Interplay between charge distribution and DNA in shaping HP1 paralog phase separation and localization

Abstract

The heterochromatin protein 1 (HP1) family is a crucial component of heterochromatin with diverse functions in gene regulation, cell cycle control, and cell differentiation. In humans, there are three paralogs, HP1α, HP1β, and HP1γ, which exhibit remarkable similarities in their domain architecture and sequence properties. Nevertheless, these paralogs display distinct behaviors in liquid-liquid phase separation (LLPS), a process linked to heterochromatin formation. Here, we employ a coarse-grained simulation framework to uncover the sequence features responsible for the observed differences in LLPS. We highlight the significance of the net charge and charge patterning along the sequence in governing paralog LLPS propensities. We also show that both highly conserved folded and less-conserved disordered domains contribute to the observed differences. Furthermore, we explore the potential co-localization of different HP1 paralogs in multicomponent assemblies and the impact of DNA on this process. Importantly, our study reveals that DNA can significantly reshape the stability of a minimal condensate formed by HP1 paralogs due to competitive interactions of HP1α with HP1β and HP1γ versus DNA. In conclusion, our work highlights the physicochemical nature of interactions that govern the distinct phase-separation behaviors of HP1 paralogs and provides a molecular framework for understanding their role in chromatin organization.

A complex network of interdomain interactions underlies the conformational ensemble of monomeric TDP‐43 and modulates its phase behavior

“…As stated in the previous section, the favorable interdomain contacts from our single‐chain, atomistic simulations may also contribute toward intermolecular interactions within the condensed phase of TDP‐43. As an example, the positive correlation between the intra‐monomer contacts and inter‐chain interactions implicated in phase separation was demonstrated recently in case of the HP1 paralogs where CG phase‐coexistence simulations were used to complement the AA simulations and study the protein phase behavior at longer timescales (Her et al, 2022 ; Phan et al, 2023 ). We used the HPS model (Dignon, Zheng, Kim, et al, 2018 ) based on the Urry hydrophobicity scale (Regy et al, 2021 ) to study the phase behavior of TDP‐43 at a single‐bead per amino acid resolution.…”

Uncovering the molecular interactions underlying MBD2 and MBD3 phase separation

“…However, upon the introduction of DNA, a notable reversal was observed: MBD2 predominantly localized within the condensate (insets of Figures 5A, 7B), likely facilitated by its enhanced affinity for both DNA and MBD3 (Figures 7C-E). These observations underscore the role of DNA in modulating the colocalization of MBD proteins, which carries significant implications for understanding their biological functions in the context of chromatin (39,72).…”

“…How do the structural elements within MBD2 and MBD3 contribute to their ability to phase separate? Like the interplay observed between the IDRs and folded domains within HP1α and MeCP2 that give rise to their LLPS behavior (5, 17, 39), we similarly aim to understand how the interactions between the folded and disordered regions within MBD2 and MBD3 contribute to their LLPS. We designed MBD2 and MBD3 truncations that include or exclude important sequence-based features or structural and functional regions to test their influence on LLPS propensity (Figures 3A and Figure 4A).…”

“…The number of chains of MBD2 and MBD3 (50 and 66, respectively) were chosen to give similar protein concentrations. The simulation box size and the number of protein chains were selected to account for the potential impact of finite-size effects as done in our previous work (39).…”

“…The coarse-grained (CG) coexistence simulations of MBD2 and MBD3 with and without DNA were performed in the HOOMD-Blue 2.9.7 software package using the slab geometry as described in previous studies (36)(37)(38)(39). The MBDs of MBD2 and MBD3 proteins were constructed using the available structural models of these domains (PDB ID: 6CNP and 2MB7) (22,40).…”