1. Cell Biology
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A systematic approach to identify recycling endocytic cargo depending on the GARP complex

  1. Sebastian Eising
  2. Lisa Thiele
  3. Florian Fröhlich  Is a corresponding author
  1. University of Osnabrück, Germany
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Cite this article as: eLife 2019;8:e42837 doi: 10.7554/eLife.42837

Abstract

Proteins and lipids of the plasma membrane underlie constant remodeling via a combination of the secretory- and the endocytic pathway. In the yeast endocytic pathway, cargo is sorted for recycling to the plasma membrane or degradation in vacuoles. Previously we have shown a role for the GARP complex in sphingolipid sorting and homeostasis (Fröhlich et al. 2015). However, the majority of cargo sorted in a GARP dependent process remain largely unknown. Here we use auxin induced degradation of GARP combined with mass spectrometry based vacuolar proteomics and lipidomics to show that recycling of two specific groups of proteins, the amino-phospholipid flippases and cell wall synthesis proteins depends on a functional GARP complex. Our results suggest that mis-sorting of flippases and remodeling of the lipid composition are the first occurring defects in GARP mutants. Our assay can be adapted to systematically map cargo of the entire endocytic pathway.

Data availability

All data generated or analysed during this study are included in the manuscript and supporting files. Source data files have been provided for Figures 3 and 4.

Article and author information

Author details

  1. Sebastian Eising

    Department of Biology/Chemistry, University of Osnabrück, Osnabrück, Germany
    Competing interests
    The authors declare that no competing interests exist.
  2. Lisa Thiele

    Department of Biology/Chemistry, University of Osnabrück, Osnabrück, Germany
    Competing interests
    The authors declare that no competing interests exist.
  3. Florian Fröhlich

    Department of Biology/Chemistry, University of Osnabrück, Osnabrück, Germany
    For correspondence
    florian.froehlich@biologie.uni-osnabrueck.de
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8307-2189

Funding

Deutsche Forschungsgemeinschaft (FR 3647/2-1)

  • Florian Fröhlich

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Peter Tontonoz, University of California, Los Angeles, United States

Publication history

  1. Received: October 19, 2018
  2. Accepted: January 29, 2019
  3. Accepted Manuscript published: January 29, 2019 (version 1)
  4. Version of Record published: February 13, 2019 (version 2)

Copyright

© 2019, Eising et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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