---
OA_place: publisher
OA_type: hybrid
PlanS_conform: '1'
_id: '20963'
abstract:
- lang: eng
text: In all domains of life, tRNAs mediate the transfer of genetic information
from mRNAs to proteins. As their depletion suppresses translation and, consequently,
viral replication, tRNAs represent long-standing and increasingly recognized targets
of innate immunity1,2,3,4,5. Here we report Cas12a3 effector nucleases from type V
CRISPR–Cas adaptive immune systems in bacteria that preferentially cleave tRNAs
after recognition of target RNA. Cas12a3 orthologues belong to one of two previously
unreported nuclease clades that exhibit RNA-mediated cleavage of non-target RNA,
and are distinct from all other known type V systems. Through cell-based and biochemical
assays and direct RNA sequencing, we demonstrate that recognition of a complementary
target RNA by the CRISPR RNA triggers Cas12a3 to cleave the conserved 5′-CCA-3′
tail of diverse tRNAs to drive growth arrest and anti-phage defence. Cryogenic
electron microscopy structures further revealed a distinct tRNA-loading domain
that positions the tRNA tail in the RuvC active site of the nuclease. By designing
synthetic reporters that mimic the tRNA acceptor stem and tail, we expanded the
capacity of current CRISPR-based diagnostics for multiplexed RNA detection. Overall,
these findings reveal widespread tRNA inactivation as a previously unrecognized
CRISPR-based immune strategy that broadens the application space of the existing
CRISPR toolbox.
acknowledgement: 'We thank Ł. Koziej for processing of the initial cryo-EM datasets,
S. Schmelz for support in cryo-EM, A. Gatzemeier for assistance in the purification
of dBa1Cas12a3, R. Rarose for support with the in vitro RNA experiments, M. Kaminski
for providing purified PsmCas13b protein, L. Schönemann for protein purification,
and C. Krempl and S. Backesfor providing the RSV and influenza A transcript-encoding
plasmids. This work was supported through funding by the European Research Council
(101001394 to S.G.; 865973 and 101158249 to C.L.B.), the R. Gaurth Hansen Family
(to R.N.J.), the National Institutes of Health (R35GM138080 to R.N.J.), the PostDoc
Plus Program from the Graduate School of Life Sciences at Julius-Maximilians-Universität
Würzburg (to O.D.), and the Deutsche Forschungsgemeinschaft (DFG, German Research
Foundation) under Germany’s Excellence Strategy–The Berlin Mathematics Research
Center MATH+ (EXC−2046/1, project ID: 390685689 to M.v.K.). Open access funding
provided by Helmholtz-Zentrum für Infektionsforschung GmbH (HZI).'
article_processing_charge: Yes (via OA deal)
article_type: original
author:
- first_name: Oleg
full_name: Dmytrenko, Oleg
last_name: Dmytrenko
- first_name: Biao
full_name: Yuan, Biao
last_name: Yuan
- first_name: Kadin T.
full_name: Crosby, Kadin T.
last_name: Crosby
- first_name: Max
full_name: Krebel, Max
last_name: Krebel
- first_name: Xiye
full_name: Chen, Xiye
last_name: Chen
- first_name: Jakub S.
full_name: Nowak, Jakub S.
last_name: Nowak
- first_name: Andrzej
full_name: Chramiec-Głąbik, Andrzej
last_name: Chramiec-Głąbik
- first_name: Bamidele
full_name: Filani, Bamidele
last_name: Filani
- first_name: Anne-Sophie
full_name: Gribling-Burrer, Anne-Sophie
last_name: Gribling-Burrer
- first_name: Wiep
full_name: van der Toorn, Wiep
last_name: van der Toorn
- first_name: Max
full_name: von Kleist, Max
last_name: von Kleist
- first_name: Tatjana
full_name: Achmedov, Tatjana
last_name: Achmedov
- first_name: Redmond P.
full_name: Smyth, Redmond P.
last_name: Smyth
- first_name: Sebastian
full_name: Glatt, Sebastian
last_name: Glatt
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
- first_name: Dirk W.
full_name: Heinz, Dirk W.
last_name: Heinz
- first_name: Ryan N.
full_name: Jackson, Ryan N.
last_name: Jackson
- first_name: Chase L.
full_name: Beisel, Chase L.
last_name: Beisel
citation:
ama: Dmytrenko O, Yuan B, Crosby KT, et al. RNA-triggered Cas12a3 cleaves tRNA tails
to execute bacterial immunity. Nature. 2026. doi:10.1038/s41586-025-09852-9
apa: Dmytrenko, O., Yuan, B., Crosby, K. T., Krebel, M., Chen, X., Nowak, J. S.,
… Beisel, C. L. (2026). RNA-triggered Cas12a3 cleaves tRNA tails to execute bacterial
immunity. Nature. Springer Nature. https://doi.org/10.1038/s41586-025-09852-9
chicago: Dmytrenko, Oleg, Biao Yuan, Kadin T. Crosby, Max Krebel, Xiye Chen, Jakub
S. Nowak, Andrzej Chramiec-Głąbik, et al. “RNA-Triggered Cas12a3 Cleaves TRNA
Tails to Execute Bacterial Immunity.” Nature. Springer Nature, 2026. https://doi.org/10.1038/s41586-025-09852-9.
ieee: O. Dmytrenko et al., “RNA-triggered Cas12a3 cleaves tRNA tails to execute
bacterial immunity,” Nature. Springer Nature, 2026.
ista: Dmytrenko O, Yuan B, Crosby KT, Krebel M, Chen X, Nowak JS, Chramiec-Głąbik
A, Filani B, Gribling-Burrer A-S, van der Toorn W, von Kleist M, Achmedov T, Smyth
RP, Glatt S, Bravo JPK, Heinz DW, Jackson RN, Beisel CL. 2026. RNA-triggered Cas12a3
cleaves tRNA tails to execute bacterial immunity. Nature.
mla: Dmytrenko, Oleg, et al. “RNA-Triggered Cas12a3 Cleaves TRNA Tails to Execute
Bacterial Immunity.” Nature, Springer Nature, 2026, doi:10.1038/s41586-025-09852-9.
short: O. Dmytrenko, B. Yuan, K.T. Crosby, M. Krebel, X. Chen, J.S. Nowak, A. Chramiec-Głąbik,
B. Filani, A.-S. Gribling-Burrer, W. van der Toorn, M. von Kleist, T. Achmedov,
R.P. Smyth, S. Glatt, J.P.K. Bravo, D.W. Heinz, R.N. Jackson, C.L. Beisel, Nature
(2026).
date_created: 2026-01-08T07:57:17Z
date_published: 2026-01-07T00:00:00Z
date_updated: 2026-01-12T10:13:56Z
day: '07'
ddc:
- '570'
department:
- _id: JaBr
doi: 10.1038/s41586-025-09852-9
external_id:
pmid:
- '41501459'
has_accepted_license: '1'
language:
- iso: eng
license: https://creativecommons.org/licenses/by/4.0/
main_file_link:
- open_access: '1'
url: https://doi.org/10.1038/s41586-025-09852-9
month: '01'
oa: 1
oa_version: Published Version
pmid: 1
publication: Nature
publication_identifier:
eissn:
- 1476-4687
issn:
- 0028-0836
publication_status: epub_ahead
publisher: Springer Nature
quality_controlled: '1'
scopus_import: '1'
status: public
title: RNA-triggered Cas12a3 cleaves tRNA tails to execute bacterial immunity
tmp:
image: /images/cc_by.png
legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode
name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)
short: CC BY (4.0)
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
year: '2026'
...
---
OA_place: publisher
OA_type: hybrid
PlanS_conform: '1'
_id: '20143'
abstract:
- lang: eng
text: Bacteria and archaea deploy diverse antiviral defense systems, many of which
remain mechanistically uncharacterized. Here, we characterize Kiwa, a widespread
two-component system composed of the transmembrane sensor KwaA and the DNA-binding
effector KwaB. Cryogenic electron microscopy (cryo-EM) analysis reveals that KwaA
and KwaB assemble into a large, membrane-associated supercomplex. Upon phage binding,
KwaA senses infection at the membrane, leading to KwaB binding of ejected phage
DNA and inhibition of replication and late transcription, without inducing host
cell death. Although KwaB can bind DNA independently, its antiviral activity requires
association with KwaA, suggesting spatial or conformational regulation. We show
that the phage-encoded DNA-mimic protein Gam directly binds and inhibits KwaB
but that co-expression with the Gam-targeted RecBCD system restores protection
by Kiwa. Our findings support a model in which Kiwa coordinates membrane-associated
detection of phage infection with downstream DNA binding by its effector, forming
a spatially coordinated antiviral mechanism.
acknowledgement: We thank Rotem Sorek (Weizmann Institute of Science) for the Lambda
Gam mutant and Ian Molineux (University of Texas) for T4Δgp2. We thank You Yu (Zhejiang
University-University of Edinburgh Institute) and J. De La Cruz (MSK) for assistance
with cryo-EM data collection and Lyuqin Zheng (MSK) for discussions on structural
analysis. We thank the Imaging and Microscopy Centre (IMC) at the University of
Southampton. This work was supported by Royal Society grant RGS\R2\222312 to F.L.N.;
Welch Foundation grant F-1938 and National Institutes of Health R35GM138348 to D.W.T.;
Wessex Medical Research Innovation grant AE06 to T.A.; and NIH grant GM145888 and
Maloris Foundation and Memorial Sloan-Kettering Core grant (P30-CA008748) to D.J.P.
In addition to MSKCC cryo-EM resources, some of this work was performed at the National
Center for CryoEM Access and Training (NCCAT) and the Simons Electron Microscopy
Center located at the New York Structural Biology Center, supported by the NIH Common
Fund Transformative High Resolution Cryo-Electron Microscopy program (U24 GM129539)
and Simons Foundation (SF349247) and NY State Assembly grants. This research used
NSLS-II MX X-ray User Resources (FMX) of the National Synchrotron Light Source II,
operated for the DOE Office of Science by Brookhaven National Laboratory under contract
no. DE-SC0012704. The Center for BioMolecular Structure (CBMS) is primarily supported
by the NIH, the National Institute of General Medical Sciences (NIGMS) through a
Center Core P30 Grant (P30GM133893), and by the DOE Office of Biological and Environmental
Research (KP1605010). R.K. and E.V.K. are supported by the Intramural Research Program
of the NIH (National Library of Medicine).
article_processing_charge: Yes (in subscription journal)
article_type: original
author:
- first_name: Zhiying
full_name: Zhang, Zhiying
last_name: Zhang
- first_name: Thomas C.
full_name: Todeschini, Thomas C.
last_name: Todeschini
- first_name: Yi
full_name: Wu, Yi
last_name: Wu
- first_name: Roman
full_name: Kogay, Roman
last_name: Kogay
- first_name: Ameena
full_name: Naji, Ameena
last_name: Naji
- first_name: Joaquin
full_name: Cardenas Rodriguez, Joaquin
last_name: Cardenas Rodriguez
- first_name: Rupavidhya
full_name: Mondi, Rupavidhya
last_name: Mondi
- first_name: Daniel
full_name: Kaganovich, Daniel
last_name: Kaganovich
- first_name: David W.
full_name: Taylor, David W.
last_name: Taylor
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
- first_name: Marianna
full_name: Teplova, Marianna
last_name: Teplova
- first_name: Triana
full_name: Amen, Triana
last_name: Amen
- first_name: Eugene
full_name: Koonin, Eugene
last_name: Koonin
- first_name: Dinshaw J.
full_name: Patel, Dinshaw J.
last_name: Patel
- first_name: Franklin L.
full_name: Nobrega, Franklin L.
last_name: Nobrega
citation:
ama: Zhang Z, Todeschini TC, Wu Y, et al. Kiwa is a membrane-embedded defense supercomplex
activated at phage attachment sites. Cell. 2025;188(21):5862-5877.e23.
doi:10.1016/j.cell.2025.07.002
apa: Zhang, Z., Todeschini, T. C., Wu, Y., Kogay, R., Naji, A., Cardenas Rodriguez,
J., … Nobrega, F. L. (2025). Kiwa is a membrane-embedded defense supercomplex
activated at phage attachment sites. Cell. Elsevier. https://doi.org/10.1016/j.cell.2025.07.002
chicago: Zhang, Zhiying, Thomas C. Todeschini, Yi Wu, Roman Kogay, Ameena Naji,
Joaquin Cardenas Rodriguez, Rupavidhya Mondi, et al. “Kiwa Is a Membrane-Embedded
Defense Supercomplex Activated at Phage Attachment Sites.” Cell. Elsevier,
2025. https://doi.org/10.1016/j.cell.2025.07.002.
ieee: Z. Zhang et al., “Kiwa is a membrane-embedded defense supercomplex
activated at phage attachment sites,” Cell, vol. 188, no. 21. Elsevier,
p. 5862–5877.e23, 2025.
ista: Zhang Z, Todeschini TC, Wu Y, Kogay R, Naji A, Cardenas Rodriguez J, Mondi
R, Kaganovich D, Taylor DW, Bravo JPK, Teplova M, Amen T, Koonin E, Patel DJ,
Nobrega FL. 2025. Kiwa is a membrane-embedded defense supercomplex activated at
phage attachment sites. Cell. 188(21), 5862–5877.e23.
mla: Zhang, Zhiying, et al. “Kiwa Is a Membrane-Embedded Defense Supercomplex Activated
at Phage Attachment Sites.” Cell, vol. 188, no. 21, Elsevier, 2025, p.
