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Sequential transport of protein between the endoplasmic-reticulum and successive golgi compartments in semi-intact cells

Academic Article
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Overview

authors

  • Schwaninger, R.
  • Beckers, C. J. M.
  • Balch, William E.

publication date

  • July 1991

journal

  • Journal of Biological Chemistry  Journal

abstract

  • The vectorial transport of vesicular stomatitis virus (VSV) G protein between the ER and the cis and medial Golgi compartments has been reconstituted using semi-intact (perforated) cells. The transport of VSV-G protein between successive compartments is measured by the sequential processing of the two N-linked oligosaccharide chains present on VSV-G protein to the endoglycosidase (endo) H-resistant structures which have unique electrophoretic mobilities during sodium dodecyl sulfate-gel electrophoresis. The appearance of a form of VSV-G which contains only one endo H-resistant oligosaccharide chain (GH1) is kinetically and biochemically indistinguishable from the appearance of the Man5, endo D-sensitive form (GD), the latter being a processing reaction diagnostic of transport from the ER to the cis Golgi compartment. These results provide evidence that the cis Golgi compartment may contain in addition to alpha-1,2-mannosidase I, both N-acetylglueosamine transferase I and alpha-1,2-mannosidase II. VSV-G protein is subsequently processed to the form which contains two endo H-resistant oligosaccharides (GH2) after a second wave of vesicular transport. Processing of GH1 to GH2 in vitro occurs only after a lag period following the appearance of GH1; processing is sensitive to N-ethylmaleimide, guanosine-5'-O-(3-thiotriphosphate), and a synthetic peptide homologous to the rab1 protein effector domain, and processing is inhibited in the absence of free Ca2+ (in the presence of EGTA), reagents which potently inhibit ER to cis Golgi transport. These results suggest that VSV-G protein proceeds through at least two rounds of vesicular transport from the ER to the medial Golgi compartment for processing to the GH2 form, providing a model system to study the regulation of the vectorial membrane fission and fusion events involved in vesicular trafficking and organelle dynamics in the early stages of the secretory pathway.

subject areas

  • Animals
  • Calcium
  • Cell Line
  • Egtazic Acid
  • Endoplasmic Reticulum
  • Ethylmaleimide
  • Golgi Apparatus
  • Guanosine 5'-O-(3-Thiotriphosphate)
  • Kinetics
  • Membrane Glycoproteins
  • Protein Processing, Post-Translational
  • Vesicular stomatitis Indiana virus
  • Viral Envelope Proteins
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Identity

International Standard Serial Number (ISSN)

  • 0021-9258

PubMed ID

  • 1649174
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Additional Document Info

start page

  • 13055

end page

  • 13063

volume

  • 266

issue

  • 20

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