Compositional Control of Phase-Separated Cellular Bodies

Abstract

SUMMARY Cellular bodies such as P bodies and PML nuclear bodies (PML NBs) appear to be phase-separated liquids organized by multivalent interactions among proteins and RNA molecules. Although many components of various cellular bodies are known, general principles that define body composition are lacking. We modeled cellular bodies using several engineered multivalent proteins and RNA. In vitro and in cells, these scaffold molecules form phase-separated liquids that concentrate low valency client proteins. Clients partition differently depending on the ratio of scaffolds, with a sharp switch across the phase diagram diagonal. Composition can switch rapidly through changes in scaffold concentration or valency. Natural PML NBs and P bodies show analogous partitioning behavior, suggesting how their compositions could be controlled by levels of PML SUMOylation or cellular mRNA concentration, respectively. The data suggest a conceptual framework for considering the composition and control thereof of cellular bodies assembled through heterotypic multivalent interactions.

Coordinating cell cycle-regulated histone gene expression through assembly and function of the Histone Locus Body

“…Multivalent interactions driven by IDRs can reduce solubility and facilitate body formation, with the strong interactions governing polymerization and the weak interactions governing solubility. 86 Interestingly, conversion over time of phase separated droplets composed of IDRs and RNAs from a liquid like to a more fiber like state has been demonstrated in vitro suggesting that IDRs may contribute to the "maturing" of the HLB in response to transcriptional activation as described above. 90,91 Post-translational modifications may also modulate phase separation associated with gene expression.…”

“…68,[85][86][87] In this process the physical chemical properties of body components promote a phase transition when the concentration of the component(s) exceeds their solubility limit, such as when molecules precipitate out of solution and form a crystal. The phase transition need not result in a solid structure: Many bodies display liquid like properties, similar to a lipid droplet within an aqueous environment.…”

Specific genomic cues regulate Cajal body assembly

“…Several groups have shown that multivalent proteins enriched with highly disordered domains are capable of forming liquid droplets in vitro that display many characteristic features of NBs, including fusion and fission events. 86,87,90,91,145 It has also been suggested that separation of nucleoli, paraspeckles 146 and Nuage bodies formed by DEAD-box helicase 4 (Ddx4) protein 86,147 is driven by these disordered domains-containing proteins. Coilin and its homologs are highly disordered but Arabidopsis thaliana-derived coilin is stabilized upon snRNA binding leading to multimerization in vitro, 114 which can be assumed to contribute to CB nucleation.…”

“…8,[103][104][105] It has been established that the assembly and structural maintenance of NBs is predominantly influenced by several prominent architectural proteins that are able to self-associate to form homo-and hetero-oligomers. 80,87 Many CB proteins have been identified as structurally essential for CB integrity, but little is known about the biochemical properties and regulation of each factor. The majority of our knowledge regarding the proteins essential for CB assembly regards coilin and SMN, which are the classical CB marker proteins, as well as a limited number of other proteins.…”

“…Modeling of NB nucleation and growth in vivo under different experimental conditions has confirmed that NBs are indeed phase separated droplet organelles. 86,87 As a result, NB domains are compositionally distinct, but not impermeable, 88 to the surrounding interchromatin space leading to optimized regulation of their specific molecular functions. This sub-nuclear phase transition may be dependent upon coilin's ability to act as a molecular hub and interact with numerous CB components via hetero-oligomerization, which stabilizes the developing structure.…”