5862–5877.e23, doi:10.1016/j.cell.2025.07.002.
short: Z. Zhang, T.C. Todeschini, Y. Wu, R. Kogay, A. Naji, J. Cardenas Rodriguez,
R. Mondi, D. Kaganovich, D.W. Taylor, J.P.K. Bravo, M. Teplova, T. Amen, E. Koonin,
D.J. Patel, F.L. Nobrega, Cell 188 (2025) 5862–5877.e23.
date_created: 2025-08-07T05:00:04Z
date_published: 2025-10-16T00:00:00Z
date_updated: 2025-12-29T14:15:58Z
day: '16'
ddc:
- '570'
department:
- _id: JaBr
doi: 10.1016/j.cell.2025.07.002
external_id:
isi:
- '001603560700005'
pmid:
- '40730155'
file:
- access_level: open_access
checksum: b944de5fbd7455f58e1ff338ad352239
content_type: application/pdf
creator: dernst
date_created: 2025-12-29T14:15:25Z
date_updated: 2025-12-29T14:15:25Z
file_id: '20875'
file_name: 2025_Cell_Zhang.pdf
file_size: 32104588
relation: main_file
success: 1
file_date_updated: 2025-12-29T14:15:25Z
has_accepted_license: '1'
intvolume: ' 188'
isi: 1
issue: '21'
language:
- iso: eng
month: '10'
oa: 1
oa_version: Published Version
page: 5862-5877.e23
pmid: 1
publication: Cell
publication_identifier:
eissn:
- 1097-4172
issn:
- 0092-8674
publication_status: published
publisher: Elsevier
quality_controlled: '1'
scopus_import: '1'
status: public
title: Kiwa is a membrane-embedded defense supercomplex activated at phage attachment
sites
tmp:
image: /images/cc_by.png
legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode
name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)
short: CC BY (4.0)
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 188
year: '2025'
...
---
DOAJ_listed: '1'
OA_place: publisher
OA_type: gold
_id: '18848'
abstract:
- lang: eng
text: Type II CRISPR endonucleases are widely used programmable genome editing tools.
Recently, CRISPR-Cas systems with highly compact nucleases have been discovered,
including Cas9d (a type II-D nuclease). Here, we report the cryo-EM structures
of a Cas9d nuclease (747 amino acids in length) in multiple functional states,
revealing a stepwise process of DNA targeting involving a conformational switch
in a REC2 domain insertion. Our structures provide insights into the intricately
folded guide RNA which acts as a structural scaffold to anchor small, flexible
protein domains for DNA recognition. The sgRNA can be truncated by up to ~25%
yet still retain activity in vivo. Using ancestral sequence reconstruction, we
generated compact nucleases capable of efficient genome editing in mammalian cells.
Collectively, our results provide mechanistic insights into the evolution and
DNA targeting of diverse type II CRISPR-Cas systems, providing a blueprint for
future re-engineering of minimal RNA-guided DNA endonucleases.
acknowledgement: We would like to thank M. Ocampo Camacho and M.F. Canedo Ocampo for
assistance with the figures. We thank M. Hooper for assistance developing the GFP
assay and operating the CE machine for in vitro cleavage analysis. We thank E. Schwartz
and A. Brilot for expert cryo-EM support in the Sauer Structural Biology Laboratory
at UT Austin. This work was funded, in part, by a sponsored research agreement with
Metagenomi, Inc. (to D.W.T), a Welch Foundation Research Grant F-1938 (to D.W.T),
and the Robert J. Kleberg, Jr. and Helen C. Kleberg Foundation Medical Research
Grant (to D.W.T), and a grant from the National Institute of Allergy and Infectious
Diseases (NIAID 1R01AI110577 to K.A.J.).
article_number: '457'
article_processing_charge: Yes
article_type: original
author:
- first_name: Rodrigo Fregoso
full_name: Ocampo, Rodrigo Fregoso
last_name: Ocampo
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
- first_name: Tyler L.
full_name: Dangerfield, Tyler L.
last_name: Dangerfield
- first_name: Isabel
full_name: Nocedal, Isabel
last_name: Nocedal
- first_name: Samatar A.
full_name: Jirde, Samatar A.
last_name: Jirde
- first_name: Lisa M.
full_name: Alexander, Lisa M.
last_name: Alexander
- first_name: Nicole C.
full_name: Thomas, Nicole C.
last_name: Thomas
- first_name: Anjali
full_name: Das, Anjali
last_name: Das
- first_name: Sarah
full_name: Nielson, Sarah
last_name: Nielson
- first_name: Kenneth A.
full_name: Johnson, Kenneth A.
last_name: Johnson
- first_name: Christopher T.
full_name: Brown, Christopher T.
last_name: Brown
- first_name: Cristina N.
full_name: Butterfield, Cristina N.
last_name: Butterfield
- first_name: Daniela S.A.
full_name: Goltsman, Daniela S.A.
last_name: Goltsman
- first_name: David W.
full_name: Taylor, David W.
last_name: Taylor
citation:
ama: Ocampo RF, Bravo JPK, Dangerfield TL, et al. DNA targeting by compact Cas9d
and its resurrected ancestor. Nature Communications. 2025;16. doi:10.1038/s41467-024-55573-4
apa: Ocampo, R. F., Bravo, J. P. K., Dangerfield, T. L., Nocedal, I., Jirde, S.
A., Alexander, L. M., … Taylor, D. W. (2025). DNA targeting by compact Cas9d and
its resurrected ancestor. Nature Communications. Springer Nature. https://doi.org/10.1038/s41467-024-55573-4
chicago: Ocampo, Rodrigo Fregoso, Jack Peter Kelly Bravo, Tyler L. Dangerfield,
Isabel Nocedal, Samatar A. Jirde, Lisa M. Alexander, Nicole C. Thomas, et al.
“DNA Targeting by Compact Cas9d and Its Resurrected Ancestor.” Nature Communications.
Springer Nature, 2025. https://doi.org/10.1038/s41467-024-55573-4.
ieee: R. F. Ocampo et al., “DNA targeting by compact Cas9d and its resurrected
ancestor,” Nature Communications, vol. 16. Springer Nature, 2025.
ista: Ocampo RF, Bravo JPK, Dangerfield TL, Nocedal I, Jirde SA, Alexander LM, Thomas
NC, Das A, Nielson S, Johnson KA, Brown CT, Butterfield CN, Goltsman DSA, Taylor
DW. 2025. DNA targeting by compact Cas9d and its resurrected ancestor. Nature
Communications. 16, 457.
mla: Ocampo, Rodrigo Fregoso, et al. “DNA Targeting by Compact Cas9d and Its Resurrected
Ancestor.” Nature Communications, vol. 16, 457, Springer Nature, 2025,
doi:10.1038/s41467-024-55573-4.
short: R.F. Ocampo, J.P.K. Bravo, T.L. Dangerfield, I. Nocedal, S.A. Jirde, L.M.
Alexander, N.C. Thomas, A. Das, S. Nielson, K.A. Johnson, C.T. Brown, C.N. Butterfield,
D.S.A. Goltsman, D.W. Taylor, Nature Communications 16 (2025).
date_created: 2025-01-19T23:01:50Z
date_published: 2025-01-07T00:00:00Z
date_updated: 2025-07-03T11:58:22Z
day: '07'
ddc:
- '570'
department:
- _id: JaBr
doi: 10.1038/s41467-024-55573-4
external_id:
pmid:
- '39774105'
file:
- access_level: open_access
checksum: 885e96690620790d5c9f188a1587b4cd
content_type: application/pdf
creator: dernst
date_created: 2025-01-22T14:35:22Z
date_updated: 2025-01-22T14:35:22Z
file_id: '18869'
file_name: 2025_NatureComm_Ocampo.pdf
file_size: 5450660
relation: main_file
success: 1
file_date_updated: 2025-01-22T14:35:22Z
has_accepted_license: '1'
intvolume: ' 16'
language:
- iso: eng
month: '01'
oa: 1
oa_version: Published Version
pmid: 1
publication: Nature Communications
publication_identifier:
eissn:
- 2041-1723
publication_status: published
publisher: Springer Nature
quality_controlled: '1'
scopus_import: '1'
status: public
title: DNA targeting by compact Cas9d and its resurrected ancestor
tmp:
image: /images/cc_by.png
legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode
name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)
short: CC BY (4.0)
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 16
year: '2025'
...
---
DOAJ_listed: '1'
OA_place: publisher
OA_type: gold
_id: '18545'
abstract:
- lang: eng
text: Clinical implementation of therapeutic genome editing relies on efficient
in vivo delivery and the safety of CRISPR-Cas tools. Previously, we identified
PsCas9 as a Type II-B family enzyme capable of editing mouse liver genome upon
adenoviral delivery without detectable off-targets and reduced chromosomal translocations.
Yet, its efficacy remains insufficient with non-viral delivery, a common challenge
for many Cas9 orthologues. Here, we sought to redesign PsCas9 for in vivo editing
using lipid nanoparticles. We solve the PsCas9 ribonucleoprotein structure with
cryo-EM and characterize it biochemically, providing a basis for its rational
engineering. Screening over numerous guide RNA and protein variants lead us to
develop engineered PsCas9 (ePsCas9) with up to 20-fold increased activity across
various targets and preserved safety advantages. We apply the same design principles
to boost the activity of FnCas9, an enzyme phylogenetically relevant to PsCas9.
Remarkably, a single administration of mRNA encoding ePsCas9 and its guide formulated
with lipid nanoparticles results in high levels of editing in the Pcsk9 gene in
mouse liver, a clinically relevant target for hypercholesterolemia treatment.
Collectively, our findings introduce ePsCas9 as a highly efficient, and precise
tool for therapeutic genome editing, in addition to the engineering strategy applicable
to other Cas9 orthologues.
article_number: '9173'
article_processing_charge: Yes
article_type: original
author:
- first_name: Dmitrii
full_name: Degtev, Dmitrii
last_name: Degtev
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
- first_name: Aikaterini
full_name: Emmanouilidi, Aikaterini
last_name: Emmanouilidi
- first_name: Aleksandar
full_name: Zdravković, Aleksandar
last_name: Zdravković
- first_name: Oi Kuan
full_name: Choong, Oi Kuan
last_name: Choong
- first_name: Julia
full_name: Liz Touza, Julia
last_name: Liz Touza
- first_name: Niklas
full_name: Selfjord, Niklas
last_name: Selfjord
- first_name: Isabel
full_name: Weisheit, Isabel
last_name: Weisheit
- first_name: Margherita
full_name: Francescatto, Margherita
last_name: Francescatto
- first_name: Pinar
full_name: Akcakaya, Pinar
last_name: Akcakaya
- first_name: Michelle
full_name: Porritt, Michelle
last_name: Porritt
- first_name: Marcello
full_name: Maresca, Marcello
last_name: Maresca
- first_name: David
full_name: Taylor, David
last_name: Taylor
- first_name: Grzegorz
full_name: Sienski, Grzegorz
last_name: Sienski
citation:
ama: Degtev D, Bravo JPK, Emmanouilidi A, et al. Engineered PsCas9 enables therapeutic
genome editing in mouse liver with lipid nanoparticles. Nature Communications.
