Biological and Medicinal Chemistry

Phosphorylation of the NBDY Microprotein Promotes Dissociation of Biomolecular Condensates

Abstract

This study reports detection of specific phosphorylation sites installed on a small open reading frame-encoded polypeptide or microprotein called NBDY. NBDY phosphorylation sites were mapped using phosphoproteomics and antibody-based validation. NBDY is phosphorylated during growth factor signaling and cell division, and quantitative fluorescence microscopy was used to show that NBDY phosphorylation is required for disappearance of cytoplasmic RNA-protein granules called P-bodies during these cellular processes. Because P-bodies have properties of liquid-liquid phase separated membraneless orgaenelles, reductionist system to investigate NBDY phase separation. Purified NBDY was shown to form complex coacervates in the presence of RNA via fluorescence microscopy and turbidity measurements, and phosphorylation by a kinase in vitro promotes liquid phase remixing.

Content

Thumbnail image of NBDY MS For Submission_ZN.pdf

Supplementary material

Thumbnail image of NBDY Phosphorylation SI.pdf
NBDY Phosphorylation SI
Thumbnail image of Movie S1 NBDY forms liquid droplets with polyU RNA.mov
Movie S1 NBDY forms liquid droplets with polyU RNA
Thumbnail image of Movie S2 Liquid droplets are dispelled by NBDY phosphorylation.mov
Movie S2 Liquid droplets are dispelled by NBDY phosphorylation
Thumbnail image of Table S1.xlsx
Table S1