title: Comparison of TALE designer transcription factors and the CRISPR/dCas9 in regulation of gene expression by targeting enhancers.
abstract: The transcription activator-like effectors (TALEs) and the RNA-guided clustered regularly interspaced short palindromic repeat (CRISPR) associated protein (Cas9) utlilize distinct molecular mechanisms in targeting site recognition. The two proteins can be modified to carry additional functional domains to regulate expression of genomic loci in mammalian cells. In this study, we have compared the two systems in activation and suppression of the Oct4 and Nanog loci by targeting their enhancers. Although both are able to efficiently activate the luciferase reporters, the CRISPR/dCas9 system is much less potent in activating the endogenous loci and in the application of reprogramming somatic cells to iPS cells. Nevertheless, repression by CRISPR/dCas9 is comparable to or even better than TALE repressors. We demonstrated that dCas9 protein binding results in significant physical interference to binding of native transcription factors at enhancer, less efficient active histone markers induction or recruitment of activating complexes in gene activation. This study thus highlighted the merits and drawbacks of transcription regulation by each system. A combined approach of TALEs and CRISPR/dCas9 should provide an optimized solution to regulate genomic loci and to study genetic elements such as enhancers in biological processes including somatic cell reprogramming and guided differentiation.
authors.affiliation: Wellcome Trust Sanger Institute, Hinxton, Cambridge, CB10 1HH, UK.
authors.forename: Xuefei
authors.initials: X
authors.lastname: Gao
authors.affiliation: Wellcome Trust Sanger Institute, Hinxton, Cambridge, CB10 1HH, UK.
authors.forename: Jason C H
authors.initials: JC
authors.lastname: Tsang
authors.affiliation: Wellcome Trust Sanger Institute, Hinxton, Cambridge, CB10 1HH, UK.
authors.forename: Fortis
authors.initials: F
authors.lastname: Gaba
authors.affiliation:  Key laboratory of Regenerative Biology, GIBH, Chinese Academy of Sciences, Guangzhou 510530, China.
authors.forename: Donghai
authors.initials: D
authors.lastname: Wu
authors.affiliation: Wellcome Trust Sanger Institute, Hinxton, Cambridge, CB10 1HH, UK  Shanghai Institute of Immunology, Shanghai Jiaotong University School of Medicine Shanghai 200025. China pl2@sanger.ac.uk.
authors.forename: Liming
authors.initials: L
authors.lastname: Lu
authors.affiliation: Wellcome Trust Sanger Institute, Hinxton, Cambridge, CB10 1HH, UK  Wellcome Trust-Medical Research Council Cambridge Stem Cell Institute, Universityof Cambridge, Tennis Court Rd, Cambridge CB2 1QR, UK pl2@sanger.ac.uk.
authors.forename: Pentao
authors.initials: P
authors.lastname: Liu
mesh_terms: D000818:Animals; D001667:Binding, Competitive; D064130:CRISPR-Associated Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D065150:Cellular Reprogramming; D004742:Enhancer Elements, Genetic; D044127:Epigenesis, Genetic; D005786:Gene Expression Regulation; D015723:Gene Library; D005822:Genetic Vectors; D057026:Induced Pluripotent Stem Cells / Q000378:metabolism; D051379:Mice; D050814:Octamer Transcription Factor-3 / Q000235:genetics; D010957:Plasmids / Q000235:genetics; D012097:Repressor Proteins / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D015533:Transcriptional Activation
chemical_list: D064130:CRISPR-Associated Proteins; D050814:Octamer Transcription Factor-3; D012097:Repressor Proteins; D014157:Transcription Factors