2024;15. doi:10.1038/s41467-024-53418-8
apa: Degtev, D., Bravo, J. P. K., Emmanouilidi, A., Zdravković, A., Choong, O. K.,
Liz Touza, J., … Sienski, G. (2024). Engineered PsCas9 enables therapeutic genome
editing in mouse liver with lipid nanoparticles. Nature Communications.
Springer Nature. https://doi.org/10.1038/s41467-024-53418-8
chicago: Degtev, Dmitrii, Jack Peter Kelly Bravo, Aikaterini Emmanouilidi, Aleksandar
Zdravković, Oi Kuan Choong, Julia Liz Touza, Niklas Selfjord, et al. “Engineered
PsCas9 Enables Therapeutic Genome Editing in Mouse Liver with Lipid Nanoparticles.”
Nature Communications. Springer Nature, 2024. https://doi.org/10.1038/s41467-024-53418-8.
ieee: D. Degtev et al., “Engineered PsCas9 enables therapeutic genome editing
in mouse liver with lipid nanoparticles,” Nature Communications, vol. 15.
Springer Nature, 2024.
ista: Degtev D, Bravo JPK, Emmanouilidi A, Zdravković A, Choong OK, Liz Touza J,
Selfjord N, Weisheit I, Francescatto M, Akcakaya P, Porritt M, Maresca M, Taylor
D, Sienski G. 2024. Engineered PsCas9 enables therapeutic genome editing in mouse
liver with lipid nanoparticles. Nature Communications. 15, 9173.
mla: Degtev, Dmitrii, et al. “Engineered PsCas9 Enables Therapeutic Genome Editing
in Mouse Liver with Lipid Nanoparticles.” Nature Communications, vol. 15,
9173, Springer Nature, 2024, doi:10.1038/s41467-024-53418-8.
short: D. Degtev, J.P.K. Bravo, A. Emmanouilidi, A. Zdravković, O.K. Choong, J.
Liz Touza, N. Selfjord, I. Weisheit, M. Francescatto, P. Akcakaya, M. Porritt,
M. Maresca, D. Taylor, G. Sienski, Nature Communications 15 (2024).
date_created: 2024-11-12T10:18:04Z
date_published: 2024-11-07T00:00:00Z
date_updated: 2024-11-13T08:19:50Z
day: '07'
ddc:
- '572'
doi: 10.1038/s41467-024-53418-8
extern: '1'
file:
- access_level: open_access
checksum: dcfadc806f4144d065eb8e2032554782
content_type: application/pdf
creator: jbravo
date_created: 2024-11-12T10:18:32Z
date_updated: 2024-11-12T10:18:32Z
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file_date_updated: 2024-11-12T10:18:32Z
has_accepted_license: '1'
intvolume: ' 15'
language:
- iso: eng
license: https://creativecommons.org/licenses/by-nc-nd/4.0/
month: '11'
oa: 1
oa_version: Published Version
publication: Nature Communications
publication_identifier:
issn:
- 2041-1723
publication_status: published
publisher: Springer Nature
quality_controlled: '1'
scopus_import: '1'
status: public
title: Engineered PsCas9 enables therapeutic genome editing in mouse liver with lipid
nanoparticles
tmp:
image: /images/cc_by_nc_nd.png
legal_code_url: https://creativecommons.org/licenses/by-nc-nd/4.0/legalcode
name: Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International
(CC BY-NC-ND 4.0)
short: CC BY-NC-ND (4.0)
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 15
year: '2024'
...
---
DOAJ_listed: '1'
_id: '15372'
abstract:
- lang: eng
text: CRISPR-Cas9 is a powerful tool for genome editing, but the strict requirement
for an NGG protospacer-adjacent motif (PAM) sequence immediately next to the DNA
target limits the number of editable genes. Recently developed Cas9 variants have
been engineered with relaxed PAM requirements, including SpG-Cas9 (SpG) and the
nearly PAM-less SpRY-Cas9 (SpRY). However, the molecular mechanisms of how SpRY
recognizes all potential PAM sequences remains unclear. Here, we combine structural
and biochemical approaches to determine how SpRY interrogates DNA and recognizes
target sites. Divergent PAM sequences can be accommodated through conformational
flexibility within the PAM-interacting region, which facilitates tight binding
to off-target DNA sequences. Nuclease activation occurs ~1000-fold slower than
for Streptococcus pyogenes Cas9, enabling us to directly visualize multiple on-pathway
intermediate states. Experiments with SpG position it as an intermediate enzyme
between Cas9 and SpRY. Our findings shed light on the molecular mechanisms of
PAMless genome editing.
acknowledgement: We thank I. Stohkendl in the Taylor group for insightful discussions.
This work was supported in part by Welch Foundation grants F-1808 (to I.J.F.), and
F-1938 (to D.W.T.), the National Institutes of Health R01GM124141 (to I.J.F.), R01AI110577
(to K.A.J.), and R35GM138348 (to D.W.T.), and a Robert J. Kleberg, Jr. and Helen
C. Kleberg Foundation Medical Research Grant (to D.W.T.). The content is solely
the responsibility of the authors and does not necessarily represent the official
views of the National Institutes of Health.
article_number: '3663'
article_processing_charge: Yes
article_type: original
author:
- first_name: Grace N.
full_name: Hibshman, Grace N.
last_name: Hibshman
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
- first_name: Matthew M.
full_name: Hooper, Matthew M.
last_name: Hooper
- first_name: Tyler L.
full_name: Dangerfield, Tyler L.
last_name: Dangerfield
- first_name: Hongshan
full_name: Zhang, Hongshan
last_name: Zhang
- first_name: Ilya J.
full_name: Finkelstein, Ilya J.
last_name: Finkelstein
- first_name: Kenneth A.
full_name: Johnson, Kenneth A.
last_name: Johnson
- first_name: David W.
full_name: Taylor, David W.
last_name: Taylor
citation:
ama: Hibshman GN, Bravo JPK, Hooper MM, et al. Unraveling the mechanisms of PAMless
DNA interrogation by SpRY-Cas9. Nature Communications. 2024;15. doi:10.1038/s41467-024-47830-3
apa: Hibshman, G. N., Bravo, J. P. K., Hooper, M. M., Dangerfield, T. L., Zhang,
H., Finkelstein, I. J., … Taylor, D. W. (2024). Unraveling the mechanisms of PAMless
DNA interrogation by SpRY-Cas9. Nature Communications. Springer Nature.
https://doi.org/10.1038/s41467-024-47830-3
chicago: Hibshman, Grace N., Jack Peter Kelly Bravo, Matthew M. Hooper, Tyler L.
Dangerfield, Hongshan Zhang, Ilya J. Finkelstein, Kenneth A. Johnson, and David
W. Taylor. “Unraveling the Mechanisms of PAMless DNA Interrogation by SpRY-Cas9.”
Nature Communications. Springer Nature, 2024. https://doi.org/10.1038/s41467-024-47830-3.
ieee: G. N. Hibshman et al., “Unraveling the mechanisms of PAMless DNA interrogation
by SpRY-Cas9,” Nature Communications, vol. 15. Springer Nature, 2024.
ista: Hibshman GN, Bravo JPK, Hooper MM, Dangerfield TL, Zhang H, Finkelstein IJ,
Johnson KA, Taylor DW. 2024. Unraveling the mechanisms of PAMless DNA interrogation
by SpRY-Cas9. Nature Communications. 15, 3663.
mla: Hibshman, Grace N., et al. “Unraveling the Mechanisms of PAMless DNA Interrogation
by SpRY-Cas9.” Nature Communications, vol. 15, 3663, Springer Nature, 2024,
doi:10.1038/s41467-024-47830-3.
short: G.N. Hibshman, J.P.K. Bravo, M.M. Hooper, T.L. Dangerfield, H. Zhang, I.J.
Finkelstein, K.A. Johnson, D.W. Taylor, Nature Communications 15 (2024).
corr_author: '1'
date_created: 2024-05-12T22:01:00Z
date_published: 2024-04-30T00:00:00Z
date_updated: 2025-05-14T09:33:21Z
day: '30'
ddc:
- '570'
department:
- _id: JaBr
doi: 10.1038/s41467-024-47830-3
external_id:
pmid:
- '38688943'
file:
- access_level: open_access
checksum: 509c65919067a03ef8ad65c7192cd860
content_type: application/pdf
creator: dernst
date_created: 2024-05-13T11:46:19Z
date_updated: 2024-05-13T11:46:19Z
file_id: '15386'
file_name: 2024_NatureComm_Hibshman.pdf
file_size: 7477013
relation: main_file
success: 1
file_date_updated: 2024-05-13T11:46:19Z
has_accepted_license: '1'
intvolume: ' 15'
language:
- iso: eng
month: '04'
oa: 1
oa_version: Published Version
pmid: 1
publication: Nature Communications
publication_identifier:
eissn:
- 2041-1723
publication_status: published
publisher: Springer Nature
quality_controlled: '1'
scopus_import: '1'
status: public
title: Unraveling the mechanisms of PAMless DNA interrogation by SpRY-Cas9
tmp:
image: /images/cc_by.png
legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode
name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)
short: CC BY (4.0)
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 15
year: '2024'
...
---
_id: '17111'
abstract:
- lang: eng
text: Membrane-associated protein phase separation plays critical roles in cell
biology, driving essential cellular phenomena from immune signaling to membrane
traffic. Importantly, by reducing dimensionality from three to two dimensions,
lipid bilayers can nucleate phase separation at far lower concentrations compared
with those required for phase separation in solution. How might other intracellular
lipid substrates, such as lipid droplets, contribute to nucleation of phase separation?
Distinct from bilayer membranes, lipid droplets consist of a phospholipid monolayer
surrounding a core of neutral lipids, and they are energy storage organelles that
protect cells from lipotoxicity and oxidative stress. Here, we show that intrinsically
disordered proteins can undergo phase separation on the surface of synthetic and
cell-derived lipid droplets. Specifically, we find that the model disordered domains
FUS LC and LAF-1 RGG separate into protein-rich and protein-depleted phases on
the surfaces of lipid droplets. Owing to the hydrophobic nature of interactions
between FUS LC proteins, increasing ionic strength drives an increase in its phase
separation on droplet surfaces. The opposite is true for LAF-1 RGG, owing to the
electrostatic nature of its interprotein interactions. In both cases, protein-rich
phases on the surfaces of synthetic and cell-derived lipid droplets demonstrate
molecular mobility indicative of a liquid-like state. Our results show that lipid
droplets can nucleate protein condensates, suggesting that protein phase separation
could be key in organizing biological processes involving lipid droplets.
article_processing_charge: No
article_type: original
author:
- first_name: Advika
full_name: Kamatar, Advika
last_name: Kamatar
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
- first_name: Feng
full_name: Yuan, Feng
last_name: Yuan
- first_name: Liping
full_name: Wang, Liping
last_name: Wang
- first_name: Eileen M.
full_name: Lafer, Eileen M.
last_name: Lafer
- first_name: David W.
full_name: Taylor, David W.
last_name: Taylor
- first_name: Jeanne C.
full_name: Stachowiak, Jeanne C.
last_name: Stachowiak
- first_name: Sapun H.
full_name: Parekh, Sapun H.
last_name: Parekh
citation:
ama: Kamatar A, Bravo JPK, Yuan F, et al. Lipid droplets as substrates for protein
phase separation. Biophysical Journal. 2024;123(11):1494-1507. doi:10.1016/j.bpj.2024.03.015
apa: Kamatar, A., Bravo, J. P. K., Yuan, F., Wang, L., Lafer, E. M., Taylor, D.
W., … Parekh, S. H. (2024). Lipid droplets as substrates for protein phase separation.
Biophysical Journal. Elsevier. https://doi.org/10.1016/j.bpj.2024.03.015
chicago: Kamatar, Advika, Jack Peter Kelly Bravo, Feng Yuan, Liping Wang, Eileen
M. Lafer, David W. Taylor, Jeanne C. Stachowiak, and Sapun H. Parekh. “Lipid Droplets
as Substrates for Protein Phase Separation.” Biophysical Journal. Elsevier,
2024. https://doi.org/10.1016/j.bpj.2024.03.015.
ieee: A. Kamatar et al., “Lipid droplets as substrates for protein phase
separation,” Biophysical Journal, vol. 123, no. 11. Elsevier, pp. 1494–1507,
2024.
ista: Kamatar A, Bravo JPK, Yuan F, Wang L, Lafer EM, Taylor DW, Stachowiak JC,
Parekh SH. 2024. Lipid droplets as substrates for protein phase separation. Biophysical
Journal. 123(11), 1494–1507.
mla: Kamatar, Advika, et al. “Lipid Droplets as Substrates for Protein Phase Separation.”
