Demonstration in yeast of the function of BP-80, a putative plant vacuolar sorting receptor.
BP-80, later renamed VSR(PS-1), is a putative receptor involved in sorting proteins such as proaleurain to the lytic vacuole, with its N-terminal domain recognizing the vacuolar sorting determinant. Although all VSR(PS-1) characteristics and in vitro binding properties described so far favored its receptor function, this function remained to be demonstrated. Here, we used green fluorescent protein (GFP) as a reporter in a yeast mutant strain defective for its own vacuolar receptor, Vps10p. By expressing VSR(PS-1) together with GFP fused to the vacuolar sorting determinant of petunia proaleurain, we were able to efficiently redirect the reporter to the yeast vacuole. VSR(PS-1) is ineffective on GFP either alone or when fused with another type of plant vacuolar sorting determinant from a chitinase. The plant VSR(PS-1) therefore interacts specifically with the proaleurain vacuolar sorting determinant in vivo, and this interaction leads to the transport of the reporter protein through the yeast secretory pathway to the vacuole. This finding demonstrates VSR(PS-1) receptor function but also emphasizes the differences in the spectrum of ligands between Vps10p and its plant equivalent.
Humair
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Laboratoire de Biochimie, rue E. Argand 9, BP2, CH-2007 Neuchâtel, Switzerland.
Hernández Felipe
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Neuhaus
J M
JM
Paris
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D015153:Blotting, Western; D002688:Chitinases / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000378:metabolism; D004792:Enzyme Precursors / Q000378:metabolism; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D008024:Ligands; D008164:Luminescent Proteins / Q000235:genetics; D018613:Microscopy, Confocal; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D010944:Plants / Q000235:genetics / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014617:Vacuoles / Q000378:metabolism*; D033921:Vesicular Transport Proteins; D015003:Yeasts / Q000235:genetics / Q000378:metabolism*
D004792:Enzyme Precursors; D008024:Ligands; D008164:Luminescent Proteins; D010940:Plant Proteins; D011956:Receptors, Cell Surface; D011993:Recombinant Fusion Proteins; D033921:Vesicular Transport Proteins; D049452:Green Fluorescent Proteins; D002688:Chitinases; D003546:Cysteine Endopeptidases; C076060:proaleurain