Concentrative sorting of secretory cargo proteins into COPII-coated vesicles.
Here, we show that efficient transport of membrane and secretory proteins from the ER of Saccharomyces cerevisiae requires concentrative and signal-mediated sorting. Three independent markers of bulk flow transport out of the ER indicate that in the absence of an ER export signal, molecules are inefficiently captured into coat protein complex II (COPII)-coated vesicles. A soluble secretory protein, glycosylated pro-alpha-factor (gpalphaf), was enriched approximately 20 fold in these vesicles relative to bulk flow markers. In the absence of Erv29p, a membrane protein that facilitates gpalphaf transport (Belden and Barlowe, 2001), gpalphaf is packaged into COPII vesicles as inefficiently as soluble bulk flow markers. We also found that a plasma membrane protein, the general amino acid permease (Gap1p), is enriched approximately threefold in COPII vesicles relative to membrane phospholipids. Mutation of a diacidic sequence present in the COOH-terminal cytosolic domain of Gap1p eliminated concentrative sorting of this protein.
Malkus
Per
P
Howard Hughes Medical Institute and Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA.
Jiang
Feng
F
Schekman
Randy
R
D000595:Amino Acid Sequence; D026905:Amino Acid Transport Systems; D022181:COP-Coated Vesicles / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D019992:Databases as Topic; D004721:Endoplasmic Reticulum / Q000378:metabolism; D015151:Immunoblotting; D008861:Microsomes / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D011498:Protein Precursors / Q000378:metabolism; D021382:Protein Sorting Signals; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D020703:ras GTPase-Activating Proteins / Q000378:metabolism
D026905:Amino Acid Transport Systems; C491145:GAP1 protein, S cerevisiae; C048632:MF(ALPHA)1 protein, S cerevisiae; D011498:Protein Precursors; D021382:Protein Sorting Signals; D029701:Saccharomyces cerevisiae Proteins; D020703:ras GTPase-Activating Proteins