Biophysical Journal, vol. 123, no. 11, Elsevier, 2024, pp. 1494–507, doi:10.1016/j.bpj.2024.03.015.
short: A. Kamatar, J.P.K. Bravo, F. Yuan, L. Wang, E.M. Lafer, D.W. Taylor, J.C.
Stachowiak, S.H. Parekh, Biophysical Journal 123 (2024) 1494–1507.
date_created: 2024-06-04T06:41:03Z
date_published: 2024-06-04T00:00:00Z
date_updated: 2025-01-13T11:03:41Z
day: '04'
doi: 10.1016/j.bpj.2024.03.015
extern: '1'
external_id:
pmid:
- '38462838'
intvolume: ' 123'
issue: '11'
language:
- iso: eng
month: '06'
oa_version: None
page: 1494-1507
pmid: 1
publication: Biophysical Journal
publication_identifier:
issn:
- 0006-3495
publication_status: published
publisher: Elsevier
quality_controlled: '1'
scopus_import: '1'
status: public
title: Lipid droplets as substrates for protein phase separation
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 123
year: '2024'
...
---
OA_place: repository
OA_type: green
_id: '17112'
abstract:
- lang: eng
text: The generation of cyclic oligoadenylates and subsequent allosteric activation
of proteins that carry sensory domains is a distinctive feature of type III CRISPR-Cas
systems. In this work, we characterize a set of associated genes of a type III-B
system from Haliangium ochraceum that contains two caspase-like proteases, SAVED-CHAT
and PCaspase (prokaryotic caspase), co-opted from a cyclic oligonucleotide–based
antiphage signaling system (CBASS). Cyclic tri–adenosine monophosphate (AMP)–induced
oligomerization of SAVED-CHAT activates proteolytic activity of the CHAT domains,
which specifically cleave and activate PCaspase. Subsequently, activated PCaspase
cleaves a multitude of proteins, which results in a strong interference phenotype
in vivo in Escherichia coli. Taken together, our findings reveal how a CRISPR-Cas–based
detection of a target RNA triggers a cascade of caspase-associated proteolytic
activities.
acknowledgement: We thank R. Fregoso Ocampo for assistance with negative-stain EM
imaging. This work was funded by Dutch Research Council (NWO) VIDI grant VI.Vidi.203.074
(R.H.J.S.), NWO Spinoza grant SPI 93-537 (J.v.d.O.), European Research Council (ERC)
Advanced grant ERC-AdG-834279 (J.v.d.O.), ERC CoG grant 817834 (T.J.G.E.), NWO VICI
grant VI.C.192.016 (T.J.G.E.), Volkswagen Foundation grant 96725 (T.J.G.E.), National
Institute of General Medical Sciences of the National Institutes of Health grant
R35GM138348 (D.W.T.), Welch Foundation research grant F-1938 (D.W.T.), a Robert
J. Kleberg, Jr. And Helen C. Kleberg Foundation medical research grant (D.W.T.),
and American Cancer Society Research Scholar grant RSG-21-050-01-DMC (D.W.T.).
article_processing_charge: No
article_type: original
author:
- first_name: Jurre A.
full_name: Steens, Jurre A.
last_name: Steens
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
- first_name: Carl Raymund P.
full_name: Salazar, Carl Raymund P.
last_name: Salazar
- first_name: Caglar
full_name: Yildiz, Caglar
last_name: Yildiz
- first_name: Afonso M.
full_name: Amieiro, Afonso M.
last_name: Amieiro
- first_name: Stephan
full_name: Köstlbacher, Stephan
last_name: Köstlbacher
- first_name: Stijn H.P.
full_name: Prinsen, Stijn H.P.
last_name: Prinsen
- first_name: Constantinos
full_name: Patinios, Constantinos
last_name: Patinios
- first_name: Andreas
full_name: Bardis, Andreas
last_name: Bardis
- first_name: Arjan
full_name: Barendregt, Arjan
last_name: Barendregt
- first_name: Richard A.
full_name: Scheltema, Richard A.
last_name: Scheltema
- first_name: Thijs J.G.
full_name: Ettema, Thijs J.G.
last_name: Ettema
- first_name: John
full_name: van der Oost, John
last_name: van der Oost
- first_name: David W.
full_name: Taylor, David W.
last_name: Taylor
- first_name: Raymond H.J.
full_name: Staals, Raymond H.J.
last_name: Staals
citation:
ama: Steens JA, Bravo JPK, Salazar CRP, et al. Type III-B CRISPR-Cas cascade of
proteolytic cleavages. Science. 2024;383(6682):512-519. doi:10.1126/science.adk0378
apa: Steens, J. A., Bravo, J. P. K., Salazar, C. R. P., Yildiz, C., Amieiro, A.
M., Köstlbacher, S., … Staals, R. H. J. (2024). Type III-B CRISPR-Cas cascade
of proteolytic cleavages. Science. American Association for the Advancement
of Science. https://doi.org/10.1126/science.adk0378
chicago: Steens, Jurre A., Jack Peter Kelly Bravo, Carl Raymund P. Salazar, Caglar
Yildiz, Afonso M. Amieiro, Stephan Köstlbacher, Stijn H.P. Prinsen, et al. “Type
III-B CRISPR-Cas Cascade of Proteolytic Cleavages.” Science. American Association
for the Advancement of Science, 2024. https://doi.org/10.1126/science.adk0378.
ieee: J. A. Steens et al., “Type III-B CRISPR-Cas cascade of proteolytic
cleavages,” Science, vol. 383, no. 6682. American Association for the Advancement
of Science, pp. 512–519, 2024.
ista: Steens JA, Bravo JPK, Salazar CRP, Yildiz C, Amieiro AM, Köstlbacher S, Prinsen
SHP, Patinios C, Bardis A, Barendregt A, Scheltema RA, Ettema TJG, van der Oost
J, Taylor DW, Staals RHJ. 2024. Type III-B CRISPR-Cas cascade of proteolytic cleavages.
Science. 383(6682), 512–519.
mla: Steens, Jurre A., et al. “Type III-B CRISPR-Cas Cascade of Proteolytic Cleavages.”
Science, vol. 383, no. 6682, American Association for the Advancement of
Science, 2024, pp. 512–19, doi:10.1126/science.adk0378.
short: J.A. Steens, J.P.K. Bravo, C.R.P. Salazar, C. Yildiz, A.M. Amieiro, S. Köstlbacher,
S.H.P. Prinsen, C. Patinios, A. Bardis, A. Barendregt, R.A. Scheltema, T.J.G.
Ettema, J. van der Oost, D.W. Taylor, R.H.J. Staals, Science 383 (2024) 512–519.
date_created: 2024-06-04T06:41:26Z
date_published: 2024-02-01T00:00:00Z
date_updated: 2025-09-24T08:31:45Z
day: '01'
doi: 10.1126/science.adk0378
extern: '1'
external_id:
pmid:
- '38301007'
intvolume: ' 383'
issue: '6682'
language:
- iso: eng
main_file_link:
- open_access: '1'
url: https://doi.org/10.1101/2023.06.23.546230
month: '02'
oa: 1
oa_version: Preprint
page: 512-519
pmid: 1
publication: Science
publication_identifier:
eissn:
- 1095-9203
issn:
- 0036-8075
publication_status: published
publisher: American Association for the Advancement of Science
quality_controlled: '1'
scopus_import: '1'
status: public
title: Type III-B CRISPR-Cas cascade of proteolytic cleavages
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 383
year: '2024'
...
---
_id: '17113'
abstract:
- lang: eng
text: CRISPR-Cas9 is a powerful tool for genome editing, but the strict requirement
for an NGG protospacer-adjacent motif (PAM) sequence immediately next to the DNA
target limits the number of editable genes. Recently developed Cas9 variants have
been engineered with relaxed PAM requirements, including SpG-Cas9 (SpG) and the
nearly PAM-less SpRY-Cas9 (SpRY). However, the molecular mechanisms of how SpRY
recognizes all potential PAM sequences remains unclear. Here, we combine structural
and biochemical approaches to determine how SpRY interrogates DNA and recognizes
target sites. Divergent PAM sequences can be accommodated through conformational
flexibility within the PAM-interacting region, which facilitates tight binding
to off-target DNA sequences. Nuclease activation occurs ~1000-fold slower than
for Streptococcus pyogenes Cas9, enabling us to directly
visualize multiple on-pathway intermediate states. Experiments with SpG position
it as an intermediate enzyme between Cas9 and SpRY. Our findings shed light on
the molecular mechanisms of PAMless genome editing.
article_number: '3663'
article_processing_charge: Yes
article_type: original
author:
- first_name: Grace N.
full_name: Hibshman, Grace N.
last_name: Hibshman
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
- first_name: Matthew M.
full_name: Hooper, Matthew M.
last_name: Hooper
- first_name: Tyler L.
full_name: Dangerfield, Tyler L.
last_name: Dangerfield
- first_name: Hongshan
full_name: Zhang, Hongshan
last_name: Zhang
- first_name: Ilya J.
full_name: Finkelstein, Ilya J.
last_name: Finkelstein
- first_name: Kenneth A.
full_name: Johnson, Kenneth A.
last_name: Johnson
- first_name: David W.
full_name: Taylor, David W.
last_name: Taylor
citation:
ama: Hibshman GN, Bravo JPK, Hooper MM, et al. Unraveling the mechanisms of PAMless
DNA interrogation by SpRY-Cas9. Nature Communications. 2024;15. doi:10.1038/s41467-024-47830-3
apa: Hibshman, G. N., Bravo, J. P. K., Hooper, M. M., Dangerfield, T. L., Zhang,
H., Finkelstein, I. J., … Taylor, D. W. (2024). Unraveling the mechanisms of PAMless
DNA interrogation by SpRY-Cas9. Nature Communications. Springer Nature.
https://doi.org/10.1038/s41467-024-47830-3
chicago: Hibshman, Grace N., Jack Peter Kelly Bravo, Matthew M. Hooper, Tyler L.
Dangerfield, Hongshan Zhang, Ilya J. Finkelstein, Kenneth A. Johnson, and David
W. Taylor. “Unraveling the Mechanisms of PAMless DNA Interrogation by SpRY-Cas9.”
Nature Communications. Springer Nature, 2024. https://doi.org/10.1038/s41467-024-47830-3.
ieee: G. N. Hibshman et al., “Unraveling the mechanisms of PAMless DNA interrogation
by SpRY-Cas9,” Nature Communications, vol. 15. Springer Nature, 2024.
ista: Hibshman GN, Bravo JPK, Hooper MM, Dangerfield TL, Zhang H, Finkelstein IJ,
Johnson KA, Taylor DW. 2024. Unraveling the mechanisms of PAMless DNA interrogation
by SpRY-Cas9. Nature Communications. 15, 3663.
mla: Hibshman, Grace N., et al. “Unraveling the Mechanisms of PAMless DNA Interrogation
by SpRY-Cas9.” Nature Communications, vol. 15, 3663, Springer Nature, 2024,
doi:10.1038/s41467-024-47830-3.
short: G.N. Hibshman, J.P.K. Bravo, M.M. Hooper, T.L. Dangerfield, H. Zhang, I.J.
Finkelstein, K.A. Johnson, D.W. Taylor, Nature Communications 15 (2024).
date_created: 2024-06-04T06:42:07Z
date_published: 2024-04-30T00:00:00Z
date_updated: 2024-10-14T12:34:26Z
day: '30'
doi: 10.1038/s41467-024-47830-3
extern: '1'
external_id:
pmid:
- '38688943'
intvolume: ' 15'
language:
- iso: eng
main_file_link:
- open_access: '1'
url: https://doi.org/10.1038/s41467-024-47830-3
month: '04'
oa: 1
oa_version: Published Version
pmid: 1
publication: Nature Communications
publication_identifier:
issn:
- 2041-1723
publication_status: published
publisher: Springer Nature
quality_controlled: '1'
scopus_import: '1'
status: public
title: Unraveling the mechanisms of PAMless DNA interrogation by SpRY-Cas9
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 15
year: '2024'
...
