biostudies-literatureRamsak MJavna Agencija za Raziskovalno Dejavnost RSNIGMS NIH HHS7973https://www.ebi.ac.uk/biostudies/studies/S-EPMC10693550biostudies-literatureUnknown14(1)Membraneless liquid compartments based on phase-separating biopolymers have been observed in diverse cell types and attributed to weak multivalent interactions predominantly based on intrinsically disordered domains. The design of liquid-liquid phase separated (LLPS) condensates based on de novo designed tunable modules that interact in a well-understood, controllable manner could improve our understanding of this phenomenon and enable the introduction of new features. Here we report the construction of CC-LLPS in mammalian cells, based on designed coiled-coil (CC) dimer-forming modules, where the stability of CC pairs, their number, linkers, and sequential arrangement govern the transition between diffuse, liquid and immobile condensates and are corroborated by coarse-grained molecular simulations. Through modular design, we achieve multiple coexisting condensates, chemical regulation of LLPS, condensate fusion, formation from either one or two polypeptide components or LLPS regulation by a third polypeptide chain. These findings provide further insights into the principles underlying LLPS formation and a design platform for controlling biological processes.Nature communicationsProgrammable de novo designed coiled coil-mediated phase separation in mammalian cells.PMC10693550T32 GM065103P4-0176, P3-0289T32 GM145437Hough LEShirts MRRamsak MRamirez DAElersic Filipic KVidmar SAnderluh GJerala RfalseProgrammable de novo designed coiled coil-mediated phase separation in mammalian cells.Membraneless liquid compartments based on phase-separating biopolymers have been observed in diverse cell types and attributed to weak multivalent interactions predominantly based on intrinsically disordered domains. The design of liquid-liquid phase separated (LLPS) condensates based on de novo designed tunable modules that interact in a well-understood, controllable manner could improve our understanding of this phenomenon and enable the introduction of new features. Here we report the construction of CC-LLPS in mammalian cells, based on designed coiled-coil (CC) dimer-forming modules, where the stability of CC pairs, their number, linkers, and sequential arrangement govern the transition between diffuse, liquid and immobile condensates and are corroborated by coarse-grained molecular simulations. Through modular design, we achieve multiple coexisting condensates, chemical regulation of LLPS, condensate fusion, formation from either one or two polypeptide components or LLPS regulation by a third polypeptide chain. These findings provide further insights into the principles underlying LLPS formation and a design platform for controlling biological processes.2023-01-01T00:00:00Z2023 Dec2026-06-22T03:16:59.668Z2025-02-19T01:17:08.378ZS-EPMC106935503804289710.1038/s41467-023-43742-w