PT  - JOURNAL ARTICLE
AU  - Yuchen Gao
AU  - Mengting Han
AU  - Stephen Shang
AU  - Haifeng Wang
AU  - Lei S. Qi
TI  - Interrogation of the Dynamic Properties of Higher-Order Heterochromatin Using CRISPR/dCas9
AID  - 10.1101/2021.06.06.447300
DP  - 2021 Jan 01
TA  - bioRxiv
PG  - 2021.06.06.447300
4099  - http://biorxiv.org/content/early/2021/06/07/2021.06.06.447300.short
4100  - http://biorxiv.org/content/early/2021/06/07/2021.06.06.447300.full
AB  - Eukaryotic chromosomes feature large regions of compact, repressed heterochromatin hallmarked by Heterochromatin Protein 1 (HP1). HP1 proteins play multi-faceted roles in shaping heterochromatin, and in cells, HP1 tethering to individual gene promoters leads to epigenetic modifications and silencing. However, emergent properties of HP1 at supranucleosomal scales remain difficult to study in cells due to lack of appropriate tools. Here, we develop CRISPR-Engineered Chromatin Organization (EChO), combining live cell CRISPR imaging with inducible large-scale recruitment of chromatin proteins to native genomic targets. We demonstrate that human HP1α tiling across kilobase-scale genomic DNA forms novel contacts with natural heterochromatin, integrates two distantly targeted regions, and reversibly changes chromatin from a diffuse to compact state. The compact state exhibits delayed disassembly kinetics and represses transcription across over 600 kilobases. These findings support a polymer model of HP1α-mediated chromatin regulation and highlight the utility of CRISPR-EChO in studying supranucleosomal chromatin organization in living cells.Competing Interest StatementThe authors have filed a U.S. Provisional Application (No. 62/744,504) via Stanford University.