Transport through the yeast endocytic pathway occurs through morphologically distinct compartments and requires an active secretory pathway and Sec18p/N-ethylmaleimide-sensitive fusion protein.
Molecules travel through the yeast endocytic pathway from the cell surface to the lysosome-like vacuole by passing through two sequential intermediates. Immunofluorescent detection of an endocytosed pheromone receptor was used to morphologically identify these intermediates, the early and late endosomes. The early endosome is a peripheral organelle that is heterogeneous in appearance, whereas the late endosome is a large perivacuolar compartment that corresponds to the prevacuolar compartment previously shown to be an endocytic intermediate. We demonstrate that inhibiting transport through the early secretory pathway in sec mutants quickly impedes transport from the early endosome. Treatment of sensitive cells with brefeldin A also blocks transport from this compartment. We provide evidence that Sec18p/N-ethylmaleimide-sensitive fusion protein, a protein required for membrane fusion, is directly required in vivo for forward transport early in the endocytic pathway. Inhibiting protein synthesis does not affect transport from the early endosome but causes endocytosed proteins to accumulate in the late endosome. As newly synthesized proteins and the late steps of secretion are not required for early to late endosome transport, but endoplasmic reticulum through Golgi traffic is, we propose that efficient forward transport in the early endocytic pathway requires delivery of lipid from secretory organelles to endosomes.
Hicke
L
L
Department of Biochemistry, Biozentrum, University of Basel, Switzerland.
Zanolari
B
B
Pypaert
M
M
Rohrer
J
J
Riezman
H
H
D000251:Adenosine Triphosphatases; D001692:Biological Transport / Q000187:drug effects; D020126:Brefeldin A; D002268:Carboxypeptidases / Q000378:metabolism; D043402:Cathepsin A; D002451:Cell Compartmentation / Q000502:physiology*; D002462:Cell Membrane; D003517:Cyclopentanes / Q000494:pharmacology; D004705:Endocytosis / Q000502:physiology*; D011992:Endosomes / Q000378:metabolism; D005033:Ethylmaleimide / Q000494:pharmacology*; D005656:Fungal Proteins / Q000187:drug effects / Q000378:metabolism*; D051964:Receptors, Mating Factor; D018000:Receptors, Peptide / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D014617:Vacuoles / Q000378:metabolism; D033921:Vesicular Transport Proteins; D015003:Yeasts / Q000235:genetics / Q000378:metabolism*
D003517:Cyclopentanes; D005656:Fungal Proteins; D051964:Receptors, Mating Factor; D018000:Receptors, Peptide; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D033921:Vesicular Transport Proteins; D020126:Brefeldin A; D002268:Carboxypeptidases; D043402:Cathepsin A; D000251:Adenosine Triphosphatases; C064404:SEC18 protein, S cerevisiae; D005033:Ethylmaleimide