Drosophilainsulator proteins exhibit in vivo liquid–liquid phase separation properties

Abstract

Mounting evidence implicates liquid–liquid phase separation (LLPS), the condensation of biomolecules into liquid-like droplets in the formation and dissolution of membraneless intracellular organelles (MLOs). Cells use MLOs or condensates for various biological processes, including emergency signaling and spatiotemporal control over steady-state biochemical reactions and heterochromatin formation. Insulator proteins are architectural elements involved in establishing independent domains of transcriptional activity within eukaryotic genomes. InDrosophila, insulator proteins form nuclear foci known as insulator bodies in response to osmotic stress. However, the mechanism through which insulator proteins assemble into bodies is yet to be investigated. Here, we identify signatures of LLPS by insulator bodies, including high disorder tendency in insulator proteins, scaffold–client–dependent assembly, extensive fusion behavior, sphericity, and sensitivity to 1,6-hexanediol. We also show that the cohesin subunit Rad21 is a component of insulator bodies, adding to the known insulator protein constituents and γH2Av. Our data suggest a concerted role of cohesin and insulator proteins in insulator body formation and under physiological conditions. We propose a mechanism whereby these architectural proteins modulate 3D genome organization through LLPS.

A mechanically regulated liquid-liquid phase separation of the transcriptional regulator Tono instructs muscle development

“…The dynamics of this change follows the typical fast fusion dynamics of smaller droplets to large ones, as described for various liquid-liquid phase separations 63,65,76 . In particular in the nucleus, liquidliquid phase separation has been proposed as a mechanism to subdivide the nucleus into different activity domains, with active and inactive DNA compartments, further subdivided into topologically associating domains (TADs) 76,77 and the formation of socalled super-enhancers that result in high expression rates 78 . Established insulator proteins, like Cp190 and Mod(mdg4)67.2 also contain BTB Zn-finger domains 77 and condense into large droplets distinct from the Tono droplets under high osmotic pressure.…”

“…In particular in the nucleus, liquidliquid phase separation has been proposed as a mechanism to subdivide the nucleus into different activity domains, with active and inactive DNA compartments, further subdivided into topologically associating domains (TADs) 76,77 and the formation of socalled super-enhancers that result in high expression rates 78 . Established insulator proteins, like Cp190 and Mod(mdg4)67.2 also contain BTB Zn-finger domains 77 and condense into large droplets distinct from the Tono droplets under high osmotic pressure. What makes Tono special is that it is also mechano-sensitive and that it displays a muscle-specific loss of function phenotype.…”

1,6-Hexanediol Is Inducing Homologous Recombination by Releasing BLM from Assemblysomes in Drosophila melanogaster

“…This reduction in parallel mitigated the DNA damage response in these otherwise radiationresistant cells [1]. HEX has been shown to disturb liquid-liquid phase separation (LLPS) in the Drosophila singe cell system [14]. Furthermore, this chemical has been shown to increase general genome instability [15].…”

“…in the Drosophila singe cell system [14]. Furthermore, this chemical has been shown to increase general genome instability [15].…”