An enhanced CRISPR repressor for targeted mammalian gene regulation

Nan Cher Yeo; Alejandro Chavez; Alissa Lance-Byrne; Yingleong Chan; David Menn; Denitsa Milanova; Chih Chung Kuo; Xiaoge Guo; Sumana Sharma; Angela Tung; Ryan J. Cecchi; Marcelle Tuttle; Swechchha Pradhan; Elaine T. Lim; Noah Davidsohn; Mo R. Ebrahimkhani; James J. Collins; Nathan E. Lewis; Samira Kiani; George M. Church

doi:10.1038/s41592-018-0048-5

An enhanced CRISPR repressor for targeted mammalian gene regulation

Nan Cher Yeo, Alejandro Chavez, Alissa Lance-Byrne, Yingleong Chan, David Menn, Denitsa Milanova, Chih Chung Kuo, Xiaoge Guo, Sumana Sharma, Angela Tung, Ryan J. Cecchi, Marcelle Tuttle, Swechchha Pradhan, Elaine T. Lim, Noah Davidsohn, Mo R. Ebrahimkhani, James J. Collins, Nathan E. Lewis, Samira Kiani, George M. Church

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

220 Scopus citations

Abstract

The RNA-guided endonuclease Cas9 can be converted into a programmable transcriptional repressor, but inefficiencies in target-gene silencing have limited its utility. Here we describe an improved Cas9 repressor based on the C-terminal fusion of a rationally designed bipartite repressor domain, KRAB–MeCP2, to nuclease-dead Cas9. We demonstrate the system’s superiority in silencing coding and noncoding genes, simultaneously repressing a series of target genes, improving the results of single and dual guide RNA library screens, and enabling new architectures of synthetic genetic circuits.

Original language	English (US)
Pages (from-to)	611-616
Number of pages	6
Journal	Nature Methods
Volume	15
Issue number	8
DOIs	https://doi.org/10.1038/s41592-018-0048-5
State	Published - Aug 1 2018

ASJC Scopus subject areas

Biotechnology
Biochemistry
Molecular Biology
Cell Biology

Access to Document

10.1038/s41592-018-0048-5

Cite this

Yeo, N. C., Chavez, A., Lance-Byrne, A., Chan, Y., Menn, D., Milanova, D., Kuo, C. C., Guo, X., Sharma, S., Tung, A., Cecchi, R. J., Tuttle, M., Pradhan, S., Lim, E. T., Davidsohn, N., Ebrahimkhani, M. R., Collins, J. J., Lewis, N. E., Kiani, S., & Church, G. M. (2018). An enhanced CRISPR repressor for targeted mammalian gene regulation. Nature Methods, 15(8), 611-616. https://doi.org/10.1038/s41592-018-0048-5

Yeo, NC, Chavez, A, Lance-Byrne, A, Chan, Y, Menn, D, Milanova, D, Kuo, CC, Guo, X, Sharma, S, Tung, A, Cecchi, RJ, Tuttle, M, Pradhan, S, Lim, ET, Davidsohn, N, Ebrahimkhani, MR, Collins, JJ, Lewis, NE, Kiani, S & Church, GM 2018, 'An enhanced CRISPR repressor for targeted mammalian gene regulation', Nature Methods, vol. 15, no. 8, pp. 611-616. https://doi.org/10.1038/s41592-018-0048-5

@article{59e6eccb2a604949b5a75b704e1f8f29,

title = "An enhanced CRISPR repressor for targeted mammalian gene regulation",

abstract = "The RNA-guided endonuclease Cas9 can be converted into a programmable transcriptional repressor, but inefficiencies in target-gene silencing have limited its utility. Here we describe an improved Cas9 repressor based on the C-terminal fusion of a rationally designed bipartite repressor domain, KRAB–MeCP2, to nuclease-dead Cas9. We demonstrate the system{\textquoteright}s superiority in silencing coding and noncoding genes, simultaneously repressing a series of target genes, improving the results of single and dual guide RNA library screens, and enabling new architectures of synthetic genetic circuits.",

author = "Yeo, {Nan Cher} and Alejandro Chavez and Alissa Lance-Byrne and Yingleong Chan and David Menn and Denitsa Milanova and Kuo, {Chih Chung} and Xiaoge Guo and Sumana Sharma and Angela Tung and Cecchi, {Ryan J.} and Marcelle Tuttle and Swechchha Pradhan and Lim, {Elaine T.} and Noah Davidsohn and Ebrahimkhani, {Mo R.} and Collins, {James J.} and Lewis, {Nathan E.} and Samira Kiani and Church, {George M.}",

