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Ribonucleoprotein (RNP) granules are membraneless liquid condensates that dynamically form,dissolve, and mature into a gel-like state in response to a changing cellular environment. RNP condensation islargely governed by promiscuous attractive inter-chain interactions mediated by low-complexity domains(LCDs). Using an archetypal disordered RNP, fused in sarcoma (FUS), here we study how molecular crowdingimpacts the RNP liquid condensation. We observe that the liquid⁻liquid coexistence boundary of FUS islowered by polymer crowders, consistent with an excluded volume model. With increasing bulk crowderconcentration, the RNP partition increases and the diffusion rate decreases in the condensed phase.Furthermore, we show that RNP condensates undergo substantial hardening wherein protein-dense dropletstransition from viscous fluid to viscoelastic gel-like states in a crowder concentration-dependent manner.Utilizing two distinct LCDs that broadly represent commonly occurring sequence motifs driving RNP phasetransitions, we reveal that the impact of crowding is largely independent of LCD charge and sequence patterns.These results are consistent with a thermodynamic model of crowder-mediated depletion interaction, whichsuggests that inter-RNP attraction is enhanced by molecular crowding. The depletion force is likely to play akey role in tuning the physical properties of RNP condensates within the crowded cellular space.
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