Serena

AU - Gaskin, Duncan

AU - Beerens, Dior

AU - Klunder, Maarten

AU - Joosten, Rob

AU - Demmers, Jeroen A.A.

AU - Van Gent, Dik

AU - Mouton, Johan W.

AU - Van Der Spek, Peter J.

AU - Van Der Oost, John

AU - Van Baarlen, Peter

AU - Louwen, Rogier

PY - 2020/6

Y1 - 2020/6

N2 - CRISPR-Cas9 systems are enriched in human pathogenic bacteria and have been linked to cytotoxicity by an unknown mechanism. Here, we show that upon infection of human cells, Campylobacter jejuni secretes its Cas9 (CjeCas9) nuclease into their cytoplasm. Next, a native nuclear localization signal enables CjeCas9 nuclear entry, where it catalyzes metal-dependent nonspecific DNA cleavage leading to cell death. Compared to CjeCas9, native Cas9 of Streptococcus pyogenes (SpyCas9) is more suitable for guide-dependent editing. However, in human cells, native SpyCas9 may still cause some DNA damage, most likely because of its ssDNA cleavage activity. This side effect can be completely prevented by saturation of SpyCas9 with an appropriate guide RNA, which is only partially effective for CjeCas9. We conclude that CjeCas9 plays an active role in attacking human cells rather than in viral defense. Moreover, these unique catalytic features may therefore make CjeCas9 less suitable for genome editing applications.

AB - CRISPR-Cas9 systems are enriched in human pathogenic bacteria and have been linked to cytotoxicity by an unknown mechanism. Here, we show that upon infection of human cells, Campylobacter jejuni secretes its Cas9 (CjeCas9) nuclease into their cytoplasm. Next, a native nuclear localization signal enables CjeCas9 nuclear entry, where it catalyzes metal-dependent nonspecific DNA cleavage leading to cell death. Compared to CjeCas9, native Cas9 of Streptococcus pyogenes (SpyCas9) is more suitable for guide-dependent editing. However, in human cells, native SpyCas9 may still cause some DNA damage, most likely because of its ssDNA cleavage activity. This side effect can be completely prevented by saturation of SpyCas9 with an appropriate guide RNA, which is only partially effective for CjeCas9. We conclude that CjeCas9 plays an active role in attacking human cells rather than in viral defense. Moreover, these unique catalytic features may therefore make CjeCas9 less suitable for genome editing applications.

U2 - 10.1126/sciadv.aaz4849

DO - 10.1126/sciadv.aaz4849

M3 - Article

C2 - 32596446

AN - SCOPUS:85086803325

SN - 2375-2548

VL - 6

JO - Science Advances

JF - Science Advances

IS - 25

M1 - eaaz4849

ER -

Guide-free Cas9 from pathogenic Campylobacter jejuni bacteria causes severe damage to DNA

Abstract

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