title: RefSOFI for Mapping Nanoscale Organization of Protein-Protein Interactions in Living Cells.
abstract: It has become increasingly clear that protein-protein interactions (PPIs) are compartmentalized in nanoscale domains that define the biochemical architecture of the cell. Despite tremendous advances in super-resolution imaging, strategies to observe PPIs at sufficient resolution to discern their organization are just emerging. Here we describe a strategy in which PPIs induce reconstitution of fluorescent proteins (FPs) that are capable of exhibiting single-molecule fluctuations suitable for stochastic optical fluctuation imaging (SOFI). Subsequently, spatial maps of these interactions can be resolved in super-resolution in living cells. Using this strategy, termed reconstituted fluorescence-based SOFI (refSOFI), we investigated the interaction between the endoplasmic reticulum (ER) Ca(2+) sensor STIM1 and the pore-forming channel subunit ORAI1, a crucial process in store-operated Ca(2+) entry (SOCE). Stimulating SOCE does not appear to change the size of existing STIM1/ORAI1 interaction puncta at the ER-plasma membrane junctions, but results in an apparent increase in the number of interaction puncta.
authors.affiliation: Department of Pharmacology, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093, USA.
authors.forename: Fabian
authors.initials: F
authors.lastname: Hertel
authors.affiliation: Department of Pharmacology, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093, USA.
authors.forename: Gary C H
authors.initials: GC
authors.lastname: Mo
authors.affiliation: Department of Chemistry, University of Leuven, Celestijnenlaan 200F, 3001 Heverlee, Belgium.
authors.forename: Sam
authors.initials: S
authors.lastname: Duwé
authors.affiliation: Department of Chemistry, University of Leuven, Celestijnenlaan 200F, 3001 Heverlee, Belgium.
authors.forename: Peter
authors.initials: P
authors.lastname: Dedecker
authors.affiliation: Department of Pharmacology, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093, USA; Department of Pharmacology and Molecular Sciences, The Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA. Electronic address: jzhang32@ucsd.edu.
authors.forename: Jin
authors.initials: J
authors.lastname: Zhang
mesh_terms: D002460:Cell Line; D002478:Cells, Cultured; D036103:Nanotechnology; D054730:Protein Interaction Domains and Motifs / Q000235:genetics*