Mechanisms of active regulation of biomolecular condensates

Abstract

Liquid-liquid phase separation is a key organizational principle in eukaryotic cells, on par with intracellular membranes. It allows cells to concentrate specific proteins into condensates, increasing reaction rates and achieving switch-like regulation. However, it is unclear how cells trigger condensate formation or dissolution and regulate their sizes. We predict from first principles two mechanisms of active regulation by post-translational modifications such as phosphorylation: In enrichment-inhibition, the regulating modifying enzyme enriches in condensates and the modifications of proteins inhibit their interactions. Stress granules, Cajal bodies, P granules, splicing speckles, and synapsin condensates obey this model. In localization-induction, condensates form around an immobilized modifying enzyme, whose modifications strengthen protein interactions. Spatially targeted condensates formed during transmembrane signaling, microtubule assembly, and actin polymerization conform to this model. The two models make testable predictions that can guide studies into the many emerging roles of biomolecular condensates. Eukaryotic cells contain numerous types of mem-1 braneless organelles, which contain between a few 2 and thousands of protein and RNA species that are 3 highly enriched in comparison to the surrounding nu-4 cleoplasm or cytoplasm. These biomolecular conden-5 sates are held together by weak, multivalent and highly 6 collaborative interactions, often between intrinsically 7 54 active promoters. Here, we propose two active mecha-55 nisms used by cells for these purposes.56 Phase separation and condensate size behaviour 57 To keep the model simple, we consider only one type of 58 condensate protein. In the dilute regime below the sat-59 1 Cellular control of liquid droplet formation, size, and localization • July 5, 2019 Figure 1: Phase separation and condensate droplet size behaviour. A When protein-protein and solvent-solventinteractions are more favorable than protein-solvent interactions, demixing into two phases can occur, a dilute phase with low protein concentration c out and a dense phase with high concentration c in . This happens when the sum of free energies of the two phases is lower (tip of blue arrow) than the energy of the single phase (base of blue arrow) . B c out is the limiting concentration for infinite condensate droplet radius R. The concentration on the outside of a condensate of radius R is larger the smaller the condensate is (green double-headed arrows), as it cannot hold on to its proteins as well as large ones. This leads to a concentration gradient (∇concentration), which fuels a diffusive flux from small to large condensates (wiggly arrows). (l c is a measure of interaction strength between proteins in comparison to the solvent.) C As a result, condensates below a radius R crit will shrink and larger ones will grow. uration protein concentration c out , condensate droplets 60 cannot form ( Figure 1A). Above c out , in the phase sepa-61 ration regime, condensates can be stable...

TopBP1 assembles nuclear condensates to switch on ATR signalling

“…The assembly of complex heterogeneous biomolecules via multivalent, weak, dynamic and cooperative interactions is emerging as a common principle of compartmentation in the nucleus (Banani et al, 2017;Soding et al, 2020). We expect the rich diversity of molecular associations in live cells to yield diverse types of condensates with distinct composition and biophysical properties.…”

“…TopBP1 is among the proteins that are most highly phosphorylated and sumoylated in response to DNA replication stress (Munk et al, 2017). Post-translational modifications play a key role in the regulation of the molecular forces that drive the formation of biomolecular condensates (Soding et al, 2020). Since TopBP1 extensive capacity to self-assemble enables optogenetic induction of TopBP1 nuclear condensates, we used this optogenetic system to explore the role of phosphorylation in the formation of TopBP1 condensates.…”

“…In recent years, application of the principles of polymer chemistry to biological molecules has accelerated spectacularly our understanding of how membraneless compartments assemble and regulate their compositions and functions (Banani et al, 2017;Bracha et al, 2019;Hyman and Simons, 2012;Soding et al, 2020). These selforganized micron-scale structures, called biomolecular condensates, assemble via multiple weak, cooperative and dynamic interactions and compartmentalize proteins and nucleic acids.…”

14-3-3β paralog is inhibited by acetylation during differentiation to the osteogenic lineage.

“…This nuclear staining for 14-3-3acK49/51 is present at all days and conditions (NT and ODM), however in the later we also observed nuclear speckles, which resemble liquidlike condensates [29]. The cytoplasmic staining was more evident at day 14 and remains until the end of the treatment on day 21.…”

14-3-3β paralog is inhibited by acetylation during differentiation to the osteogenic lineage

“…Images were quantified by using a python scripts to show the enrichment in nuclear localization for 14-3-3acK49/51 for many conditions ( Fig 3B). This nuclear staining for 14-3-3acK49/51 is present at all days and conditions (NT and ODM), however in the later we also observed nuclear speckles, which resemble liquid-like condensates [29]. The cytoplasmic staining was more evident at day 14 and remains until the end of the treatment on day 21.…”