Yeast Kex1p is a Golgi-associated membrane protein: deletions in a cytoplasmic targeting domain result in mislocalization to the vacuolar membrane.
We have investigated the localization of Kex1p, a type I transmembrane carboxypeptidase involved in precursor processing within the yeast secretory pathway. Indirect immunofluorescence demonstrated the presence of Kex1p in a punctate organelle resembling the yeast Golgi apparatus as identified by Kex2p and Sec7p (Franzusoff, A., K. Redding, J. Crosby, R. S. Fuller, and R. Schekman. 1991. J. Cell Biol. 112:27-37). Glycosylation studies of Kex1p were consistent with a Golgi location, as Kex1p was progressively N-glycosylated in an MNN1-dependent manner. To address the basis of Kex1p targeting to the Golgi apparatus, we examined the cellular location of a series of carboxy-terminal truncations of the protein. The results indicate that a cytoplasmically exposed carboxy-terminal domain is required for retention of this membrane protein within the Golgi apparatus. Deletions of the retention region or overproduction of wild-type Kex1p led to mislocalization of Kex1p to the vacuolar membrane. This unexpected finding is discussed in terms of models involving either the vacuole as a default destination for membrane proteins, or by endocytosis to the vacuole following their default localization to the plasma membrane.
Cooper
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A
Department of Biology, McGill University, Montreal, Canada.
Bussey
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H
D000595:Amino Acid Sequence; D001483:Base Sequence; D001692:Biological Transport; D002240:Carbohydrate Sequence; D002268:Carboxypeptidases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D002451:Cell Compartmentation; D004252:DNA Mutational Analysis; D005455:Fluorescent Antibody Technique; D006031:Glycosylation; D006056:Golgi Apparatus / Q000378:metabolism*; D055646:Killer Factors, Yeast; D008954:Models, Biological; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009183:Mycotoxins / Q000235:genetics / Q000378:metabolism; D011487:Protein Conformation; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D014617:Vacuoles / Q000378:metabolism*
C527157:KHR1 protein, S cerevisiae; D055646:Killer Factors, Yeast; D009183:Mycotoxins; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D002268:Carboxypeptidases; C078874:carboxypeptidase D