Selective endocytosis controls slit diaphragm maintenance and dynamics in Drosophila nephrocytes

  1. Konrad Lang
  2. Julian Milosavljevic
  3. Helena Heinkele
  4. Mengmeng Chen
  5. Lea Gerstner
  6. Dominik Spitz
  7. Severine Kayser
  8. Martin Helmstädter
  9. Gerd Walz
  10. Michael Köttgen
  11. Andrew Spracklen
  12. John Poulton
  13. Tobias Hermle  Is a corresponding author
  1. University of Freiburg, Germany
  2. University of North Carolina at Chapel Hill, United States

Abstract

The kidneys generate about 180 liters of primary urine per day by filtration of plasma. An essential part of the filtration barrier is the slit diaphragm, a multiprotein complex containing nephrin as major component. Filter dysfunction typically manifests with proteinuria and mutations in endocytosis regulating genes were discovered as causes of proteinuria. However, it is unclear how endocytosis regulates the slit diaphragm and how the filtration barrier is maintained without either protein leakage or filter clogging. Here we study nephrin dynamics in podocyte-like nephrocytes of Drosophila and show that selective endocytosis either by dynamin- or flotillin-mediated pathways regulates a stable yet highly dynamic architecture. Short-term manipulation of endocytic functions indicates that dynamin-mediated endocytosis of ectopic nephrin restricts slit diaphragm formation spatially while flotillin-mediated turnover of nephrin within the slit diaphragm is needed to maintain filter permeability by shedding of molecules bound to nephrin in endosomes. Since slit diaphragms cannot be studied in vitro and are poorly accessible in mouse models, this is the first analysis of their dynamics within the slit diaphragm multiprotein complex. Identification of the mechanisms of slit diaphragm maintenance will help to develop novel therapies for proteinuric renal diseases that are frequently limited to symptomatic treatment.

Data availability

Transgenic Drosophila lines are available from the corresponding author upon reasonable request. Unprocessed image files were submitted to a public repository (zenodo.org, DOI: 10.5281/zenodo.6418762). Access is not restricted for scientific purposes.

The following data sets were generated

Article and author information

Author details

  1. Konrad Lang

    Department of Medicine, University of Freiburg, Freiburg, Germany
    Competing interests
    The authors declare that no competing interests exist.
  2. Julian Milosavljevic

    Department of Medicine, University of Freiburg, Freiburg, Germany
    Competing interests
    The authors declare that no competing interests exist.
  3. Helena Heinkele

    Department of Medicine, University of Freiburg, Freiburg, Germany
    Competing interests
    The authors declare that no competing interests exist.
  4. Mengmeng Chen

    Department of Medicine, University of Freiburg, Freiburg, Germany
    Competing interests
    The authors declare that no competing interests exist.
  5. Lea Gerstner

    Department of Medicine, University of Freiburg, Freiburg, Germany
    Competing interests
    The authors declare that no competing interests exist.
  6. Dominik Spitz

    Department of Medicine, University of Freiburg, Freiburg, Germany
    Competing interests
    The authors declare that no competing interests exist.
  7. Severine Kayser

    Department of Medicine, University of Freiburg, Freiburg, Germany
    Competing interests
    The authors declare that no competing interests exist.
  8. Martin Helmstädter

    Department of Medicine, University of Freiburg, Freiburg, Germany
    Competing interests
    The authors declare that no competing interests exist.
  9. Gerd Walz

    Department of Medicine, University of Freiburg, Freiburg, Germany
    Competing interests
    The authors declare that no competing interests exist.
  10. Michael Köttgen

    Department of Medicine, University of Freiburg, Freiburg, Germany
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-2406-5039
  11. Andrew Spracklen

    Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-5550-8595
  12. John Poulton

    Department of Medicine, University of North Carolina at Chapel Hill, Chapel Hill, United States
    Competing interests
    The authors declare that no competing interests exist.
  13. Tobias Hermle

    Department of Medicine, University of Freiburg, Freiburg, Germany
    For correspondence
    tobias.hermle@uniklinik-freiburg.de
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-0441-7749

Funding

Deutsche Forschungsgemeinschaft (project-ID 431984000)

  • Tobias Hermle

Deutsche Forschungsgemeinschaft (HE 7456/3-1)

  • Tobias Hermle

Deutsche Forschungsgemeinschaft (HE 7456/4-1)

  • Tobias Hermle

Deutsche Gesellschaft für Innere Medizin (Clinician Scientist Fellowship)

  • Tobias Hermle

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

Reviewing Editor

  1. Ilse S Daehn, Icahn School of Medicine at Mount Sinai, United States

Publication history

  1. Preprint posted: March 2, 2022 (view preprint)
  2. Received: March 28, 2022
  3. Accepted: July 24, 2022
  4. Accepted Manuscript published: July 25, 2022 (version 1)
  5. Version of Record published: August 4, 2022 (version 2)

Copyright

© 2022, Lang 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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  1. Konrad Lang
  2. Julian Milosavljevic
  3. Helena Heinkele
  4. Mengmeng Chen
  5. Lea Gerstner
  6. Dominik Spitz
  7. Severine Kayser
  8. Martin Helmstädter
  9. Gerd Walz
  10. Michael Köttgen
  11. Andrew Spracklen
  12. John Poulton
  13. Tobias Hermle
(2022)
Selective endocytosis controls slit diaphragm maintenance and dynamics in Drosophila nephrocytes
eLife 11:e79037.
https://doi.org/10.7554/eLife.79037

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