HP1 proteins compact DNA into mechanically and positionally stable phase separated domains

Abstract

In mammals, HP1-mediated heterochromatin forms positionally and mechanically stable genomic domains even though the component HP1 paralogs, HP1α, HP1β, and HP1γ, display rapid on-off dynamics. Here, we investigate whether phase-separation by HP1 proteins can explain these biological observations. Using bulk and single-molecule methods, we show that, within phase-separated HP1α-DNA condensates, HP1α acts as a dynamic liquid, while compacted DNA molecules are constrained in local territories. These condensates are resistant to large forces yet can be readily dissolved by HP1β. Finally, we find that differences in each HP1 paralog’s DNA compaction and phase-separation properties arise from their respective disordered regions. Our findings suggest a generalizable model for genome organization in which a pool of weakly bound proteins collectively capitalize on the polymer properties of DNA to produce self-organizing domains that are simultaneously resistant to large forces at the mesoscale and susceptible to competition at the molecular scale.

HP1α is a chromatin crosslinker that controls nuclear and mitotic chromosome mechanics

“…Similarly, HP1α has been shown to provide mechanical resistance to a single DNA fiber (Keenen et al, 2021). Furthermore, consistent with HP1α's newfound role in chromatin-based mechanics, we find that HP1α degradation results in the loss of nuclear shape stability (Figure 2 E, G), similar to the effects of other chromatin perturbations that soften the cell nucleus (Furusawa et al, 2015;Stephens et al, 2019aStephens et al, , 2019bStephens et al, , 2018Wang et al, 2018).…”

“…These results are consistent with prior experiments showing that chromatin dominates the mechanical response to small deformations, while lamins underlie strain stiffening to large deformations ( Stephens et al, 2017 ). Similarly, HP1α has been shown to provide mechanical resistance for a single DNA fiber ( Keenen et al, 2021 ). Furthermore, consistent with HP1α’s newfound role in chromatin-based mechanics, we find that HP1α degradation results in the loss of nuclear shape stability ( Figure 2E,G ), similar to the effects of other chromatin perturbations that soften the cell nucleus ( Furusawa et al, 2015 ; Stephens et al, 2019a ; Stephens et al, 2019b ; Stephens et al, 2018 ; Wang et al, 2018 ).…”

“…Furthermore, HP1α binding to methylated histones is known to alter the structure of the nucleosome core, which could promote nucleosome-nucleosome interactions, and induce polymer-polymer phase separation of the chromatin fiber ( Sanulli et al, 2019 ). Additionally, purified HP1α protein in vitro exhibits liquid-liquid phase separation by itself, with naked DNA, and with nucleosome arrays ( Larson et al, 2017 ; Shakya et al, 2020 ), and HP1 condensates bound to dsDNA in vitro can lend mechanical strength ( Keenen et al, 2021 ). The material properties of these in vitro condensates varies depending on the chromatin content ( Larson et al, 2017 ; Shakya et al, 2020 ).…”

Co-condensation of proteins with single- and double-stranded DNA

“…We speculate that the mechanism we describe here is relevant for other processes of DNA compaction, such as heterochromatin formation driven by HP1α (Larson et al ., 2017; Strom et al ., 2017; Larson and Narlikar, 2018; Sanulli et al ., 2019; Keenen et al ., 2021). We further suggest that generalized capillary forces arising in liquid-like co-condensates play an important role in other biological processes such as the transcription-dependent organization of chromatin (Cho et al ., 2018; Sabari et al ., 2018; Thompson et al ., 2018; Henninger et al ., 2021) (Figure S6A), the formation of viral replication compartments (Schmid et al ., 2014; Heinrich et al ., 2018; McSwiggen et al ., 2019; Nevers et al ., 2020) (Figure S6C), and the DNA damage response (Altmeyer et al ., 2015; Aleksandrov et al ., 2018; Naumann et al ., 2018; Levone et al ., 2021) (Figure S6B).…”

“…Many condensed structures play essential roles in nuclear organization. For example, heterochromatin is a dense form of chromatin in which DNA co-condenses with specific factors as well as nucleosomes to form transcriptionally silent domains of chromatin (Larson et al ., 2017; Strom et al ., 2017; Larson and Narlikar, 2018; Sanulli et al ., 2019; Keenen et al ., 2021). Furthermore, transcriptional condensates are dense and dynamic assemblies of transcription factors, associated proteins, DNA and RNA.…”