---
_id: '17114'
abstract:
- lang: eng
text: CRISPR-Cas are adaptive immune systems in bacteria and archaea that utilize
CRISPR RNA-guided surveillance complexes to target complementary RNA or DNA for
destruction1–5. Target RNA cleavage at regular intervals
is characteristic of type III effector complexes6–8. Here,
we determine the structures of the Synechocystis type
III-Dv complex, an apparent evolutionary intermediate from multi-protein to single-protein
type III effectors9,10, in pre- and post-cleavage states.
The structures show how multi-subunit fusion proteins in the effector are tethered
together in an unusual arrangement to assemble into an active and programmable
RNA endonuclease and how the effector utilizes a distinct mechanism for target
RNA seeding from other type III effectors. Using structural, biochemical, and
quantum/classical molecular dynamics simulation, we study the structure and dynamics
of the three catalytic sites, where a 2′-OH of the ribose on the target RNA acts
as a nucleophile for in line self-cleavage of the upstream scissile phosphate.
Strikingly, the arrangement at the catalytic residues of most type III complexes
resembles the active site of ribozymes, including the hammerhead, pistol, and
Varkud satellite ribozymes. Our work provides detailed molecular insight into
the mechanisms of RNA targeting and cleavage by an important intermediate in the
evolution of type III effector complexes.
article_number: '3324'
article_processing_charge: Yes
article_type: original
author:
- first_name: Evan A.
full_name: Schwartz, Evan A.
last_name: Schwartz
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
- first_name: Mohd
full_name: Ahsan, Mohd
last_name: Ahsan
- first_name: Luis A.
full_name: Macias, Luis A.
last_name: Macias
- first_name: Caitlyn L.
full_name: McCafferty, Caitlyn L.
last_name: McCafferty
- first_name: Tyler L.
full_name: Dangerfield, Tyler L.
last_name: Dangerfield
- first_name: Jada N.
full_name: Walker, Jada N.
last_name: Walker
- first_name: Jennifer S.
full_name: Brodbelt, Jennifer S.
last_name: Brodbelt
- first_name: Giulia
full_name: Palermo, Giulia
last_name: Palermo
- first_name: Peter C.
full_name: Fineran, Peter C.
last_name: Fineran
- first_name: Robert D.
full_name: Fagerlund, Robert D.
last_name: Fagerlund
- first_name: David W.
full_name: Taylor, David W.
last_name: Taylor
citation:
ama: Schwartz EA, Bravo JPK, Ahsan M, et al. RNA targeting and cleavage by the type
III-Dv CRISPR effector complex. Nature Communications. 2024;15. doi:10.1038/s41467-024-47506-y
apa: Schwartz, E. A., Bravo, J. P. K., Ahsan, M., Macias, L. A., McCafferty, C.
L., Dangerfield, T. L., … Taylor, D. W. (2024). RNA targeting and cleavage by
the type III-Dv CRISPR effector complex. Nature Communications. Springer
Nature. https://doi.org/10.1038/s41467-024-47506-y
chicago: Schwartz, Evan A., Jack Peter Kelly Bravo, Mohd Ahsan, Luis A. Macias,
Caitlyn L. McCafferty, Tyler L. Dangerfield, Jada N. Walker, et al. “RNA Targeting
and Cleavage by the Type III-Dv CRISPR Effector Complex.” Nature Communications.
Springer Nature, 2024. https://doi.org/10.1038/s41467-024-47506-y.
ieee: E. A. Schwartz et al., “RNA targeting and cleavage by the type III-Dv
CRISPR effector complex,” Nature Communications, vol. 15. Springer Nature,
2024.
ista: Schwartz EA, Bravo JPK, Ahsan M, Macias LA, McCafferty CL, Dangerfield TL,
Walker JN, Brodbelt JS, Palermo G, Fineran PC, Fagerlund RD, Taylor DW. 2024.
RNA targeting and cleavage by the type III-Dv CRISPR effector complex. Nature
Communications. 15, 3324.
mla: Schwartz, Evan A., et al. “RNA Targeting and Cleavage by the Type III-Dv CRISPR
Effector Complex.” Nature Communications, vol. 15, 3324, Springer Nature,
2024, doi:10.1038/s41467-024-47506-y.
short: E.A. Schwartz, J.P.K. Bravo, M. Ahsan, L.A. Macias, C.L. McCafferty, T.L.
Dangerfield, J.N. Walker, J.S. Brodbelt, G. Palermo, P.C. Fineran, R.D. Fagerlund,
D.W. Taylor, Nature Communications 15 (2024).
date_created: 2024-06-04T06:43:02Z
date_published: 2024-04-18T00:00:00Z
date_updated: 2024-06-04T07:05:26Z
day: '18'
doi: 10.1038/s41467-024-47506-y
extern: '1'
external_id:
pmid:
- '38637512'
intvolume: ' 15'
language:
- iso: eng
main_file_link:
- open_access: '1'
url: https://doi.org/10.1038/s41467-024-47506-y
month: '04'
oa: 1
oa_version: Published Version
pmid: 1
publication: Nature Communications
publication_identifier:
issn:
- 2041-1723
publication_status: published
publisher: Springer Nature
quality_controlled: '1'
scopus_import: '1'
status: public
title: RNA targeting and cleavage by the type III-Dv CRISPR effector complex
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 15
year: '2024'
...
---
OA_place: repository
OA_type: green
_id: '17442'
abstract:
- lang: eng
text: "Although eukaryotic Argonautes have a pivotal role in post-transcriptional
gene regulation through nucleic acid cleavage, some short prokaryotic Argonaute
variants (pAgos) rely on auxiliary nuclease factors for efficient foreign DNA
degradation1. Here we reveal the activation pathway of the DNA defence module
DdmDE system, which rapidly eliminates small, multicopy plasmids from the Vibrio
cholerae seventh pandemic strain (7PET)2. Through a combination of cryo-electron
microscopy, biochemistry and in vivo plasmid clearance assays, we demonstrate
that DdmE is a catalytically inactive, DNA-guided, DNA-targeting pAgo with a distinctive
insertion domain. We observe that the helicase-nuclease DdmD transitions from
an autoinhibited, dimeric complex to a monomeric state upon loading of single-stranded
DNA targets. Furthermore, the complete structure of the DdmDE–guide–target handover
complex provides a comprehensive view into how DNA recognition triggers processive
plasmid destruction. Our work establishes a mechanistic foundation for how pAgos
utilize ancillary factors to achieve plasmid clearance, and provides insights
into anti-plasmid immunity in bacteria.\r\n\r\n"
acknowledgement: We thank K. Kiernan, G. Hibshman and I. Strohkendl for insightful
discussions and comments on the manuscript, and R. Lin for assistance with the ATPase
assay. Data were collected at the Sauer Structural Biology Laboratory at the University
of Texas at Austin. This work was supported in part by the National Institute of
General Medical Sciences (NIGMS) of the National Institutes of Health (NIH) R35GM138348
(to D.W.T.) and Welch Foundation research grant F-1938 (to D.W.T.).
article_processing_charge: No
article_type: original
author:
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
- first_name: Delisa A.
full_name: Ramos, Delisa A.
last_name: Ramos
- first_name: Rodrigo
full_name: Fregoso Ocampo, Rodrigo
last_name: Fregoso Ocampo
- first_name: Caiden
full_name: Ingram, Caiden
last_name: Ingram
- first_name: David W.
full_name: Taylor, David W.
last_name: Taylor
citation:
ama: Bravo JPK, Ramos DA, Fregoso Ocampo R, Ingram C, Taylor DW. Plasmid targeting
and destruction by the DdmDE bacterial defence system. Nature. 2024;630(8018):961-967.
doi:10.1038/s41586-024-07515-9
apa: Bravo, J. P. K., Ramos, D. A., Fregoso Ocampo, R., Ingram, C., & Taylor,
D. W. (2024). Plasmid targeting and destruction by the DdmDE bacterial defence
system. Nature. Springer Nature. https://doi.org/10.1038/s41586-024-07515-9
chicago: Bravo, Jack Peter Kelly, Delisa A. Ramos, Rodrigo Fregoso Ocampo, Caiden
Ingram, and David W. Taylor. “Plasmid Targeting and Destruction by the DdmDE Bacterial
Defence System.” Nature. Springer Nature, 2024. https://doi.org/10.1038/s41586-024-07515-9.
ieee: J. P. K. Bravo, D. A. Ramos, R. Fregoso Ocampo, C. Ingram, and D. W. Taylor,
“Plasmid targeting and destruction by the DdmDE bacterial defence system,” Nature,
vol. 630, no. 8018. Springer Nature, pp. 961–967, 2024.
ista: Bravo JPK, Ramos DA, Fregoso Ocampo R, Ingram C, Taylor DW. 2024. Plasmid
targeting and destruction by the DdmDE bacterial defence system. Nature. 630(8018),
961–967.
mla: Bravo, Jack Peter Kelly, et al. “Plasmid Targeting and Destruction by the DdmDE
Bacterial Defence System.” Nature, vol. 630, no. 8018, Springer Nature,
2024, pp. 961–67, doi:10.1038/s41586-024-07515-9.
short: J.P.K. Bravo, D.A. Ramos, R. Fregoso Ocampo, C. Ingram, D.W. Taylor, Nature
630 (2024) 961–967.
corr_author: '1'
date_created: 2024-08-19T09:41:18Z
date_published: 2024-06-27T00:00:00Z
date_updated: 2025-06-24T12:47:21Z
day: '27'
department:
- _id: JaBr
doi: 10.1038/s41586-024-07515-9
external_id:
pmid:
- '38740055'
intvolume: ' 630'
issue: '8018'
language:
- iso: eng
main_file_link:
- open_access: '1'
url: https://pmc.ncbi.nlm.nih.gov/articles/PMC11649018/
month: '06'
oa: 1
oa_version: Submitted Version
page: 961-967
pmid: 1
publication: Nature
publication_identifier:
eissn:
- 1476-4687
issn:
- 0028-0836
publication_status: published
publisher: Springer Nature
quality_controlled: '1'
scopus_import: '1'
status: public
title: Plasmid targeting and destruction by the DdmDE bacterial defence system
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 630
year: '2024'
...
---
OA_place: publisher
OA_type: free access
_id: '17494'
acknowledgement: I would like to thank K Kiernan for insightful comments and feedback.
J P K Bravo is supported by IST Austria.
article_processing_charge: No
article_type: letter_note
author:
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
citation:
ama: 'Bravo JPK. Anti-plasmid immunity: A key to pathogen success? Future Microbiology.
2024;19(15):1269-1272. doi:10.1080/17460913.2024.2389720'
apa: 'Bravo, J. P. K. (2024). Anti-plasmid immunity: A key to pathogen success?
Future Microbiology. Taylor & Francis. https://doi.org/10.1080/17460913.2024.2389720'
chicago: 'Bravo, Jack Peter Kelly. “Anti-Plasmid Immunity: A Key to Pathogen Success?”
Future Microbiology. Taylor & Francis, 2024. https://doi.org/10.1080/17460913.2024.2389720.'
ieee: 'J. P. K. Bravo, “Anti-plasmid immunity: A key to pathogen success?,” Future
Microbiology, vol. 19, no. 15. Taylor & Francis, pp. 1269–1272, 2024.'
ista: 'Bravo JPK. 2024. Anti-plasmid immunity: A key to pathogen success? Future
Microbiology. 19(15), 1269–1272.'
mla: 'Bravo, Jack Peter Kelly. “Anti-Plasmid Immunity: A Key to Pathogen Success?”
Future Microbiology, vol. 19, no. 15, Taylor & Francis, 2024, pp. 1269–72,
doi:10.1080/17460913.2024.2389720.'
short: J.P.K. Bravo, Future Microbiology 19 (2024) 1269–1272.
corr_author: '1'
date_created: 2024-09-05T07:32:00Z
date_published: 2024-10-01T00:00:00Z
date_updated: 2025-09-08T09:03:00Z
day: '01'
department:
- _id: JaBr
doi: 10.1080/17460913.2024.2389720
external_id:
isi:
- '001306115400001'
pmid:
- '39230568'
has_accepted_license: '1'
intvolume: ' 19'
isi: 1
issue: '15'
language:
- iso: eng
main_file_link:
- open_access: '1'
url: https://doi.org/10.1080/17460913.2024.2389720
month: '10'
oa: 1
oa_version: Published Version
page: 1269-1272
pmid: 1
publication: Future Microbiology
publication_identifier:
eissn:
- 1746-0921
issn:
- 1746-0913
publication_status: published
publisher: Taylor & Francis
quality_controlled: '1'
scopus_import: '1'
status: public
title: 'Anti-plasmid immunity: A key to pathogen success?'
type: journal_article
user_id: 317138e5-6ab7-11ef-aa6d-ffef3953e345
volume: 19
year: '2024'
...
