Structural organization of a Type III-A CRISPR effector subcomplex determined by X-ray crystallography and cryo-EM

Bryan W. Dorsey; Lei Huang; Alfonso Mondragón

doi:10.1093/nar/gkz079

Structural organization of a Type III-A CRISPR effector subcomplex determined by X-ray crystallography and cryo-EM

Bryan W. Dorsey, Lei Huang, Alfonso Mondragón^*

^*Corresponding author for this work

Molecular Biosciences

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

9 Scopus citations

Abstract

Clustered regularly interspaced short palindromic repeats (CRISPR) and their associated Cas proteins provide an immune-like response in many prokaryotes against extraneous nucleic acids. CRISPR-Cas systems are classified into different classes and types. Class 1 CRISPR-Cas systems form multi-protein effector complexes that includes a guide RNA (crRNA) used to identify the target for destruction. Here we present crystal structures of Staphylococcus epidermidis Type III-A CRISPR subunits Csm2 and Csm3 and a 5.2 Å resolution single-particle cryo-electron microscopy (cryo-EM) reconstruction of an in vivo assembled effector subcomplex including the crRNA. The structures help to clarify the quaternary architecture of Type III-A effector complexes, and provide details on crRNA binding, target RNA binding and cleavage, and intermolecular interactions essential for effector complex assembly. The structures allow a better understanding of the organization of Type III-A CRISPR effector complexes as well as highlighting the overall similarities and differences with other Class 1 effector complexes.

Original language	English (US)
Pages (from-to)	3765-3783
Number of pages	19
Journal	Nucleic acids research
Volume	47
Issue number	7
DOIs	https://doi.org/10.1093/nar/gkz079
State	Published - Apr 23 2019

Funding

National Institutes of Health (NIH) [R35 GM118108 to A.M.]; LS-CAT/Sector 21 at the Advanced Photon Source, Argonne National Laboratory was supported by the Michigan Economic Development Corporation and the Michigan Technology Tri-Corridor. Support from the R.H. Lurie Comprehensive Cancer Center of Northwestern University to the Structural Biology, Keck Biophysics, and High Throughput Analysis Facilities is acknowledged. The Gatan K2 DDE at Northwestern University was purchased with funds provided by the Chicago Biomedical Consortium with support from the Searle Funds at The Chicago Community Trust. The GE Healthcare Typhoon 9400 Flu-orimager at Northwestern University was purchased with funds provided by award [S10RR027842] from the National Center for Research Resources. Funding for open access charge: National Institutes of Health [R35 GM118108]. Conflict of interest statement. None declared.

ASJC Scopus subject areas

Genetics

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10.1093/nar/gkz079

Cite this

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title = "Structural organization of a Type III-A CRISPR effector subcomplex determined by X-ray crystallography and cryo-EM",

abstract = "Clustered regularly interspaced short palindromic repeats (CRISPR) and their associated Cas proteins provide an immune-like response in many prokaryotes against extraneous nucleic acids. CRISPR-Cas systems are classified into different classes and types. Class 1 CRISPR-Cas systems form multi-protein effector complexes that includes a guide RNA (crRNA) used to identify the target for destruction. Here we present crystal structures of Staphylococcus epidermidis Type III-A CRISPR subunits Csm2 and Csm3 and a 5.2 {\AA} resolution single-particle cryo-electron microscopy (cryo-EM) reconstruction of an in vivo assembled effector subcomplex including the crRNA. The structures help to clarify the quaternary architecture of Type III-A effector complexes, and provide details on crRNA binding, target RNA binding and cleavage, and intermolecular interactions essential for effector complex assembly. The structures allow a better understanding of the organization of Type III-A CRISPR effector complexes as well as highlighting the overall similarities and differences with other Class 1 effector complexes.",

author = "Dorsey, \{Bryan W.\} and Lei Huang and Alfonso Mondrag{\'o}n",

note = "Publisher Copyright: {\textcopyright} The Author(s) 2019. Published by Oxford University Press on behalf of Nucleic Acids Research.",

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AU - Dorsey, Bryan W.

AU - Huang, Lei

AU - Mondragón, Alfonso

N1 - Publisher Copyright: © The Author(s) 2019. Published by Oxford University Press on behalf of Nucleic Acids Research.

PY - 2019/4/23

Y1 - 2019/4/23

N2 - Clustered regularly interspaced short palindromic repeats (CRISPR) and their associated Cas proteins provide an immune-like response in many prokaryotes against extraneous nucleic acids. CRISPR-Cas systems are classified into different classes and types. Class 1 CRISPR-Cas systems form multi-protein effector complexes that includes a guide RNA (crRNA) used to identify the target for destruction. Here we present crystal structures of Staphylococcus epidermidis Type III-A CRISPR subunits Csm2 and Csm3 and a 5.2 Å resolution single-particle cryo-electron microscopy (cryo-EM) reconstruction of an in vivo assembled effector subcomplex including the crRNA. The structures help to clarify the quaternary architecture of Type III-A effector complexes, and provide details on crRNA binding, target RNA binding and cleavage, and intermolecular interactions essential for effector complex assembly. The structures allow a better understanding of the organization of Type III-A CRISPR effector complexes as well as highlighting the overall similarities and differences with other Class 1 effector complexes.

AB - Clustered regularly interspaced short palindromic repeats (CRISPR) and their associated Cas proteins provide an immune-like response in many prokaryotes against extraneous nucleic acids. CRISPR-Cas systems are classified into different classes and types. Class 1 CRISPR-Cas systems form multi-protein effector complexes that includes a guide RNA (crRNA) used to identify the target for destruction. Here we present crystal structures of Staphylococcus epidermidis Type III-A CRISPR subunits Csm2 and Csm3 and a 5.2 Å resolution single-particle cryo-electron microscopy (cryo-EM) reconstruction of an in vivo assembled effector subcomplex including the crRNA. The structures help to clarify the quaternary architecture of Type III-A effector complexes, and provide details on crRNA binding, target RNA binding and cleavage, and intermolecular interactions essential for effector complex assembly. The structures allow a better understanding of the organization of Type III-A CRISPR effector complexes as well as highlighting the overall similarities and differences with other Class 1 effector complexes.

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Structural organization of a Type III-A CRISPR effector subcomplex determined by X-ray crystallography and cryo-EM

Abstract

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CRISPR Complex Subunit Csm2 from Staphylococcus epidermidis RP62a

CRISPR Complex Subunit Csm3 from Staphylococcus epidermidis RP62a

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Structural organization of a Type III-A CRISPR effector subcomplex determined by X-ray crystallography and cryo-EM

Abstract

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ASJC Scopus subject areas

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Datasets

CRISPR Complex Subunit Csm2 from Staphylococcus epidermidis RP62a

CRISPR Complex Subunit Csm3 from Staphylococcus epidermidis RP62a

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