title: Electron tomography reveals Rab6 is essential to the trafficking of trans-Golgi clathrin and COPI-coated vesicles and the maintenance of Golgi cisternal number.
abstract: We have shown previously that Rab6, a small, trans-Golgi-localized GTPase, acts upstream of the conserved oligomeric Golgi complex (COG) and ZW10/RINT1 retrograde tether complexes to maintain Golgi homeostasis. In this article, we present evidence from the unbiased and high-resolution approach of electron microscopy and electron tomography that Rab6 is essential to the trans-Golgi trafficking of two morphological classes of coated vesicles; the larger corresponds to clathrin-coated vesicles and the smaller to coat protein I (COPI)-coated vesicles. On the basis of the site of coated vesicle accumulation, cisternal dilation and the normal kinetics of cargo transport from the endoplasmic reticulum (ER) to Golgi followed by delayed Golgi to cell surface transport, we suggest that Golgi function in cargo transport is preferentially inhibited at the trans-Golgi/trans-Golgi network (TGN). The >50% increase in Golgi cisternae number in Rab6-depleted HeLa cells that we observed may well be coupled to the trans-Golgi accumulation of COPI-coated vesicles; depletion of the individual Rab6 effector, myosin IIA, produced an accumulation of uncoated vesicles with if anything a decrease in cisternal number. These results are the first evidence for a Rab6-dependent protein machine affecting Golgi-proximal, coated vesicle accumulation and probably transport at the trans-Golgi and the first example of concomitant cisternal proliferation and increased Golgi stack organization under inhibited transport conditions.
authors.affiliation: Department of Physiology and Biophysics, University of Arkansas for Medical Sciences, Little Rock, AR 72205, USA.
authors.forename: Brian
authors.initials: B
authors.lastname: Storrie
authors.forename: Massimo
authors.initials: M
authors.lastname: Micaroni
authors.forename: Garry P
authors.initials: GP
authors.lastname: Morgan
authors.forename: Nick
authors.initials: N
authors.lastname: Jones
authors.forename: Jeffrey A
authors.initials: JA
authors.lastname: Kamykowski
authors.forename: Ngozi
authors.initials: N
authors.lastname: Wilkins
authors.forename: Timothy H
authors.initials: TH
authors.lastname: Pan
authors.forename: Brad J
authors.initials: BJ
authors.lastname: Marsh
mesh_terms: D001692:Biological Transport; D022181:COP-Coated Vesicles / Q000378:metabolism*; D055032:Electron Microscope Tomography / Q000379:methods*; D006056:Golgi Apparatus / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006367:HeLa Cells; D006706:Homeostasis; D006801:Humans; D007091:Image Processing, Computer-Assisted; D008854:Microscopy, Electron / Q000379:methods; D024462:Myosin Type II / Q000378:metabolism; D010641:Phenotype; D020033:Protein Isoforms; D021381:Protein Transport; D034741:RNA, Small Interfering / Q000378:metabolism; D020691:rab GTP-Binding Proteins / Q000378:metabolism*; D021601:trans-Golgi Network / Q000378:metabolism
chemical_list: D020033:Protein Isoforms; D034741:RNA, Small Interfering; C094135:Rab6 protein; D049452:Green Fluorescent Proteins; D024462:Myosin Type II; D020691:rab GTP-Binding Proteins