---
_id: '15129'
abstract:
- lang: eng
text: Type I CRISPR-Cas systems employ multi-subunit Cascade effector complexes
to target foreign nucleic acids for destruction. Here, we present structures of
D. vulgaris type I-C Cascade at various stages of double-stranded (ds)DNA target
capture, revealing mechanisms that underpin PAM recognition and Cascade allosteric
activation. We uncover an interesting mechanism of non-target strand (NTS) DNA
stabilization via stacking interactions with the “belly” subunits, securing the
NTS in place. This “molecular seatbelt” mechanism facilitates efficient R-loop
formation and prevents dsDNA reannealing. Additionally, we provide structural
insights into how two anti-CRISPR (Acr) proteins utilize distinct strategies to
achieve a shared mechanism of type I-C Cascade inhibition by blocking PAM scanning.
These observations form a structural basis for directional R-loop formation and
reveal how different Acr proteins have converged upon common molecular mechanisms
to efficiently shut down CRISPR immunity.
article_processing_charge: Yes (in subscription journal)
article_type: original
author:
- first_name: Roisin E.
full_name: O’Brien, Roisin E.
last_name: O’Brien
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
- first_name: Delisa
full_name: Ramos, Delisa
last_name: Ramos
- first_name: Grace N.
full_name: Hibshman, Grace N.
last_name: Hibshman
- first_name: Jacquelyn T.
full_name: Wright, Jacquelyn T.
last_name: Wright
- first_name: David W.
full_name: Taylor, David W.
last_name: Taylor
citation:
ama: O’Brien RE, Bravo JPK, Ramos D, Hibshman GN, Wright JT, Taylor DW. Structural
snapshots of R-loop formation by a type I-C CRISPR Cascade. Molecular Cell.
2023;83(5):746-758.e5. doi:10.1016/j.molcel.2023.01.024
apa: O’Brien, R. E., Bravo, J. P. K., Ramos, D., Hibshman, G. N., Wright, J. T.,
& Taylor, D. W. (2023). Structural snapshots of R-loop formation by a type
I-C CRISPR Cascade. Molecular Cell. Elsevier. https://doi.org/10.1016/j.molcel.2023.01.024
chicago: O’Brien, Roisin E., Jack Peter Kelly Bravo, Delisa Ramos, Grace N. Hibshman,
Jacquelyn T. Wright, and David W. Taylor. “Structural Snapshots of R-Loop Formation
by a Type I-C CRISPR Cascade.” Molecular Cell. Elsevier, 2023. https://doi.org/10.1016/j.molcel.2023.01.024.
ieee: R. E. O’Brien, J. P. K. Bravo, D. Ramos, G. N. Hibshman, J. T. Wright, and
D. W. Taylor, “Structural snapshots of R-loop formation by a type I-C CRISPR Cascade,”
Molecular Cell, vol. 83, no. 5. Elsevier, p. 746–758.e5, 2023.
ista: O’Brien RE, Bravo JPK, Ramos D, Hibshman GN, Wright JT, Taylor DW. 2023. Structural
snapshots of R-loop formation by a type I-C CRISPR Cascade. Molecular Cell. 83(5),
746–758.e5.
mla: O’Brien, Roisin E., et al. “Structural Snapshots of R-Loop Formation by a Type
I-C CRISPR Cascade.” Molecular Cell, vol. 83, no. 5, Elsevier, 2023, p.
746–758.e5, doi:10.1016/j.molcel.2023.01.024.
short: R.E. O’Brien, J.P.K. Bravo, D. Ramos, G.N. Hibshman, J.T. Wright, D.W. Taylor,
Molecular Cell 83 (2023) 746–758.e5.
date_created: 2024-03-20T10:40:56Z
date_published: 2023-03-02T00:00:00Z
date_updated: 2024-06-04T06:33:54Z
day: '02'
doi: 10.1016/j.molcel.2023.01.024
extern: '1'
external_id:
pmid:
- '36805026'
intvolume: ' 83'
issue: '5'
keyword:
- Cell Biology
- Molecular Biology
language:
- iso: eng
main_file_link:
- open_access: '1'
url: https://doi.org/10.1016/j.molcel.2023.01.024
month: '03'
oa: 1
oa_version: Published Version
page: 746-758.e5
pmid: 1
publication: Molecular Cell
publication_identifier:
issn:
- 1097-2765
publication_status: published
publisher: Elsevier
quality_controlled: '1'
scopus_import: '1'
status: public
title: Structural snapshots of R-loop formation by a type I-C CRISPR Cascade
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 83
year: '2023'
...
---
_id: '15130'
abstract:
- lang: eng
text: Cas12a2 is a CRISPR-associated nuclease that performs RNA-guided, sequence-nonspecific
degradation of single-stranded RNA, single-stranded DNA and double-stranded DNA
following recognition of a complementary RNA target, culminating in abortive infection1.
Here we report structures of Cas12a2 in binary, ternary and quaternary complexes
to reveal a complete activation pathway. Our structures reveal that Cas12a2 is
autoinhibited until binding a cognate RNA target, which exposes the RuvC active
site within a large, positively charged cleft. Double-stranded DNA substrates
are captured through duplex distortion and local melting, stabilized by pairs
of ‘aromatic clamp’ residues that are crucial for double-stranded DNA degradation
and in vivo immune system function. Our work provides a structural basis for this
mechanism of abortive infection to achieve population-level immunity, which can
be leveraged to create rational mutants that degrade a spectrum of collateral
substrates.
article_processing_charge: Yes (in subscription journal)
article_type: original
author:
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
- first_name: Thomson
full_name: Hallmark, Thomson
last_name: Hallmark
- first_name: Bronson
full_name: Naegle, Bronson
last_name: Naegle
- first_name: Chase L.
full_name: Beisel, Chase L.
last_name: Beisel
- first_name: Ryan N.
full_name: Jackson, Ryan N.
last_name: Jackson
- first_name: David W.
full_name: Taylor, David W.
last_name: Taylor
citation:
ama: Bravo JPK, Hallmark T, Naegle B, Beisel CL, Jackson RN, Taylor DW. RNA targeting
unleashes indiscriminate nuclease activity of CRISPR–Cas12a2. Nature. 2023;613(7944):582-587.
doi:10.1038/s41586-022-05560-w
apa: Bravo, J. P. K., Hallmark, T., Naegle, B., Beisel, C. L., Jackson, R. N., &
Taylor, D. W. (2023). RNA targeting unleashes indiscriminate nuclease activity
of CRISPR–Cas12a2. Nature. Springer Nature. https://doi.org/10.1038/s41586-022-05560-w
chicago: Bravo, Jack Peter Kelly, Thomson Hallmark, Bronson Naegle, Chase L. Beisel,
Ryan N. Jackson, and David W. Taylor. “RNA Targeting Unleashes Indiscriminate
Nuclease Activity of CRISPR–Cas12a2.” Nature. Springer Nature, 2023. https://doi.org/10.1038/s41586-022-05560-w.
ieee: J. P. K. Bravo, T. Hallmark, B. Naegle, C. L. Beisel, R. N. Jackson, and D.
W. Taylor, “RNA targeting unleashes indiscriminate nuclease activity of CRISPR–Cas12a2,”
Nature, vol. 613, no. 7944. Springer Nature, pp. 582–587, 2023.
ista: Bravo JPK, Hallmark T, Naegle B, Beisel CL, Jackson RN, Taylor DW. 2023. RNA
targeting unleashes indiscriminate nuclease activity of CRISPR–Cas12a2. Nature.
613(7944), 582–587.
mla: Bravo, Jack Peter Kelly, et al. “RNA Targeting Unleashes Indiscriminate Nuclease
Activity of CRISPR–Cas12a2.” Nature, vol. 613, no. 7944, Springer Nature,
2023, pp. 582–87, doi:10.1038/s41586-022-05560-w.
short: J.P.K. Bravo, T. Hallmark, B. Naegle, C.L. Beisel, R.N. Jackson, D.W. Taylor,
Nature 613 (2023) 582–587.
date_created: 2024-03-20T10:41:36Z
date_published: 2023-01-04T00:00:00Z
date_updated: 2024-06-04T06:30:59Z
day: '04'
doi: 10.1038/s41586-022-05560-w
extern: '1'
external_id:
pmid:
- '36599980'
intvolume: ' 613'
issue: '7944'
language:
- iso: eng
main_file_link:
- open_access: '1'
url: https://doi.org/10.1038/s41586-022-05560-w
month: '01'
oa: 1
oa_version: Published Version
page: 582-587
pmid: 1
publication: Nature
publication_identifier:
eissn:
- 1476-4687
issn:
- 0028-0836
publication_status: published
publisher: Springer Nature
quality_controlled: '1'
scopus_import: '1'
status: public
title: RNA targeting unleashes indiscriminate nuclease activity of CRISPR–Cas12a2
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 613
year: '2023'
...
---
_id: '15131'
abstract:
- lang: eng
text: RNA modifications are widespread in biology and abundant in ribosomal RNA.
However, the importance of these modifications is not well understood. We show
that methylation of a single nucleotide, in the catalytic center of the large
subunit, gates ribosome assembly. Massively parallel mutational scanning of the
essential nuclear GTPase Nog2 identified important interactions with rRNA, particularly
with the 2′-O-methylated A-site base Gm2922. We found
that methylation of G2922 is needed for assembly and efficient nuclear export
of the large subunit. Critically, we identified single amino acid changes in Nog2
that completely bypass dependence on G2922 methylation and used cryoelectron microscopy
to directly visualize how methylation flips Gm2922 into the active site channel
of Nog2. This work demonstrates that a single RNA modification is a critical checkpoint
in ribosome biogenesis, suggesting that such modifications can play an important
role in regulation and assembly of macromolecular machines.
article_processing_charge: Yes (in subscription journal)
article_type: original
author:
- first_name: James N.
full_name: Yelland, James N.
last_name: Yelland
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
- first_name: Joshua J.
full_name: Black, Joshua J.
last_name: Black
- first_name: David W.
full_name: Taylor, David W.
last_name: Taylor
- first_name: Arlen W.
full_name: Johnson, Arlen W.
last_name: Johnson
citation:
ama: Yelland JN, Bravo JPK, Black JJ, Taylor DW, Johnson AW. A single 2′-O-methylation
of ribosomal RNA gates assembly of a functional ribosome. Nature Structural
& Molecular Biology. 2022;30:91-98. doi:10.1038/s41594-022-00891-8
apa: Yelland, J. N., Bravo, J. P. K., Black, J. J., Taylor, D. W., & Johnson,
A. W. (2022). A single 2′-O-methylation of ribosomal RNA gates assembly of a functional
ribosome. Nature Structural & Molecular Biology. Springer Nature. https://doi.org/10.1038/s41594-022-00891-8
chicago: Yelland, James N., Jack Peter Kelly Bravo, Joshua J. Black, David W. Taylor,
and Arlen W. Johnson. “A Single 2′-O-Methylation of Ribosomal RNA Gates Assembly
of a Functional Ribosome.” Nature Structural & Molecular Biology. Springer
Nature, 2022. https://doi.org/10.1038/s41594-022-00891-8.
ieee: J. N. Yelland, J. P. K. Bravo, J. J. Black, D. W. Taylor, and A. W. Johnson,
“A single 2′-O-methylation of ribosomal RNA gates assembly of a functional ribosome,”
Nature Structural & Molecular Biology, vol. 30. Springer Nature, pp.
