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PMID: 7565740 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Endocytosis and vacuolar degradation of the plasma membrane-localized Pdr5 ATP-binding cassette multidrug transporter in Saccharomyces cerevisiae.

Molecular and cellular biology ·Vol. 15 ·No. 11 ·1995-11-00 ·Pages 5879-87

Egner R, Mahé Y, Pandjaitan R, Kuchler K

Abstract

Multidrug resistance (MDR) to different cytotoxic compounds in the yeast Saccharomyces cerevisiae can arise from overexpression of the Pdr5 (Sts1, Ydr1, or Lem1) ATP-binding cassette (ABC) multidrug transporter. We have raised polyclonal antibodies recognizing the yeast Pdr5 ABC transporter to study its biogenesis and to analyze the molecular mechanisms underlying MDR development. Subcellular fractionation and indirect immunofluorescence experiments showed that Pdr5 is localized in the plasma membrane. In addition, pulse-chase radiolabeling of cells and immunoprecipitation indicated that Pdr5 is a short-lived membrane protein with a half-life of about 60 to 90 min. A dramatic metabolic stabilization of Pdr5 was observed in delta pep4 mutant cells defective in vacuolar proteinases, and indirect immunofluorescence showed that Pdr5 accumulates in vacuoles of stationary-phase delta pep4 mutant cells, demonstrating that Pdr5 turnover requires vacuolar proteolysis. However, Pdr5 turnover does not require a functional proteasome, since the half-life of Pdr5 was unaffected in either pre1-1 or pre1-1 pre2-1 mutants defective in the multicatalytic cytoplasmic proteasome that is essential for cytoplasmic protein degradation. Immunofluorescence analysis revealed that vacuolar delivery of Pdr5 is blocked in conditional end4 endocytosis mutants at the restrictive temperature, showing that endocytosis delivers Pdr5 from the plasma membrane to the vacuole.

MeSH Terms
ATP-Binding Cassette Transporters/genetics,metabolism Adenosine Triphosphate/metabolism Base Sequence Biological Transport, Active Cell Compartmentation Cell Membrane/metabolism DNA Primers/chemistry Drug Resistance, Multiple Endocytosis Genes, Fungal Membrane Proteins/genetics Molecular Sequence Data Saccharomyces cerevisiae/metabolism Saccharomyces cerevisiae Proteins Subcellular Fractions/chemistry Vacuoles/pathology
Chemicals
ATP-Binding Cassette Transporters DNA Primers Membrane Proteins PDR5 protein, S cerevisiae Saccharomyces cerevisiae Proteins Adenosine Triphosphate
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Egner R
Department of Molecular Genetics, University and Biocenter of Vienna, Austria.
Mahé Y
Pandjaitan R
Kuchler K
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/MCB.15.11.5879
Published
1995-11-00
Pages
5879-87
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC230839
Subset
IM
External Links
PubMed source DOI: 10.1128/MCB.15.11.5879 PMC Full Text View journal details
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