Determinants of Disordered Protein Co-Assembly Into Discrete Condensed Phases

Abstract

Cells harbor numerous mesoscale membraneless compartments that house specific biochemical processes and perform distinct cellular functions. These protein and RNA-rich bodies are thought to form through multivalent interactions among proteins and nucleic acids resulting in demixing via liquid-liquid phase separation (LLPS). Proteins harboring intrinsically disordered regions (IDRs) predominate in membraneless organelles. However, it is not known whether IDR sequence alone can dictate the formation of distinct condensed phases. We identified a pair of IDRs capable of forming spatially distinct condensates when expressed in cells. When reconstituted in vitro, these model proteins do not co-partition, suggesting condensation specificity is encoded directly in the polypeptide sequences. Through computational modeling and mutagenesis, we identified the amino acids and chain properties governing homotypic and heterotypic interactions that direct selective condensation. These results form the basis of physicochemical principles that may direct subcellular organization of IDRs into specific condensates and reveal an IDR code that can guide construction of orthogonal membraneless compartments.

Phase Separation in Mixtures of Prion-Like Low Complexity Domains is Driven by the Interplay of Homotypic and Heterotypic Interactions

“…8C ). Similar findings were recently reported by Welles et al, (71). Finally, adding asymmetry to the energetics by keeping A-B and B-B interactions equivalent, but weakening A-A interactions leads to a strong preference for A molecules to interact with B molecules over other A molecules, whereas B molecules show indifference at low temperatures, but a preference for other B molecules at high temperatures ( Fig.…”

“…Our results set the stage for understanding how dynamical control over compositions in complex mixtures might be achieved and how this can influence condensate formation of bodies such as 40S hnRNP particles (84) or stress granules. There is also growing interest in understanding how the interplay of homotypic and heterotypic interactions impact the dynamically controlled (85) or purely thermodynamically influenced miscibility of condensates formed in multicomponent systems (71,(86)(87)(88).…”

Phase Separation in Mixtures of Prion-Like Low Complexity Domains is Driven by the Interplay of Homotypic and Heterotypic Interactions

“…8C). Similar ndings were recently reported by Welles et al, 71 . Finally, adding asymmetry to the energetics by keeping A-B and B-B interactions equivalent, but weakening A-A interactions leads to a strong preference for A molecules to interact with B molecules over other A molecules, whereas B molecules show indifference at low temperatures, but a preference for other B molecules at high temperatures (Fig.…”

Heterotypic interactions in the dilute phase can drive co-condensation of prion-like low-complexity domains of FET proteins and mammalian SWI/SNF complex

“…S10) and the discussion above. If the homotypic chain interactions were dominant over heterotypic interactions in the ternary mixture, multi-phasic condensate morphologies may emerge 47,49,62,63 with spatially co-existing individual PLD phases, which is the opposite of our experimental observations (Fig. 5b-d & Fig.…”