91–98, 2022.
ista: Yelland JN, Bravo JPK, Black JJ, Taylor DW, Johnson AW. 2022. A single 2′-O-methylation
of ribosomal RNA gates assembly of a functional ribosome. Nature Structural &
Molecular Biology. 30, 91–98.
mla: Yelland, James N., et al. “A Single 2′-O-Methylation of Ribosomal RNA Gates
Assembly of a Functional Ribosome.” Nature Structural & Molecular Biology,
vol. 30, Springer Nature, 2022, pp. 91–98, doi:10.1038/s41594-022-00891-8.
short: J.N. Yelland, J.P.K. Bravo, J.J. Black, D.W. Taylor, A.W. Johnson, Nature
Structural & Molecular Biology 30 (2022) 91–98.
date_created: 2024-03-20T10:41:45Z
date_published: 2022-12-19T00:00:00Z
date_updated: 2024-06-04T06:27:09Z
day: '19'
doi: 10.1038/s41594-022-00891-8
extern: '1'
external_id:
pmid:
- '36536102'
intvolume: ' 30'
keyword:
- Molecular Biology
- Structural Biology
language:
- iso: eng
main_file_link:
- open_access: '1'
url: https://doi.org/10.1038/s41594-022-00891-8
month: '12'
oa: 1
oa_version: Published Version
page: 91-98
pmid: 1
publication: Nature Structural & Molecular Biology
publication_identifier:
eissn:
- 1545-9985
issn:
- 1545-9993
publication_status: published
publisher: Springer Nature
quality_controlled: '1'
scopus_import: '1'
status: public
title: A single 2′-O-methylation of ribosomal RNA gates assembly of a functional ribosome
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 30
year: '2022'
...
---
_id: '15132'
abstract:
- lang: eng
text: Clustered regularly interspaced short palindromic repeats - CRISPR-associated
protein (CRISPR-Cas) systems are a critical component of the bacterial adaptive
immune response. Since the discovery that they can be reengineered as programmable
RNA-guided nucleases, there has been significant interest in using these systems
to perform diverse and precise genetic manipulations. Here, we outline recent
advances in the mechanistic understanding of CRISPR-Cas9, how these findings have
been leveraged in the rational redesign of Cas9 variants with altered activities,
and how these novel tools can be exploited for biotechnology and therapeutics.
We also discuss the potential of the ubiquitous, yet often-overlooked, multisubunit
CRISPR effector complexes for large-scale genomic deletions. Furthermore, we highlight
how future structural studies will bolster these technologies.
article_number: '102839'
article_processing_charge: No
article_type: review
author:
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
- first_name: Grace N
full_name: Hibshman, Grace N
last_name: Hibshman
- first_name: David W
full_name: Taylor, David W
last_name: Taylor
citation:
ama: Bravo JPK, Hibshman GN, Taylor DW. Constructing next-generation CRISPR–Cas
tools from structural blueprints. Current Opinion in Biotechnology. 2022;78.
doi:10.1016/j.copbio.2022.102839
apa: Bravo, J. P. K., Hibshman, G. N., & Taylor, D. W. (2022). Constructing
next-generation CRISPR–Cas tools from structural blueprints. Current Opinion
in Biotechnology. Elsevier. https://doi.org/10.1016/j.copbio.2022.102839
chicago: Bravo, Jack Peter Kelly, Grace N Hibshman, and David W Taylor. “Constructing
Next-Generation CRISPR–Cas Tools from Structural Blueprints.” Current Opinion
in Biotechnology. Elsevier, 2022. https://doi.org/10.1016/j.copbio.2022.102839.
ieee: J. P. K. Bravo, G. N. Hibshman, and D. W. Taylor, “Constructing next-generation
CRISPR–Cas tools from structural blueprints,” Current Opinion in Biotechnology,
vol. 78. Elsevier, 2022.
ista: Bravo JPK, Hibshman GN, Taylor DW. 2022. Constructing next-generation CRISPR–Cas
tools from structural blueprints. Current Opinion in Biotechnology. 78, 102839.
mla: Bravo, Jack Peter Kelly, et al. “Constructing Next-Generation CRISPR–Cas Tools
from Structural Blueprints.” Current Opinion in Biotechnology, vol. 78,
102839, Elsevier, 2022, doi:10.1016/j.copbio.2022.102839.
short: J.P.K. Bravo, G.N. Hibshman, D.W. Taylor, Current Opinion in Biotechnology
78 (2022).
date_created: 2024-03-20T10:41:53Z
date_published: 2022-12-01T00:00:00Z
date_updated: 2024-10-14T12:34:11Z
day: '01'
doi: 10.1016/j.copbio.2022.102839
extern: '1'
external_id:
pmid:
- '36371895'
intvolume: ' 78'
keyword:
- Biomedical Engineering
- Bioengineering
- Biotechnology
language:
- iso: eng
month: '12'
oa_version: None
pmid: 1
publication: Current Opinion in Biotechnology
publication_identifier:
issn:
- 0958-1669
publication_status: published
publisher: Elsevier
quality_controlled: '1'
scopus_import: '1'
status: public
title: Constructing next-generation CRISPR–Cas tools from structural blueprints
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 78
year: '2022'
...
---
_id: '15133'
abstract:
- lang: eng
text: In the evolutionary arms race against phage, bacteria have assembled a diverse
arsenal of antiviral immune strategies. While the recently discovered DISARM (Defense
Island System Associated with Restriction-Modification) systems can provide protection
against a wide range of phage, the molecular mechanisms that underpin broad antiviral
targeting but avoiding autoimmunity remain enigmatic. Here, we report cryo-EM
structures of the core DISARM complex, DrmAB, both alone and in complex with an
unmethylated phage DNA mimetic. These structures reveal that DrmAB core complex
is autoinhibited by a trigger loop (TL) within DrmA and binding to DNA substrates
containing a 5′ overhang dislodges the TL, initiating a long-range structural
rearrangement for DrmAB activation. Together with structure-guided in vivo studies,
our work provides insights into the mechanism of phage DNA recognition and specific
activation of this widespread antiviral defense system.
article_number: '2987'
article_processing_charge: Yes
article_type: original
author:
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
- first_name: Cristian
full_name: Aparicio-Maldonado, Cristian
last_name: Aparicio-Maldonado
- first_name: Franklin L.
full_name: Nobrega, Franklin L.
last_name: Nobrega
- first_name: Stan J. J.
full_name: Brouns, Stan J. J.
last_name: Brouns
- first_name: David W.
full_name: Taylor, David W.
last_name: Taylor
citation:
ama: Bravo JPK, Aparicio-Maldonado C, Nobrega FL, Brouns SJJ, Taylor DW. Structural
basis for broad anti-phage immunity by DISARM. Nature Communications. 2022;13.
doi:10.1038/s41467-022-30673-1
apa: Bravo, J. P. K., Aparicio-Maldonado, C., Nobrega, F. L., Brouns, S. J. J.,
& Taylor, D. W. (2022). Structural basis for broad anti-phage immunity by
DISARM. Nature Communications. Springer Nature. https://doi.org/10.1038/s41467-022-30673-1
chicago: Bravo, Jack Peter Kelly, Cristian Aparicio-Maldonado, Franklin L. Nobrega,
Stan J. J. Brouns, and David W. Taylor. “Structural Basis for Broad Anti-Phage
Immunity by DISARM.” Nature Communications. Springer Nature, 2022. https://doi.org/10.1038/s41467-022-30673-1.
ieee: J. P. K. Bravo, C. Aparicio-Maldonado, F. L. Nobrega, S. J. J. Brouns, and
D. W. Taylor, “Structural basis for broad anti-phage immunity by DISARM,” Nature
Communications, vol. 13. Springer Nature, 2022.
ista: Bravo JPK, Aparicio-Maldonado C, Nobrega FL, Brouns SJJ, Taylor DW. 2022.
Structural basis for broad anti-phage immunity by DISARM. Nature Communications.
13, 2987.
mla: Bravo, Jack Peter Kelly, et al. “Structural Basis for Broad Anti-Phage Immunity
by DISARM.” Nature Communications, vol. 13, 2987, Springer Nature, 2022,
doi:10.1038/s41467-022-30673-1.
short: J.P.K. Bravo, C. Aparicio-Maldonado, F.L. Nobrega, S.J.J. Brouns, D.W. Taylor,
Nature Communications 13 (2022).
date_created: 2024-03-20T10:41:59Z
date_published: 2022-05-27T00:00:00Z
date_updated: 2024-06-04T06:16:38Z
day: '27'
doi: 10.1038/s41467-022-30673-1
extern: '1'
external_id:
pmid:
- '35624106'
intvolume: ' 13'
keyword:
- General Physics and Astronomy
- General Biochemistry
- Genetics and Molecular Biology
- General Chemistry
- Multidisciplinary
language:
- iso: eng
main_file_link:
- open_access: '1'
url: https://doi.org/10.1038/s41467-022-30673-1
month: '05'
oa: 1
oa_version: Published Version
pmid: 1
publication: Nature Communications
publication_identifier:
issn:
- 2041-1723
publication_status: published
publisher: Springer Nature
quality_controlled: '1'
scopus_import: '1'
status: public
title: Structural basis for broad anti-phage immunity by DISARM
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 13
year: '2022'
...
---
_id: '15134'
abstract:
- lang: eng
text: CRISPR-Cas systems are adaptive immune systems that protect prokaryotes from
foreign nucleic acids, such as bacteriophages. Two of the most prevalent CRISPR-Cas
systems include type I and type III. Interestingly, the type I-D interference
proteins contain characteristic features of both type I and type III systems.
Here, we present the structures of type I-D Cascade bound to both a double-stranded
(ds)DNA and a single-stranded (ss)RNA target at 2.9 and 3.1 Å, respectively. We
show that type I-D Cascade is capable of specifically binding ssRNA and reveal
how PAM recognition of dsDNA targets initiates long-range structural rearrangements
that likely primes Cas10d for Cas3′ binding and subsequent non-target strand DNA
cleavage. These structures allow us to model how binding of the anti-CRISPR protein
AcrID1 likely blocks target dsDNA binding via competitive inhibition of the DNA
substrate engagement with the Cas10d active site. This work elucidates the unique
mechanisms used by type I-D Cascade for discrimination of single-stranded and
double stranded targets. Thus, our data supports a model for the hybrid nature
of this complex with features of type III and type I systems.
article_number: '2829'
article_processing_charge: Yes
article_type: original
author:
- first_name: Evan A.
full_name: Schwartz, Evan A.
last_name: Schwartz
- first_name: Tess M.
full_name: McBride, Tess M.
last_name: McBride
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
- first_name: Daniel
full_name: Wrapp, Daniel
last_name: Wrapp
- first_name: Peter C.
full_name: Fineran, Peter C.
last_name: Fineran
- first_name: Robert D.
full_name: Fagerlund, Robert D.
last_name: Fagerlund
- first_name: David W.
full_name: Taylor, David W.
last_name: Taylor
citation:
ama: Schwartz EA, McBride TM, Bravo JPK, et al. Structural rearrangements allow
nucleic acid discrimination by type I-D Cascade. Nature Communications.
2022;13. doi:10.1038/s41467-022-30402-8
apa: Schwartz, E. A., McBride, T. M., Bravo, J. P. K., Wrapp, D., Fineran, P. C.,
Fagerlund, R. D., & Taylor, D. W. (2022). Structural rearrangements allow
nucleic acid discrimination by type I-D Cascade. Nature Communications.
Springer Nature. https://doi.org/10.1038/s41467-022-30402-8
chicago: Schwartz, Evan A., Tess M. McBride, Jack Peter Kelly Bravo, Daniel Wrapp,
Peter C. Fineran, Robert D. Fagerlund, and David W. Taylor. “Structural Rearrangements
Allow Nucleic Acid Discrimination by Type I-D Cascade.” Nature Communications.
Springer Nature, 2022. https://doi.org/10.1038/s41467-022-30402-8.
ieee: E. A. Schwartz et al., “Structural rearrangements allow nucleic acid
discrimination by type I-D Cascade,” Nature Communications, vol. 13. Springer
Nature, 2022.
ista: Schwartz EA, McBride TM, Bravo JPK, Wrapp D, Fineran PC, Fagerlund RD, Taylor
DW. 2022. Structural rearrangements allow nucleic acid discrimination by type
I-D Cascade. Nature Communications. 13, 2829.
mla: Schwartz, Evan A., et al. “Structural Rearrangements Allow Nucleic Acid Discrimination
by Type I-D Cascade.” Nature Communications, vol. 13, 2829, Springer Nature,
2022, doi:10.1038/s41467-022-30402-8.
short: E.A. Schwartz, T.M. McBride, J.P.K. Bravo, D. Wrapp, P.C. Fineran, R.D. Fagerlund,
D.W. Taylor, Nature Communications 13 (2022).
date_created: 2024-03-20T10:42:05Z
date_published: 2022-05-20T00:00:00Z
date_updated: 2024-06-04T06:14:28Z
day: '20'
doi: 10.1038/s41467-022-30402-8
extern: '1'
external_id:
pmid:
- '35595728'
intvolume: ' 13'
keyword:
- General Physics and Astronomy
- General Biochemistry
- Genetics and Molecular Biology
- General Chemistry
- Multidisciplinary
language:
- iso: eng
main_file_link:
- open_access: '1'
url: https://doi.org/10.1038/s41467-022-30402-8
month: '05'
oa: 1
oa_version: Published Version
pmid: 1
publication: Nature Communications
publication_identifier:
issn:
- 2041-1723
publication_status: published
publisher: Springer Nature
quality_controlled: '1'
scopus_import: '1'
status: public
title: Structural rearrangements allow nucleic acid discrimination by type I-D Cascade
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 13
year: '2022'
...
