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High-Throughput and Proteome-wide Discovery of Endogenous Biomolecular Condensates

ABSTRACT: Phase separation inside mammalian cells regulates the formation of biomolecular condensates that are related to gene expression, signalling, development, and diseases. However, a large population of endogenous condensates and their candidate phase separating proteins have yet to be discovered in a quantitative and high-throughput manner. Here, we demonstrate that endogenously-expressed biomolecular condensates can be identified across a cell's proteome by sorting proteins across varying oligomeric states. We employ volumetric compression to modulate the concentrations of intracellular proteins and the degree of crowdedness, which are physical regulators of cellular biomolecular condensates. The changes in degree of the partition of proteins into condensates or phase separation lead to varying oligomeric states of the proteins, which can be detected by coupling density gradient ultracentrifugation and quantitative mass spectrometry. In total, we identified 1,518 endogenous-expression condensate proteins, of which 538 have not been reported before. Furthermore, we demonstrate that our strategy can identify condensate proteins that respond to specific biological processes.

INSTRUMENT(S): Orbitrap Exploris 480

ORGANISM(S): Homo Sapiens (human) Mus Musculus (mouse)

TISSUE(S): Epithelial Cell

SUBMITTER: Pengjie Li

LAB HEAD: Yiwei Li

PROVIDER: PXD048218 | Pride | 2024-03-18

REPOSITORIES: Pride

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