Investigators at Scripps Research have discovered that droplet-like compartments that form without membranes called biomolecular condensates can contain an internal filament architecture that is required for their function, a discovery that is counter to current views that they are simple liquids. The findings, published in Nature Structural & Molecular Biology, suggest that these defined structural features could serve as drug targets in diseases such as cancer and neurodegenerative disorders, where disruption of condensate formation and material properties contribute to disease pathology.
“Ever since we realized that disruptions in condensate formation are at the heart of many diseases, it has been challenging to target them therapeutically because they appeared to lack structure—there were no specific features for a drug to latch onto,” says senior author Keren Lasker, PhD, an associate professor at Scripps Research. “This work changes that. We can now see that some condensates have an internal architecture, and that, importantly, this structure is required for function, opening the door to targeting these membrane-less assemblies much like we target individual proteins.”
As described in the study, biomolecular condensates are nonstoichiometric assemblies of macromolecules that concentrate specific components while excluding others. They are driven by multivalent interactions and serve as a mechanism of cellular organization, regulating aspects of transcription, signal transduction, and protein quality control. Because they can merge, flow and exchange contents, it has been assumed they lack an organized internal structure.
“A particularly understudied question is how condensate function relates to its internal structure,” the researchers wrote. “By internal structure, we refer to stable structures on the molecular and supramolecular level, larger than single molecules but smaller than the condensate itself.”
To better understand the composition of biomolecular condensates, the Scripps researchers focused on the polar organizing protein Z (PopZ), which was first identified in the bacterium Caulobacter crescentus. In these rod-shaped bacteria, PopZ forms condensates at the cell poles, where they recruit proteins needed for asymmetric cell division. Deletion of PopZ leads to abnormal cell division, disrupted chromosome segregation and loss of cellular asymmetry. Prior work had shown that aberrations in condensate material properties, if they were either too fluid or too solid, could lead to growth defects, indicating that their physical properties are linked to biological function.
In order to map PopZ structure, the investigators used cryo-electron tomography to visualize condensates at high resolution. From this they learned that PopZ assembles hierarchically into trimers, hexamers and short filaments to form a scaffold within the droplet. Additional experiments showed that PopZ adopts different conformations depending on whether it is in a dilute phase or within a dense condensate. Phase-dependent conformational changes prevent interfilament contacts in the dilute phase and expose client-binding sites in the dense phase. “Realizing that protein conformation depends on location gives us multiple ways to engineer cellular function,” said first Daniel Scholl, PhD, a former postdoctoral researcher at Scripps.
While the research relied on a bacterial system, the researcher said the implications of their findings extend to human biology. Similar filament-based condensates in human cells help remove damaged or toxic proteins and regulate cell growth. These condensates fail to clear proteins, toxic proteins can accumulate, which is a hallmark of many neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS). Additionally, when growth-regulating condensates do not function properly, tumor suppression can break down, which can lead to the growth to prostate cancer, breast cancer, and endometrial malignancies.
The authors contend that their study provides a framework for drug development. “By demonstrating that condensate architecture is both definable and functionally critical, the work raises the possibility of designing therapies that act directly on condensate structure and correct the underlying disorganization that allows disease to take hold,” said Lasker.
Targeting specific domains that regulate filament formation, modulate electrostatic repulsion or control phase-dependent conformations could offer strategies to restore proper condensate behavior. For cancer, drugs might reinforce growth-suppressive condensates or prevent formation of assemblies that support tumor progression. For neurodegenerative disorders, therapies might stabilize condensates involved in protein quality control to enhance clearance of toxic aggregates.
Next steps for the team will be to broaden their understanding of this mechanistic analysis to other filament-driven condensates in human cells and testing whether modulating their internal architecture can correct disease-associated defects.
