Biophysical Principles Governing HP1a-DNA Condensate Properties: Regulatory Roles of Histone Tails and RNA

Author

Priyasha Deshpande, The Graduate Center, City University of New YorkFollow

Date of Degree

2-2025

Document Type

Dissertation

Degree Name

Ph.D.

Program

Biochemistry

Advisor

Shana Elbaum-Garfinkle

Advisor

Patrizia Casaccia

Committee Members

Kevin Ryan

Pinar Ayata

Alexandra Zidovska

Rein Ulijn

Subject Categories

Biochemistry, Biophysics, and Structural Biology

Keywords

Heterochromatin, Biomolecular Condensates, Phase separation

Abstract

Biomolecular condensates formed via phase separation of proteins and nucleic acids play crucial roles in chromatin organization and regulation. The biological functional outcomes of condensates are dictated by their emergent network properties that span the viscoelastic spectrum. Characterizing these material properties is essential for decoding the mechanisms underlying condensate function and dysfunction.

Eukaryotic genome is broadly compartmentalized into two functionally and morphologically distinct domains – euchromatin and heterochromatin. Heterochromatin is a critical structural component of the nucleus and is suggested to form via phase separation of Heterochromatin Protein 1α (HP1α) and underlying DNA. This thesis investigates how distinct epigenetic modifications and non-coding RNA fine-tune the material properties of HP1α-DNA condensates which are implicated in heterochromatin assembly.

Through a combination of biophysical and rheological approaches, we demonstrate that the heterochromatin-associated H3K9me3 mark specifically promotes and maintains HP1α-DNA phase separation while simultaneously modulating the condensates' viscoelastic properties. In contrast, unmodified H3 tails and activation-associated H3K4me3 marked tails are incompatible with phase separation due to their inability to bind HP1α.

We further demonstrate that telomeric repeat-containing RNA (TERRA) introduces an additional layer of complexity by forming distinct multiphase condensates in HP1α-DNA systems. These TERRA-enriched condensates exhibit solid-like properties and preferentially localize to the interfaces of larger HP1α-DNA condensates, thereby modulating their interfacial properties. This finding suggests a novel mechanism for how non-coding RNAs can influence heterochromatin organization through control of condensate interfaces.

Finally, we demonstrate that even in the absence of HP1α, histone H3 tails with different modifications display distinct phase behaviors with DNA. Specifically, H3K9me3 tails undergo phase separation at elevated DNA concentrations, while unmodified H3 and H3K4me3 tails form simple aggregates. These results highlight how specific epigenetic marks can directly influence chromatin phase behavior independent of reader proteins.

Together, this work provides mechanistic insights into how cells might utilize epigenetic modifications and non-coding RNAs to precisely control the material properties of chromatin-associated condensates, with important implications for understanding heterochromatin organization and regulation.

Recommended Citation

Deshpande, Priyasha, "Biophysical Principles Governing HP1a-DNA Condensate Properties: Regulatory Roles of Histone Tails and RNA" (2025). CUNY Academic Works.
https://academicworks.cuny.edu/gc_etds/6140