Reprogrammable CRISPR/Cas9-based system for inducing site-specific DNA methylation
Year: 2016
Type: article
Abstract: Advances in sequencing technology allow researchers to map genome-wide changes in DNA methylation in development and disease. However, there is a lack of experimental tools to site-specifically manipulate DNA methylation to discern the functional consequences. We developed a CRISPR/Cas9 DNA methyltransferase 3A (DNMT3A) fusion to induce DNA methylation at specific loci in the genome. We induced DNA methylation at up to 50% of alleles for targeted CpG dinucleotides. DNA methylation levels peaked within 50 bp of the short guide RNA (sgRNA) binding site and between pairs of sgRNAs. We used our approach to target methylation across the entire CpG island at the CDKN2A promoter, three CpG dinucleotides at the ARF promoter, and the CpG island within the Cdkn1a promoter to decrease expression of the target gene. These tools permit mechanistic studies of DNA methylation and its role in guiding molecular processes that determine cellular fate.
Source: Biology Open
DOI: 10.1242/bio.019067
Institution: Washington University in St. Louis
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $1,995
Funders: V Foundation for Cancer Research, American Society of Hematology, Alex's Lemonade Stand Foundation for Childhood Cancer, Children's Discovery Institute, Edward Mallinckrodt, Jr. Foundation, Alvin J. Siteman Cancer Center, National Institutes of Health, National Cancer Institute, National Institute of General Medical Sciences, National Institute of Diabetes and Digestive and Kidney Diseases, U.S. National Library of Medicine, Congressionally Directed Medical Research Programs
Awards: MODELING DNA METHYLATION'S ROLE IN GENE REGULATION, EPIGENTIC REGULATION OF HEMATOPOIETIC STEM CELL FUNCTION AND TRANSFORMATION, Training Program in Cellular and Molecular Biology, A MACHINE LEARNING APPROACH FOR FINE-SCALE GENOME WIDE DNA METHYLATION ANALYSIS, Mammary Cancer and Activation of Transposable Elements, Cancer Center Support Grant
OpenAlex ID: W2351565907