title: Genetic visualization of protein interactions harnessing liquid phase transitions.
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.
authors.affiliation: Laboratory for Cell Function and Dynamics, Brain Science Institute, RIKEN, Saitama 351-0198, Japan.|Ina Laboratory, Medical &Biological Laboratories Co., Ltd., Ina, Nagano, Japan.|Amalgaam Co. Ltd., Ina, Nagano, Japan.
authors.forename: Taku
authors.initials: T
authors.lastname: Watanabe
authors.affiliation: Laboratory for Cell Function and Dynamics, Brain Science Institute, RIKEN, Saitama 351-0198, Japan.|Ina Laboratory, Medical &Biological Laboratories Co., Ltd., Ina, Nagano, Japan.|Amalgaam Co. Ltd., Ina, Nagano, Japan.
authors.forename: Tatsuya
authors.initials: T
authors.lastname: Seki
authors.affiliation: Laboratory for Cell Function and Dynamics, Brain Science Institute, RIKEN, Saitama 351-0198, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Fukano
authors.affiliation: Laboratory for Cell Function and Dynamics, Brain Science Institute, RIKEN, Saitama 351-0198, Japan.
authors.forename: Asako
authors.initials: A
authors.lastname: Sakaue-Sawano
authors.affiliation: Laboratory for Cell Function and Dynamics, Brain Science Institute, RIKEN, Saitama 351-0198, Japan.|Ina Laboratory, Medical &Biological Laboratories Co., Ltd., Ina, Nagano, Japan.|Amalgaam Co. Ltd., Ina, Nagano, Japan.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Karasawa
authors.affiliation: Laboratory for Cell Function and Dynamics, Brain Science Institute, RIKEN, Saitama 351-0198, Japan.
authors.forename: Misaki
authors.initials: M
authors.lastname: Kubota
authors.affiliation: Laboratory for Cell Function and Dynamics, Brain Science Institute, RIKEN, Saitama 351-0198, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Kurokawa
authors.affiliation: Ina Laboratory, Medical &Biological Laboratories Co., Ltd., Ina, Nagano, Japan.
authors.forename: Ken
authors.initials: K
authors.lastname: Inoue
authors.affiliation: Ina Laboratory, Medical &Biological Laboratories Co., Ltd., Ina, Nagano, Japan.
authors.forename: Junichi
authors.initials: J
authors.lastname: Akatsuka
authors.affiliation: Laboratory for Cell Function and Dynamics, Brain Science Institute, RIKEN, Saitama 351-0198, Japan.|Biotechnological Optics Research Team, Center for Advanced Photonics, RIKEN, Saitama 351-0198, Japan.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Miyawaki
mesh_terms: D051017:Apoptosis Regulatory Proteins; D001665:Binding Sites; D055592:Biophysical Phenomena; D005456:Fluorescent Dyes / Q000737:chemistry*; D006367:HeLa Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D024101:Mitochondrial Proteins / Q000737:chemistry / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000737:chemistry / Q000378:metabolism; D000072417:Protein Domains; D025941:Protein Interaction Mapping / Q000379:methods*; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D051636:X-Linked Inhibitor of Apoptosis Protein / Q000737:chemistry / Q000378:metabolism
chemical_list: D051017:Apoptosis Regulatory Proteins; C412403:DIABLO protein, human; D005456:Fluorescent Dyes; D047908:Intracellular Signaling Peptides and Proteins; D024101:Mitochondrial Proteins; C118585:P62 protein, human; D011506:Proteins; D016601:RNA-Binding Proteins; D051636:X-Linked Inhibitor of Apoptosis Protein; C496866:XIAP protein, human; C535150:MAPK1 protein, human; D019950:Mitogen-Activated Protein Kinase 1