---
_id: '15136'
abstract:
- lang: eng
text: CRISPR–Cas9 as a programmable genome editing tool is hindered by off-target
DNA cleavage1,2,3,4, and the underlying mechanisms by which Cas9 recognizes mismatches
are poorly understood5,6,7. Although Cas9 variants with greater discrimination
against mismatches have been designed8,9,10, these suffer from substantially reduced
rates of on-target DNA cleavage5,11. Here we used kinetics-guided cryo-electron
microscopy to determine the structure of Cas9 at different stages of mismatch
cleavage. We observed a distinct, linear conformation of the guide RNA–DNA duplex
formed in the presence of mismatches, which prevents Cas9 activation. Although
the canonical kinked guide RNA–DNA duplex conformation facilitates DNA cleavage,
we observe that substrates that contain mismatches distal to the protospacer adjacent
motif are stabilized by reorganization of a loop in the RuvC domain. Mutagenesis
of mismatch-stabilizing residues reduces off-target DNA cleavage but maintains
rapid on-target DNA cleavage. By targeting regions that are exclusively involved
in mismatch tolerance, we provide a proof of concept for the design of next-generation
high-fidelity Cas9 variants.
article_processing_charge: Yes (in subscription journal)
article_type: original
author:
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
- first_name: Mu-Sen
full_name: Liu, Mu-Sen
last_name: Liu
- first_name: Grace N.
full_name: Hibshman, Grace N.
last_name: Hibshman
- first_name: Tyler L.
full_name: Dangerfield, Tyler L.
last_name: Dangerfield
- first_name: Kyungseok
full_name: Jung, Kyungseok
last_name: Jung
- first_name: Ryan S.
full_name: McCool, Ryan S.
last_name: McCool
- first_name: Kenneth A.
full_name: Johnson, Kenneth A.
last_name: Johnson
- first_name: David W.
full_name: Taylor, David W.
last_name: Taylor
citation:
ama: Bravo JPK, Liu M-S, Hibshman GN, et al. Structural basis for mismatch surveillance
by CRISPR–Cas9. Nature. 2022;603(7900):343-347. doi:10.1038/s41586-022-04470-1
apa: Bravo, J. P. K., Liu, M.-S., Hibshman, G. N., Dangerfield, T. L., Jung, K.,
McCool, R. S., … Taylor, D. W. (2022). Structural basis for mismatch surveillance
by CRISPR–Cas9. Nature. Springer Nature. https://doi.org/10.1038/s41586-022-04470-1
chicago: Bravo, Jack Peter Kelly, Mu-Sen Liu, Grace N. Hibshman, Tyler L. Dangerfield,
Kyungseok Jung, Ryan S. McCool, Kenneth A. Johnson, and David W. Taylor. “Structural
Basis for Mismatch Surveillance by CRISPR–Cas9.” Nature. Springer Nature,
2022. https://doi.org/10.1038/s41586-022-04470-1.
ieee: J. P. K. Bravo et al., “Structural basis for mismatch surveillance
by CRISPR–Cas9,” Nature, vol. 603, no. 7900. Springer Nature, pp. 343–347,
2022.
ista: Bravo JPK, Liu M-S, Hibshman GN, Dangerfield TL, Jung K, McCool RS, Johnson
KA, Taylor DW. 2022. Structural basis for mismatch surveillance by CRISPR–Cas9.
Nature. 603(7900), 343–347.
mla: Bravo, Jack Peter Kelly, et al. “Structural Basis for Mismatch Surveillance
by CRISPR–Cas9.” Nature, vol. 603, no. 7900, Springer Nature, 2022, pp.
343–47, doi:10.1038/s41586-022-04470-1.
short: J.P.K. Bravo, M.-S. Liu, G.N. Hibshman, T.L. Dangerfield, K. Jung, R.S. McCool,
K.A. Johnson, D.W. Taylor, Nature 603 (2022) 343–347.
date_created: 2024-03-20T10:42:21Z
date_published: 2022-03-02T00:00:00Z
date_updated: 2024-06-04T06:36:59Z
day: '02'
doi: 10.1038/s41586-022-04470-1
extern: '1'
external_id:
pmid:
- '35236982'
intvolume: ' 603'
issue: '7900'
language:
- iso: eng
main_file_link:
- open_access: '1'
url: https://doi.org/10.1038/s41586-022-04470-1
month: '03'
oa: 1
oa_version: Published Version
page: 343-347
pmid: 1
publication: Nature
publication_identifier:
eissn:
- 1476-4687
issn:
- 0028-0836
publication_status: published
publisher: Springer Nature
quality_controlled: '1'
related_material:
link:
- relation: erratum
url: https://doi.org/10.1038/s41586-022-04655-8
scopus_import: '1'
status: public
title: Structural basis for mismatch surveillance by CRISPR–Cas9
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 603
year: '2022'
...
---
_id: '15144'
article_processing_charge: No
article_type: letter_note
author:
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
citation:
ama: Bravo JPK. SuperFi-Cas9 exceeds fidelity, matches speed of original Cas9. Genetic
Engineering & Biotechnology News. 2022;42(4):12. doi:10.1089/gen.42.04.03
apa: Bravo, J. P. K. (2022). SuperFi-Cas9 exceeds fidelity, matches speed of original
Cas9. Genetic Engineering & Biotechnology News. Mary Ann Liebert. https://doi.org/10.1089/gen.42.04.03
chicago: Bravo, Jack Peter Kelly. “SuperFi-Cas9 Exceeds Fidelity, Matches Speed
of Original Cas9.” Genetic Engineering & Biotechnology News. Mary Ann
Liebert, 2022. https://doi.org/10.1089/gen.42.04.03.
ieee: J. P. K. Bravo, “SuperFi-Cas9 exceeds fidelity, matches speed of original
Cas9,” Genetic Engineering & Biotechnology News, vol. 42, no. 4. Mary
Ann Liebert, p. 12, 2022.
ista: Bravo JPK. 2022. SuperFi-Cas9 exceeds fidelity, matches speed of original
Cas9. Genetic Engineering & Biotechnology News. 42(4), 12.
mla: Bravo, Jack Peter Kelly. “SuperFi-Cas9 Exceeds Fidelity, Matches Speed of Original
Cas9.” Genetic Engineering & Biotechnology News, vol. 42, no. 4, Mary
Ann Liebert, 2022, p. 12, doi:10.1089/gen.42.04.03.
short: J.P.K. Bravo, Genetic Engineering & Biotechnology News 42 (2022) 12.
date_created: 2024-03-20T10:43:19Z
date_published: 2022-04-01T00:00:00Z
date_updated: 2024-10-14T12:32:14Z
day: '01'
doi: 10.1089/gen.42.04.03
extern: '1'
intvolume: ' 42'
issue: '4'
keyword:
- Management of Technology and Innovation
- Biomedical Engineering
- Bioengineering
- Biotechnology
language:
- iso: eng
month: '04'
oa_version: None
page: '12'
publication: Genetic Engineering & Biotechnology News
publication_identifier:
eissn:
- 1937-8661
issn:
- 1935-472X
publication_status: published
publisher: Mary Ann Liebert
quality_controlled: '1'
scopus_import: '1'
status: public
title: SuperFi-Cas9 exceeds fidelity, matches speed of original Cas9
type: journal_article
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 42
year: '2022'
...
---
_id: '17115'
abstract:
- lang: eng
text: Cascades are RNA-guided multi-subunit CRISPR-Cas surveillances complexes that
target foreign nucleic acids for destruction. Here, we present a 2.9-Å resolution
cryo-electron (cryo-EM) structure of the D. vulgaris
type I-C Cascade bound to a double-stranded (ds)DNA target. Our data shows how
the 5’-TTC-3’ protospacer adjacent motif (PAM) sequence is recognized, and provides
a unique mechanism through which the displaced, single-stranded non-target strand
(NTS) is stabilized via stacking interactions with protein subunits in order to
favor R-loop formation and prevent dsDNA re-annealing. Additionally, we provide
structural insights into how diverse anti-CRISPR (Acr) proteins utilize distinct
strategies to achieve a shared mechanism of type I-C Cascade inhibition by blocking
initial DNA binding. These observations provide a structural basis for directional
R-loop formation and reveal how divergent Acr proteins have converged upon common
molecular mechanisms to efficiently shut down CRISPR immunity.
article_processing_charge: No
author:
- first_name: Roisin E.
full_name: O’Brien, Roisin E.
last_name: O’Brien
- first_name: Jack Peter Kelly
full_name: Bravo, Jack Peter Kelly
id: 96aecfa5-8931-11ee-af30-aa6a5d6eee0e
last_name: Bravo
orcid: 0000-0003-0456-0753
- first_name: Delisa
full_name: Ramos, Delisa
last_name: Ramos
- first_name: Grace N.
full_name: Hibshman, Grace N.
last_name: Hibshman
- first_name: Jacquelyn T.
full_name: Wright, Jacquelyn T.
last_name: Wright
- first_name: David W.
full_name: Taylor, David W.
last_name: Taylor
citation:
ama: O’Brien RE, Bravo JPK, Ramos D, Hibshman GN, Wright JT, Taylor DW. Modes of
inhibition used by phage anti-CRISPRs to evade type I-C Cascade. bioRxiv.
2022. doi:10.1101/2022.06.15.496202
apa: O’Brien, R. E., Bravo, J. P. K., Ramos, D., Hibshman, G. N., Wright, J. T.,
& Taylor, D. W. (2022). Modes of inhibition used by phage anti-CRISPRs to
evade type I-C Cascade. bioRxiv. Cold Spring Harbor Laboratory. https://doi.org/10.1101/2022.06.15.496202
chicago: O’Brien, Roisin E., Jack Peter Kelly Bravo, Delisa Ramos, Grace N. Hibshman,
Jacquelyn T. Wright, and David W. Taylor. “Modes of Inhibition Used by Phage Anti-CRISPRs
to Evade Type I-C Cascade.” BioRxiv. Cold Spring Harbor Laboratory, 2022.
https://doi.org/10.1101/2022.06.15.496202.
ieee: R. E. O’Brien, J. P. K. Bravo, D. Ramos, G. N. Hibshman, J. T. Wright, and
D. W. Taylor, “Modes of inhibition used by phage anti-CRISPRs to evade type I-C
Cascade,” bioRxiv. Cold Spring Harbor Laboratory, 2022.
ista: O’Brien RE, Bravo JPK, Ramos D, Hibshman GN, Wright JT, Taylor DW. 2022. Modes
of inhibition used by phage anti-CRISPRs to evade type I-C Cascade. bioRxiv, 10.1101/2022.06.15.496202.
mla: O’Brien, Roisin E., et al. “Modes of Inhibition Used by Phage Anti-CRISPRs
to Evade Type I-C Cascade.” BioRxiv, Cold Spring Harbor Laboratory, 2022,
doi:10.1101/2022.06.15.496202.
short: R.E. O’Brien, J.P.K. Bravo, D. Ramos, G.N. Hibshman, J.T. Wright, D.W. Taylor,
BioRxiv (2022).
date_created: 2024-06-04T06:43:30Z
date_published: 2022-06-15T00:00:00Z
date_updated: 2024-06-04T07:03:02Z
day: '15'
doi: 10.1101/2022.06.15.496202
extern: '1'
language:
- iso: eng
main_file_link:
- open_access: '1'
url: https://doi.org/10.1101/2022.06.15.496202
month: '06'
oa: 1
oa_version: Preprint
publication: bioRxiv
publication_status: published
publisher: Cold Spring Harbor Laboratory
status: public
title: Modes of inhibition used by phage anti-CRISPRs to evade type I-C Cascade
type: preprint
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
year: '2022'
...