Open Access
Editing DNA Methylation in the Mammalian Genome
X. Shawn Liu,Hao Wu,Xiong Ji,Yonatan Stelzer,Xuebing Wu,Szymon Czauderna,Szymon Czauderna,Jian Shu,Daniel B. Dadon,Richard A. Young,Rudolf Jaenisch +10 more
TLDR
Fusion of Tet1 or Dnmt3a with a catalytically inactive Cas9 enables targeted DNA methylation editing and these tools can editDNA methylation in mice, demonstrating their wide utility for functional studies of epigenetic regulation.Abstract:
Mammalian DNA methylation is a critical epigenetic mechanism orchestrating gene expression networks in many biological processes. However, investigation of the functions of specific methylation events remains challenging. Here, we demonstrate that fusion of Tet1 or Dnmt3a with a catalytically inactive Cas9 (dCas9) enables targeted DNA methylation editing. Targeting of the dCas9-Tet1 or -Dnmt3a fusion protein to methylated or unmethylated promoter sequences caused activation or silencing, respectively, of an endogenous reporter. Targeted demethylation of the BDNF promoter IV or the MyoD distal enhancer by dCas9-Tet1 induced BDNF expression in post-mitotic neurons or activated MyoD facilitating reprogramming of fibroblasts into myoblasts, respectively. Targeted de novo methylation of a CTCF loop anchor site by dCas9-Dnmt3a blocked CTCF binding and interfered with DNA looping, causing altered gene expression in the neighboring loop. Finally, we show that these tools can edit DNA methylation in mice, demonstrating their wide utility for functional studies of epigenetic regulation.read more
Citations
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Journal ArticleDOI
The CRISPR tool kit for genome editing and beyond.
TL;DR: A brief history of gene-editing tools is presented and the wide range of CRISPR-based genome-targeting tools are described, to conclude with future directions and the broader impact ofCRISPR technologies.
Journal ArticleDOI
The diverse roles of DNA methylation in mammalian development and disease
TL;DR: The mechanisms and functions of DNA methylation and demethylation in both mice and humans at CpG-rich promoters, gene bodies and transposable elements are discussed and the dynamic erasure and re-establishment in embryonic, germline and somatic cell development is highlighted.
Journal ArticleDOI
Integrative single-cell analysis
Tim Stuart,Rahul Satija +1 more
TL;DR: Diverse approaches for integrative single-cell analysis are discussed, including experimental methods for profiling multiple omics types from the same cells, analytical approaches for extracting additional layers of information directly from scRNA-seq data and computational integration of omics data collected across different cell samples.
Journal ArticleDOI
CRISPR-Based Technologies for the Manipulation of Eukaryotic Genomes.
Alexis C. Komor,Alexis C. Komor,Alexis C. Komor,Ahmed H. Badran,Ahmed H. Badran,Ahmed H. Badran,David R. Liu,David R. Liu,David R. Liu +8 more
TL;DR: Recent developments that extend the generality, DNA specificity, product selectivity, and fundamental capabilities of natural CRISPR systems are described.
Journal ArticleDOI
The DNA methyltransferase family: a versatile toolkit for epigenetic regulation
TL;DR: Analysis of molecular interactions and changes in gene copy numbers modulate the activity of DNMTs in diverse gene regulatory functions, including transcriptional silencing, transcriptional activation and post-transcriptional regulation by DNMT2-dependent tRNA methylation enables the DNMT family to function as a versatile toolkit for epigenetic regulation.
References
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STAR: ultrafast universal RNA-seq aligner
Alexander Dobin,Carrie A. Davis,Felix Schlesinger,Jorg Drenkow,Chris Zaleski,Sonali Jha,Philippe Batut,Mark Chaisson,Thomas R. Gingeras +8 more
TL;DR: The Spliced Transcripts Alignment to a Reference (STAR) software based on a previously undescribed RNA-seq alignment algorithm that uses sequential maximum mappable seed search in uncompressed suffix arrays followed by seed clustering and stitching procedure outperforms other aligners by a factor of >50 in mapping speed.
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Model-based Analysis of ChIP-Seq (MACS)
Yong Zhang,Tao Liu,Clifford A. Meyer,Jérôme Eeckhoute,David S. Johnson,Bradley E. Bernstein,Bradley E. Bernstein,Chad Nusbaum,Richard M. Myers,Myles Brown,Wei Li,X. Shirley Liu +11 more
TL;DR: This work presents Model-based Analysis of ChIP-Seq data, MACS, which analyzes data generated by short read sequencers such as Solexa's Genome Analyzer, and uses a dynamic Poisson distribution to effectively capture local biases in the genome, allowing for more robust predictions.
Journal ArticleDOI
A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity.
Martin Jinek,Krzysztof Chylinski,Krzysztof Chylinski,Ines Fonfara,Michael H. Hauer,Jennifer A. Doudna,Emmanuelle Charpentier +6 more
TL;DR: This study reveals a family of endonucleases that use dual-RNAs for site-specific DNA cleavage and highlights the potential to exploit the system for RNA-programmable genome editing.
Multiplex Genome Engineering Using CRISPR/Cas Systems
Le Cong,F. A. Ran,David Benjamin Turitz Cox,Shuailiang Lin,Robert P. J. Barretto,Naomi Habib,Patrick D. Hsu,Xuebing Wu,Wenyan Jiang,Luciano A. Marraffini,Feng Zhang +10 more
TL;DR: Two different type II CRISPR/Cas systems are engineered and it is demonstrated that Cas9 nucleases can be directed by short RNAs to induce precise cleavage at endogenous genomic loci in human and mouse cells, demonstrating easy programmability and wide applicability of the RNA-guided nuclease technology.
Journal ArticleDOI
RNA-Guided Human Genome Engineering via Cas9
Prashant Mali,Luhan Yang,Kevin M. Esvelt,John Aach,Marc Güell,James E. DiCarlo,Julie E. Norville,George M. Church,George M. Church +8 more
TL;DR: The type II bacterial CRISPR system is engineer to function with custom guide RNA (gRNA) in human cells to establish an RNA-guided editing tool for facile, robust, and multiplexable human genome engineering.