Department of iPS Cell Research & Epigenetic Medicine, Keio University School of Medicine, Tokyo, Japan; Department of Physiology, Keio University School of Medicine, Tokyo, Japan
Osamu Takase
Department of iPS Cell Research & Epigenetic Medicine, Keio University School of Medicine, Tokyo, Japan; Department of Physiology, Keio University School of Medicine, Tokyo, Japan
Masahiro Yoshikawa
Department of iPS Cell Research & Epigenetic Medicine, Keio University School of Medicine, Tokyo, Japan; Department of Physiology, Keio University School of Medicine, Tokyo, Japan
Etsuko Sano
Department of iPS Cell Research & Epigenetic Medicine, Keio University School of Medicine, Tokyo, Japan; Department of Physiology, Keio University School of Medicine, Tokyo, Japan
Matsuhiko Hayashi
Apheresis and Dialysis Center, Keio University School of Medicine, Tokyo, Japan
Kazuto Hoshi
Division of Tissue Engineering, University of Tokyo Hospital, Tokyo, Japan; Department of Oral and Maxillofacial Surgery, University of Tokyo Hospital, Tokyo, Japan
Tsuyoshi Takato
Division of Tissue Engineering, University of Tokyo Hospital, Tokyo, Japan; Department of Oral and Maxillofacial Surgery, University of Tokyo Hospital, Tokyo, Japan
Department of Physiology, Keio University School of Medicine, Tokyo, Japan
Keiichi Hishikawa
Department of iPS Cell Research & Epigenetic Medicine, Keio University School of Medicine, Tokyo, Japan; Department of Physiology, Keio University School of Medicine, Tokyo, Japan
While regulation of gene-enhancer interaction is intensively studied, its application remains limited. Here, we reconstituted arrays of CTCF-binding sites and devised a synthetic topological insulator with tetO for chromatin-engineering (STITCH). By coupling STITCH with tetR linked to the KRAB domain to induce heterochromatin and disable the insulation, we developed a drug-inducible system to control gene activation by enhancers. In human induced pluripotent stem cells, STITCH inserted between MYC and the enhancer down-regulated MYC. Progressive mutagenesis of STITCH led to a preferential escalation of the gene-enhancer interaction, corroborating the strong insulation ability of STITCH. STITCH also altered epigenetic states around MYC. Time-course analysis by drug induction uncovered deposition and removal of H3K27me3 repressive marks follows and reflects, but does not precede and determine, the expression change. Finally, STITCH inserted near NEUROG2 impaired the gene activation in differentiating neural progenitor cells. Thus, STITCH should be broadly useful for functional genetic studies.
