title: Mobility of tethering factor EEA1 on endosomes is decreased upon stimulation of EGF receptor endocytosis in HeLa cells.
abstract: Tethering factor EEA1, mediating homotypic fusion of early endosomes, was shown to be localized in membrane-bound state both in serum-deprived and stimulated for EGF receptor endocytosis cells. However, it is not known whether dynamics behavior of EEA1 is affected by EGF stimulation. We investigated EEA1 cytosol-to-membrane exchange rate in interphase HeLa cells by FRAP analysis. The data obtained fitted two-states binding model, with the bulk of membrane-associated EEA1 protein represented by the mobile fraction both in serum-starved and EGF-stimulated cells. Fast recovery state had similar half-times in the two cases: about 1.6 s and 2.8 s, respectively. However, the recovery half-time of slowly cycled EEA1 fraction significantly increased in EGF-stimulated comparing to serum-starved cells (from 21 to 99 s). We suppose that the retardation of EEA1 fluorescence recovery upon EGF-stimulation may be due to the increase of activated Rab5 on endosomal membranes, the growth of the number of tethering events between EEA1-positive vesicles and their clustering.
authors.affiliation: Institute of Cytology of RAS, 4, Tikhoretsky Ave, St. Petersburg, 194064, Russia. Electronic address: kosheverova_vera@incras.ru.
authors.forename: Vera V
authors.initials: VV
authors.lastname: Kosheverova
authors.affiliation: Institute of Cytology of RAS, 4, Tikhoretsky Ave, St. Petersburg, 194064, Russia; St. Petersburg State University, 7-9, Universitetskaya nab, St. Petersburg, 199034, Russia. Electronic address: rkamentseva@yandex.ru.
authors.forename: Rimma S
authors.initials: RS
authors.lastname: Kamentseva
authors.affiliation: Institute of Cytology of RAS, 4, Tikhoretsky Ave, St. Petersburg, 194064, Russia. Electronic address: ample@mail.ru.
authors.forename: Ilya V
authors.initials: IV
authors.lastname: Gonchar
authors.affiliation: Institute of Cytology of RAS, 4, Tikhoretsky Ave, St. Petersburg, 194064, Russia. Electronic address: mariannakharchenko@gmail.com.
authors.forename: Marianna V
authors.initials: MV
authors.lastname: Kharchenko
authors.affiliation: Institute of Cytology of RAS, 4, Tikhoretsky Ave, St. Petersburg, 194064, Russia; St. Petersburg State University, 7-9, Universitetskaya nab, St. Petersburg, 199034, Russia; Department of Medical Physics, Peter the Great St. Petersburg Polytechnic University, 29, Polytechnicheskaya, St.Petersburg, 195251, Russia. Electronic address: lenkor@mail.cytspb.rssi.ru.
authors.forename: Elena S
authors.initials: ES
authors.lastname: Kornilova
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004285:Dogs; D004705:Endocytosis; D011992:Endosomes / Q000378:metabolism; D004815:Epidermal Growth Factor / Q000737:chemistry*; D066246:ErbB Receptors / Q000737:chemistry; D036681:Fluorescence Recovery After Photobleaching; D049452:Green Fluorescent Proteins / Q000737:chemistry; D006367:HeLa Cells; D006801:Humans; D061985:Madin Darby Canine Kidney Cells; D008856:Microscopy, Fluorescence; D010957:Plasmids / Q000378:metabolism; D033921:Vesicular Transport Proteins / Q000737:chemistry*
chemical_list: D033921:Vesicular Transport Proteins; C093846:early endosome antigen 1; D049452:Green Fluorescent Proteins; D004815:Epidermal Growth Factor; C512478:EGFR protein, human; D066246:ErbB Receptors
keywords: EEA1; EGF; Endosome; Fluorescent recovery after photobleaching; HeLa cells