Fig. 3.

A comprehensive overview of intrinsic and extrinsic factors regulating biomolecular phase separation. a Increasing the intracellular concentration of phase-separating proteins can shift the system beyond the saturation threshold, promoting phase separation through multivalent interactions. b Disease-associated mutations may alter the interaction strength, structure, or charge distribution of proteins, which can either enhance or impair their ability to undergo phase separation, sometimes leading to abnormal condensate behavior. c Fusion proteins generated by chromosomal translocations often link domains with high phase-separation potential. For example, the FUS–CHOP fusion combines the prion-like domain of FUS with a leucine zipper from CHOP, enabling aberrant condensate formation independent of physiological regulatory mechanisms. d PTMs such as phosphorylation, methylation, ubiquitination, acetylation, SUMOylation, and lactylation modulate the interaction properties of proteins by altering their charge state, hydrophobicity, or domain accessibility, thereby fine-tuning condensation propensity and condensate architecture. e Intracellular condensates are typically composed of multiple components, and the phase behavior of a given protein is profoundly influenced by its interaction partners due to the multicomponent nature of these assemblies. f Temperature affects molecular diffusion and interaction kinetics, thereby modulating biomolecular phase separation. g The pH of the surrounding environment affects the protonation state of charged residues, thereby modulating electrostatic complementarity and hydrophobic interactions. These changes can either promote or inhibit phase separation. h Salt modulates the ionic strength of the cellular milieu, thereby influencing electrostatic shielding between charged domains and ultimately regulating condensate formation. i ATP functions not only as a metabolic energy source but also as a regulator of biomolecular phase separation, owing to its amphiphilic nature. j Various stress conditions such as heat shock, oxidative stress, energy deprivation, and endomembrane damage can trigger the formation of SGs, integrating biomolecular condensation into the cellular stress response. k Metabolites including glutamine and glycogen influence condensate formation by affecting macromolecular crowding, hydration shells, and interaction specificity, thereby linking metabolic status to phase separation regulation. l Molecular chaperones can modulate the phase separation behavior of their client proteins. m Nucleic acids such as RNA and DNA contribute to condensate formation by serving as scaffolds or client molecules. Additionally, modifications such as m6A enhance the ability of RNA to recruit specific reader proteins like YTHDFs, thereby promoting the formation of YTHDF condensates