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Dissecting the mechanism of HP1 mediated chromatin compaction


Heterochromatin protein-1 (HP1) maintains condensed, stable heterochromatin domains throughout interphase despite the weak binding and rapid exchange of HP1 within heterochromatin. I utilize two methods, DNA curtains and liquid-liquid phase separation (LLPS) assays, to decode a mechanistic understanding of HP1 behavior and directly test if dynamic HP1 binding can maintain static DNA compaction in vitro. Within droplets, we find HP1α and DNA have distinct material properties: HP1α rapidly exchanges within and between droplets while simultaneously condensing DNA into stable domains within a droplet. Further, we show HP1α compacted DNA puncta are resistant to 40pN of force, over twice that required to stall RNA polymerase. I find the disordered regions of the three human HP1 paralogs - HP1α, HP1β, and HP1γ – dictate their DNA compaction and LLPS phenotypes. The HP1α hinge is necessary and sufficient for these activities, and we determine a network of hinge autoregulation within the Nand C- terminal extensions. I demonstrate dynamic HP1α binding primes droplets for regulation as the addition of HP1β, which exhibits minimal DNA compaction and LLPS behavior, invades and dissolves preformed HP1α droplets. Finally, I utilize chromatin substrates and find HP1 maintains separate domains of unmodified and H3K9me3 chromatin. Together this data describes how a pool of weakly bound HP1 proteins exploits both the collective behavior of proteins and the polymer properties of DNA to produce self-organizing domains that are simultaneously stable and fragile.

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