note = "Funding Information: This work was supported by the NIH (grants RM1 HG008525 and P50 HG005550 to G.M.C.), the National Cancer Institute (grant 5T32CA009216-34 to A.C.), the Burroughs Wellcome Fund (Career Award for Medical Scientists to A.C.), NIGMS (R35 GM119850 to N.E.L.), the Novo Nordisk Foundation (NNF10CC1016517 to N.E.L.), DARPA (Young Faculty Award D16AP00047 to S.K.), the Arizona State University Fulton Schools of Engineering startup fund (S.K.), and the Paul G. Allen Frontiers Group (J.J.C.). HEK293T cells were a gift from P. Mali (University of California, San Diego, San Diego, CA, USA). psPAX2 was a gift from D. Trono (Ecole Polytechnique Federale de Lausanne, Lausanne, Switzerland). pCMV-VSV-G was a gift from B. Weinberg (Massachusetts Institute of Technology, Boston, MA, USA). The plasmid containing a single gRNA library targeting essential genes was a gift from R. Bernards (The Netherlands Cancer Institute, Amsterdam, the Netherlands). Publisher Copyright: {\textcopyright} 2018, The Author(s).",

year = "2018",

month = aug,

day = "1",

doi = "10.1038/s41592-018-0048-5",

language = "English (US)",

volume = "15",

pages = "611--616",

journal = "Nature Methods",

issn = "1548-7091",

publisher = "Nature Publishing Group",

number = "8",

}

TY - JOUR

T1 - An enhanced CRISPR repressor for targeted mammalian gene regulation

AU - Yeo, Nan Cher

AU - Chavez, Alejandro

AU - Lance-Byrne, Alissa

AU - Chan, Yingleong

AU - Menn, David

AU - Milanova, Denitsa

AU - Kuo, Chih Chung

AU - Guo, Xiaoge

AU - Sharma, Sumana

AU - Tung, Angela

AU - Cecchi, Ryan J.

AU - Tuttle, Marcelle

AU - Pradhan, Swechchha

AU - Lim, Elaine T.

AU - Davidsohn, Noah

AU - Ebrahimkhani, Mo R.

AU - Collins, James J.

AU - Lewis, Nathan E.

AU - Kiani, Samira

AU - Church, George M.

N1 - Funding Information: This work was supported by the NIH (grants RM1 HG008525 and P50 HG005550 to G.M.C.), the National Cancer Institute (grant 5T32CA009216-34 to A.C.), the Burroughs Wellcome Fund (Career Award for Medical Scientists to A.C.), NIGMS (R35 GM119850 to N.E.L.), the Novo Nordisk Foundation (NNF10CC1016517 to N.E.L.), DARPA (Young Faculty Award D16AP00047 to S.K.), the Arizona State University Fulton Schools of Engineering startup fund (S.K.), and the Paul G. Allen Frontiers Group (J.J.C.). HEK293T cells were a gift from P. Mali (University of California, San Diego, San Diego, CA, USA). psPAX2 was a gift from D. Trono (Ecole Polytechnique Federale de Lausanne, Lausanne, Switzerland). pCMV-VSV-G was a gift from B. Weinberg (Massachusetts Institute of Technology, Boston, MA, USA). The plasmid containing a single gRNA library targeting essential genes was a gift from R. Bernards (The Netherlands Cancer Institute, Amsterdam, the Netherlands). Publisher Copyright: © 2018, The Author(s).

PY - 2018/8/1

Y1 - 2018/8/1

N2 - The RNA-guided endonuclease Cas9 can be converted into a programmable transcriptional repressor, but inefficiencies in target-gene silencing have limited its utility. Here we describe an improved Cas9 repressor based on the C-terminal fusion of a rationally designed bipartite repressor domain, KRAB–MeCP2, to nuclease-dead Cas9. We demonstrate the system’s superiority in silencing coding and noncoding genes, simultaneously repressing a series of target genes, improving the results of single and dual guide RNA library screens, and enabling new architectures of synthetic genetic circuits.

AB - The RNA-guided endonuclease Cas9 can be converted into a programmable transcriptional repressor, but inefficiencies in target-gene silencing have limited its utility. Here we describe an improved Cas9 repressor based on the C-terminal fusion of a rationally designed bipartite repressor domain, KRAB–MeCP2, to nuclease-dead Cas9. We demonstrate the system’s superiority in silencing coding and noncoding genes, simultaneously repressing a series of target genes, improving the results of single and dual guide RNA library screens, and enabling new architectures of synthetic genetic circuits.

UR - http://www.scopus.com/inward/record.url?scp=85049982902&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85049982902&partnerID=8YFLogxK

U2 - 10.1038/s41592-018-0048-5

DO - 10.1038/s41592-018-0048-5

M3 - Article

C2 - 30013045

AN - SCOPUS:85049982902

SN - 1548-7091

VL - 15

SP - 611

EP - 616

JO - Nature Methods

JF - Nature Methods

IS - 8

ER -

An enhanced CRISPR repressor for targeted mammalian gene regulation

Abstract

ASJC Scopus subject areas

Access to Document

Other files and links

Fingerprint

Cite this