title: Synchronization of secretory protein traffic in populations of cells.
abstract: To dissect secretory traffic, we developed the retention using selective hooks (RUSH) system. RUSH is a two-state assay based on the reversible interaction of a hook protein fused to core streptavidin and stably anchored in the donor compartment with a reporter protein of interest fused to streptavidin-binding peptide (SBP). Biotin addition causes a synchronous release of the reporter from the hook. Using the RUSH system, we analyzed different transport characteristics of various Golgi and plasma membrane reporters at physiological temperature in living cells. Using dual-color simultaneous live-cell imaging of two cargos, we observed intra- and post-Golgi segregation of cargo traffic, consistent with observation in other systems. We show preliminarily that the RUSH system is usable for automated screening. The system should help increase the understanding of the mechanisms of trafficking and enable screens for molecules that perturb pathological protein transport.
authors.affiliation: Institut Curie, Centre de Recherche, Paris, France.
authors.forename: Gaelle
authors.initials: G
authors.lastname: Boncompain
authors.forename: Severine
authors.initials: S
authors.lastname: Divoux
authors.forename: Nelly
authors.initials: N
authors.lastname: Gareil
authors.forename: Helene
authors.initials: H
authors.lastname: de Forges
authors.forename: Aurianne
authors.initials: A
authors.lastname: Lescure
authors.forename: Lynda
authors.initials: L
authors.lastname: Latreche
authors.forename: Valentina
authors.initials: V
authors.lastname: Mercanti
authors.forename: Florence
authors.initials: F
authors.lastname: Jollivet
authors.forename: Graça
authors.initials: G
authors.lastname: Raposo
authors.forename: Franck
authors.initials: F
authors.lastname: Perez
mesh_terms: D001692:Biological Transport; D002352:Carrier Proteins / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism* / Q000648:ultrastructure; D006056:Golgi Apparatus / Q000378:metabolism* / Q000648:ultrastructure; D049452:Green Fluorescent Proteins / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D018613:Microscopy, Confocal / Q000379:methods*; D016253:Microscopy, Immunoelectron; D019809:Streptavidin / Q000378:metabolism*; D014162:Transfection / Q000379:methods
chemical_list: D002352:Carrier Proteins; C452700:streptavidin-binding peptide; D049452:Green Fluorescent Proteins; D019809:Streptavidin