Research
Quantitative biophysics of nuclear organization
Life out of equilibrium
Cells organize functional internal compartments in the cytoplasm largely through membrane-bound organelles, which are selectively permeable, allowing the cell to expend energy to concentrate specific factors inside the organelle to achieve localized functions. In the last decade, researchers have come to understand that similar sub-cellular compartments can also exist without a bounding membrane. These micron scale membraneless structures can provide the cell with similar properties including selective permeability and functional compartmentalization, are formed through macromolecular self-assembly, and are increasingly referred to as biomolecular condensates.
Processes of macromolecular self-assembly have been extensively described in equilibrium inorganic systems, but how the living cell deploys the principles of self-assembly to organize its internal structures, and what happens when these equilibrium processes are dysregulated are yet to be completely understood.
A section of this poster is recreated in the 7th edition of the textbook Molecular Biology of the Cell by Alberts et al (Ch6p354)
Mesoscale organization of the nucleus
Within the mammalian cell nucleus, subcompartments including the nucleolus, nuclear speckle and heterochromatin (among other structures) concentrate unique subsets of molecules and perform independent functions. Many of these compartments need to form at specific regions of the genome to complete their function, and mistargeting or aberrant formation of nuclear structures are associated with disease states including aging and cancer. Still, how condensates interact with the genome to achieve targeted functional compartmentalization is incompletely understood.
Contributions of Dr. Amy Strom to nuclear organization and function
In her research, Dr. Strom has worked to define the basic molecular interactions that build emergent functional nuclear compartments at targeted loci. She has collaborated with theorists to adapt physical descriptions of inorganic systems to describe biological systems, and engineered optogenetic techniques that utilize these biophysical principles to address biological questions in new ways. She has applied this novel perspective to better understand how mutations in a nuclear compartment-forming protein leads to nuclear dysfunction and ultimately cancer.
The Liquid Nucleome (left, JCS 2019) is a model perspective proposed by Dr. Strom and colleagues that suggests basic biophysical principles including condensation can describe the fundamental organization of multiple functional nuclear structures.
Precision Genome Organization
An under-appreciated feature of biomolecular condensates is that they create interfaces with other cellular objects, and through interfacial or capillary forces, can do work. Dr. Strom and co-first-author Yoonji Kim rationally designed a biologically encoded system that creates interfacial interactions between light-inducible protein condensates and specific chromatin loci, and was able to exert force on the loci to reposition them across microns of space on demand in living cells. This system, called VECTOR, was utilized to induce collision of telomeres and establish whether they coalesce in a publication in Dev Cell, 2022, and was developed further to measure mechanical heterogeneity of the local chromatin network in hetero- and euchromatic areas, in a publication in Cell, August 2024. These results highlight the power of rational engineering in tool development to address complex problems, and establish a role for condensates in doing work to organize cells, all without ATP-driven motors.
Capillary forces between light-induced condensates and chromatin loci are capable of applying forces to rearrange the genome. This technology can potentially be further adapted to reposition any locus of interest, for applications in studies of enhancer-promoter interactions, DNA damage repair templates, and beyond.
Specificity from disorder
Disordered regions are associated with biomolecular condensation and are particularly enriched in nuclear proteins, so better understanding of the molecular grammar of disordered regions may allow new insights into mesoscale nuclear compartmentalization, and potentially be able to redirect efforts to develop small molecule therapeutics toward their most probable targets.
A disordered region controls cBAF activity via condensation and partner recruitment
25% of the human proteins contain at least 30% disorder. This lack of structure has been equated to a lack of specificity in protein-protein interactions and has led disordered regions to be considered poor targets for therapeutics, but there is growing evidence that disordered regions do have a 'molecular grammar' that confers specificity.
We demonstrated that condensate formation and heterotypic interactions are distinct and separable features of an IDR within the ARID1A/B subunits of the mSWI/SNF chromatin remodeler, cBAF, and establish distinct "sequence grammars" underlying each contribution.
Human disease-associated perturbations in ARID1B IDR sequence grammars disrupt cBAF function in cells. Together, these data identify IDR contributions to chromatin remodeling and explain how phase separation provides a mechanism through which both genomic loclaization and functional partner recruitment are achieved.
This work was a close collaboration with co-first author Ajinkya Patil, in the lab of Dr. Strom's K99 co-mentor Cigall Kadoch at the Dana Farber Cancer Institute. It was published in Cell in October 2023.
How do nuclear compartments interact with chromatin?
Phase separation drives heterochromatin domain formation
Constitutive heterochromatin is an important component of eukaryotic genomes that has essential roles in nuclear architecture, DNA repair and genome stability, and silencing of transposon and gene expression. Heterochromatic domains form via phase separation, and mature into a structure that includes liquid and stable compartments. Emergent biophysical properties associated with phase-separated systems are critical to understanding the unusual behaviors of heterochromatin, and how chromatin domains in general regulate essential nuclear functions.
Phase separation of heterochromatin was the focus of Dr. Strom's graduate work, published in Nature in 2017, and has represented a perspective shift in the field of nuclear organization to understand that condensation organizes nuclear compartments.
Interplay of condensation and chromatin binding underlies BRD4 targeting
While protein self-interactions, especially within low-complexity and intrinsically disordered regions, are known to mediate condensation, the role of substrate-binding interactions in regulating the formation and function of biomolecular condensates is under-explored. Both kinetic and thermodynamic properties of BRD4 condensation are affected by chromatin binding: nucleation rate is sensitive to BRD4-chromatin interactions, providing an explanation for the selective formation of BRD4 condensates at acetylated chromatin regions, and thermodynamically, multivalent acetylated chromatin sites provide a platform for BRD4 clustering below the concentration required for off-chromatin condensation.
This recent work from Dr. Strom and co-first-author Dr. Jorine Eeftens, alongside colleagues in the lab of Dr. Will Jacobs at Princeton was recently published in MBoC in May 2024.
Nuclear Mechanics
Both chromatin and the nuclear lamina provide mechanical structure to the nucleus, which is implicated in permature aging and constrained migration, especially during metastasis. It is known that increased histone methylation leads to increased nuclear rigidity, but is unclear whether this is a direct effect of the modification on chromatin structure, or indirect, through binding of heterochromatic protein HP1a. Alongside a team of 4D Nucleome consortium members, I created a U2OS cell line with auxin-inducible degradation of the endogenous HP1a protein. Upon HP1a degradation, we found nuclei were significantly softer and more deformable by micropipette manipulation. This work emphasizes that chromatin-binding proteins contribute to rigidity separately from lamins. It was published in eLife in 2021.