RNA-guided transcriptional regulation in planta via synthetic dCas9-based transcription factors
AuthorsPiatek, Agnieszka Anna
Abulfaraj, Aala A.
Mahfouz, Magdy M.
KAUST DepartmentBiological and Environmental Sciences and Engineering (BESE) Division
Center for Desert Agriculture
MetadataShow full item record
AbstractTargeted genomic regulation is a powerful approach to accelerate trait discovery and development in agricultural biotechnology. Bacteria and archaea use clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated (Cas) regulatory systems for adaptive molecular immunity against foreign nucleic acids introduced by invading phages and conjugative plasmids. The type II CRISPR/Cas system has been adapted for genome editing in many cell types and organisms. A recent study used the catalytically inactive Cas9 (dCas9) protein combined with guide-RNAs (gRNAs) as a DNA-targeting platform to modulate gene expression in bacterial, yeast, and human cells. Here, we modified this DNA-targeting platform for targeted transcriptional regulation in planta by developing chimeric dCas9-based transcriptional activators and repressors. To generate transcriptional activators, we fused the dCas9 C-terminus with the activation domains of EDLL and TAL effectors. To generate a transcriptional repressor, we fused the dCas9 C-terminus with the SRDX repression domain. Our data demonstrate that dCas9 fusion with the EDLL activation domain (dCas9:EDLL) and the TAL activation domain (dCas9:TAD), guided by gRNAs complementary to selected promoter elements, induce strong transcriptional activation on Bs3
SponsorsThis study is funded by King Abdullah University of Science and Technology (KAUST). The authors thank Lei S. Qi for providing the Cas9 and dCas9 plasmid constructs. The authors would like to thank Moussa Benhamed and Axel de Julien de Zelicourt for their helpful comments on qPCR analysis. Moreover, authors thank the anonymous reviewers for their valuable comments which helped improve the manuscript. No conflict of interest declared.
JournalPlant Biotechnology Journal
- Multiplexed Transcriptional Activation or Repression in Plants Using CRISPR-dCas9-Based Systems.
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- CRISPR RNA-guided activation of endogenous human genes.
- Authors: Maeder ML, Linder SJ, Cascio VM, Fu Y, Ho QH, Joung JK
- Issue date: 2013 Oct
- Genome engineering via TALENs and CRISPR/Cas9 systems: challenges and perspectives.
- Authors: Mahfouz MM, Piatek A, Stewart CN Jr
- Issue date: 2014 Oct
- RNA-guided transcriptional activation via CRISPR/dCas9 mimics overexpression phenotypes in Arabidopsis.
- Authors: Park JJ, Dempewolf E, Zhang W, Wang ZY
- Issue date: 2017
- Gene transcription repression in Clostridium beijerinckii using CRISPR-dCas9.
- Authors: Wang Y, Zhang ZT, Seo SO, Lynn P, Lu T, Jin YS, Blaschek HP
- Issue date: 2016 Dec