The field of epigenetics tends to be very “fluid” with new studies and paradigm shifts seemingly the norm! Now two new studies in Nature have described a new liquid model for the formation and function of silent chromatin domains known as heterochromatin.
The predominant view of heterochromatin formation and function tells us that repressive epigenetic modifications recruit adaptor proteins that help to compact chromatin into large, distinct, condensed domains. These dense domains remain inaccessible to the transcriptional machinery, and hence heterochromatin contributes to the repression of unwanted/unneeded transcription. However, many of the observed characteristics of heterochromatin – such as the accessibility of bulky DNA repair proteins and certain dynamic features – do not fit with this compaction model.
In the search for a new model, two sets of researchers hit upon the same idea: heterochromatin formation driven by biological “phase-separation”. In this strange sounding, but actually common scenario, certain proteins come together to form distinct liquid-like droplets that selectively concentrate some molecules and exclude others. This separation creates non-membrane-bound intracellular compartments such as nucleoli or P cells. For a simpler picture of phase-separation, consider what happens minutes after mixing motor oil and water or dunking some sourdough into a dish of peppery olive oil and aged balsamic vinegar, whichever takes your fancy!
Both teams concentrated on Heterochromatin Protein 1 alpha (or HP1a), an adaptor protein that binds the methylated lysine 9 of histone H3 (H3K9me) and other HP1a monomers, as a potential driver of biological phase-separation in what would represent a new mode of chromatin-mediated transcriptional control.
The first jaw drop-ping study, from the lab of Gary H. Karpen (Lawrence Berkeley National Laboratory/University of California, USA), assessed HP1a dynamics in fruit fly (Drosophila) embryos and mammalian cells, and discovered that:
- Purified Drosophila HP1a protein spontaneously phase-separated in solution to form spherical liquid-like droplets
- Droplets reversibly dissolved at 37 °C, as observed for other phase-separation-associated proteins
- HP1a proteins also nucleated into liquid-like foci during heterochromatin domain formation in Drosophila embryos
- These foci displayed many signs of phase-separation behavior; they grew by fusion, repelled large, inert macromolecules, and dissolved after the disruption of weak hydrophobic interactions
- Furthermore, phase-separated heterochromatin foci dissolved after HP1a removal
- Further analysis of HP1a dynamics in both Drosophila and mammalian nuclei found additional evidence of heterochromatin protein containment within liquid-like, phase-separated compartments
First author Amy Strom shares, “We are excited about these findings because they explain a mystery that’s existed in the field for a decade. That is, if compaction controls access to silenced sequences, how are other large proteins still able to get in? Chromatin organization by phase separation means that proteins are targeted to one liquid or the other based not on size, but on other physical traits, like charge, flexibility, and interaction partners.”
- Human HP1α also phase-separated in solution to form liquid-like droplets
- Phase-separating behavior required HP1α phosphorylation or the presence of DNA
- Both situations drove conformational changes allowing HP1α monomers to bind to each other
- Mutation of a conserved region of HP1a leading to a loss of HP1-HP1 interaction and hence phase-separating behavior inhibited the formation of nucleating HP1α foci
- Heterochromatin-associated factors preferentially concentrated within phase-separated liquid-like droplets
- Although certain molecules, such as the TFIIB transcription factor, did not localize to these foci
- A high-tech single molecule DNA curtain assay confirmed that DNA-bound HP1α monomers bind to adjacent DNA-bound monomers to nucleate heterochromatin into phase-separated liquid-like droplets
Karpen concludes, “Gene therapy, or any treatment that relies on tight regulation of gene expression, could be improved by precisely targeting molecules to the right place in the nucleus. It is very difficult to target genes located in heterochromatin, but this understanding of the properties linked to phase separation and liquid behaviors could help change that and open up a third of the genome that we couldn’t get to before.”
So has the penny finally “dropped” and have these studies uncovered a better model for heterochromatin formation and function? Ongoing studies from the groups aim to fully appreciate the consequences of this new liquid phase-separation model to genome function by delineating just how specific molecules are concentrated into these liquid-like droplets, the exact physicochemical environment within droplets, and the role silencing RNAs.
Overall, we are sure that these studies represent a lot more than just another “drop in the ocean”; indeed, these two exciting studies may represent a paradigm shift in our understanding of genome organization, chromatin dynamics, and the regulation of gene expression! See these amazing new studies in Nature at Larson et al. and Strom et al.