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Vol. 12, Issue 4, 981-995, April 2001
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and¶
*Department of Pediatrics, Division of Gastroenterology and
Nutrition, Vesicles carrying recycling plasma membrane proteins from early
endosomes have not yet been characterized. Using Chinese hamster ovary
cells transfected with the facilitative glucose transporter, GLUT4, we identified two classes of discrete, yet similarly
sized, small vesicles that are derived from early endosomes. We refer to these postendosomal vesicles as endocytic small vesicles or ESVs.
One class of ESVs contains a sizable fraction of the pool of the
transferrin receptor, and the other contains 40% of the total
cellular pool of GLUT4 and is enriched in the insulin-responsive aminopeptidase (IRAP). The ESVs contain cellubrevin and
Rab4 but are lacking other early endosomal markers, such as EEA1 or
syntaxin13. The ATP-, temperature-, and cytosol-dependent formation of
ESVs has been reconstituted in vitro from endosomal membranes.
Guanosine 5'-[The Hormone Research Institute,
Department of Anatomy, and §Department of
Neurology University of California, San Francisco, San Francisco,
California 94143
-thio]triphosphate and neomycin, but not brefeldin
A, inhibit budding of the ESVs in vitro. A monoclonal antibody
recognizing the GLUT4 cytoplasmic tail perturbs the in vitro targeting
of GLUT4 to the ESVs without interfering with the incorporation of IRAP
or TfR. We suggest that cytosolic proteins mediate the incorporation of
recycling membrane proteins into discrete populations of ESVs that
serve as carrier vesicles to store and then transport the cargo from
early endosomes, either directly or indirectly, to the cell surface.
Corresponding author and present address:
Merck Research Laboratories, RY 33-672, Rahway, NJ 07065; e-mail:
Gary_Herman{at}Merck.com.
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