Genetic visualization of protein interactions harnessing liquid phase transitions

Sci Rep. 2017 Apr 13:7:46380. doi: 10.1038/srep46380.

Abstract

Protein-protein interactions (PPIs) are essential components of cellular function. Current fluorescence-based technologies to measure PPIs have limited dynamic range and quantitative reproducibility. Here, we describe a genetically-encoded PPI visualization system that harnesses the dynamics of condensed liquid-phase transitions to analyze protein interactions in living cells. The fluorescent protein Azami-Green and p62-PB1 domain when fused to PPI partners triggered a rapid concatenation/oligomerization process that drove the condensation of liquid-phase droplets for real-time analysis of the interaction with unlimited dynamic range in the fluorescence signal. Proof-of-principle studies revealed novel insights on the live cell dynamics of XIAP-Smac and ERK2-dimer interactions. A photoconvertible variant allowed time-resolved optical highlighting for PPI kinetic analysis. Our system, called Fluoppi, demonstrates the unique signal amplification properties of liquid-phase condensation to detect PPIs. The findings introduce a general method for discovery of novel PPIs and modulators of established PPIs.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Apoptosis Regulatory Proteins
  • Binding Sites
  • Biophysical Phenomena
  • Fluorescent Dyes / chemistry*
  • HeLa Cells
  • Humans
  • Intracellular Signaling Peptides and Proteins / chemistry
  • Intracellular Signaling Peptides and Proteins / metabolism
  • Kinetics
  • Mitochondrial Proteins / chemistry
  • Mitochondrial Proteins / metabolism
  • Mitogen-Activated Protein Kinase 1 / chemistry
  • Mitogen-Activated Protein Kinase 1 / metabolism
  • Protein Domains
  • Protein Interaction Mapping / methods*
  • Proteins / chemistry*
  • Proteins / metabolism
  • RNA-Binding Proteins / chemistry
  • RNA-Binding Proteins / metabolism
  • X-Linked Inhibitor of Apoptosis Protein / chemistry
  • X-Linked Inhibitor of Apoptosis Protein / metabolism

Substances

  • Apoptosis Regulatory Proteins
  • DIABLO protein, human
  • Fluorescent Dyes
  • Intracellular Signaling Peptides and Proteins
  • Mitochondrial Proteins
  • P62 protein, human
  • Proteins
  • RNA-Binding Proteins
  • X-Linked Inhibitor of Apoptosis Protein
  • XIAP protein, human
  • MAPK1 protein, human
  • Mitogen-Activated Protein Kinase 1