Format

Send to

Choose Destination
Cell. 2016 Sep 22;167(1):219-232.e14. doi: 10.1016/j.cell.2016.09.006.

Inheritable Silencing of Endogenous Genes by Hit-and-Run Targeted Epigenetic Editing.

Author information

1
San Raffaele Telethon Institute for Gene Therapy (SR-Tiget), IRCCS San Raffaele Scientific Institute, Via Olgettina 58, 20132 Milan, Italy; Vita-Salute San Raffaele University, Via Olgettina 58, 20132 Milan, Italy.
2
San Raffaele Telethon Institute for Gene Therapy (SR-Tiget), IRCCS San Raffaele Scientific Institute, Via Olgettina 58, 20132 Milan, Italy.
3
Center for Translational Genomics and Bioinformatics, IRCCS San Raffaele Scientific Institute, Via Olgettina 58, 20132 Milan, Italy.
4
San Raffaele Telethon Institute for Gene Therapy (SR-Tiget), IRCCS San Raffaele Scientific Institute, Via Olgettina 58, 20132 Milan, Italy; Vita-Salute San Raffaele University, Via Olgettina 58, 20132 Milan, Italy. Electronic address: naldini.luigi@hsr.it.
5
San Raffaele Telethon Institute for Gene Therapy (SR-Tiget), IRCCS San Raffaele Scientific Institute, Via Olgettina 58, 20132 Milan, Italy; Vita-Salute San Raffaele University, Via Olgettina 58, 20132 Milan, Italy. Electronic address: lombardo.angelo@hsr.it.

Abstract

Gene silencing is instrumental to interrogate gene function and holds promise for therapeutic applications. Here, we repurpose the endogenous retroviruses' silencing machinery of embryonic stem cells to stably silence three highly expressed genes in somatic cells by epigenetics. This was achieved by transiently expressing combinations of engineered transcriptional repressors that bind to and synergize at the target locus to instruct repressive histone marks and de novo DNA methylation, thus ensuring long-term memory of the repressive epigenetic state. Silencing was highly specific, as shown by genome-wide analyses, sharply confined to the targeted locus without spreading to nearby genes, resistant to activation induced by cytokine stimulation, and relieved only by targeted DNA demethylation. We demonstrate the portability of this technology by multiplex gene silencing, adopting different DNA binding platforms and interrogating thousands of genomic loci in different cell types, including primary T lymphocytes. Targeted epigenome editing might have broad application in research and medicine.

KEYWORDS:

B2M-null cells; CRISPR/Cas9; DNA methylation; DNMT3L; KRAB-ZFP/KAP1; TALE; TET1; epigenetic editing; gene therapy; permanent gene silencing

PMID:
27662090
PMCID:
PMC5039111
DOI:
10.1016/j.cell.2016.09.006
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Elsevier Science Icon for PubMed Central
Loading ...
Support Center