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Purification of age-distinct insulin secretory granules through antigen restriction

View ORCID ProfileMartin Neukam, Pia Sala, Andreas-David Brunner, Katharina Ganß, Alessandra Palladini, Michal Grzybek, View ORCID ProfileJovana Vasiljević, View ORCID ProfileJohannes Broichhagen, View ORCID ProfileKai Johnsson, View ORCID ProfileThomas Kurth, Matthias Mann, Ünal Coskun, View ORCID ProfileMichele Solimena
doi: https://doi.org/10.1101/2020.06.03.103770
Martin Neukam
1Molecular Diabetology, University Hospital and Faculty of Medicine Carl Gustav Carus, TU Dresden, Dresden, Germany
2Paul Langerhans Institute Dresden (PLID) of the Helmholtz Center Munich at the University Hospital Carl Gustav Carus and Faculty of Medicine of the TU Dresden, Dresden, Germany
3German Center for Diabetes Research (DZD), Neuherberg, Germany
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  • ORCID record for Martin Neukam
  • For correspondence: martin.neukam@tu-dresden.de michele.solimena@uniklinikum-dresden.de
Pia Sala
2Paul Langerhans Institute Dresden (PLID) of the Helmholtz Center Munich at the University Hospital Carl Gustav Carus and Faculty of Medicine of the TU Dresden, Dresden, Germany
3German Center for Diabetes Research (DZD), Neuherberg, Germany
4Center of Membrane Biochemistry and Lipid Research, University Hospital and Faculty of Medicine Carl Gustav Carus, TU Dresden, Dresden, Germany
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Andreas-David Brunner
5Max Planck Institute of Biochemistry, Martinsried, Germany
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Katharina Ganß
1Molecular Diabetology, University Hospital and Faculty of Medicine Carl Gustav Carus, TU Dresden, Dresden, Germany
2Paul Langerhans Institute Dresden (PLID) of the Helmholtz Center Munich at the University Hospital Carl Gustav Carus and Faculty of Medicine of the TU Dresden, Dresden, Germany
3German Center for Diabetes Research (DZD), Neuherberg, Germany
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Alessandra Palladini
2Paul Langerhans Institute Dresden (PLID) of the Helmholtz Center Munich at the University Hospital Carl Gustav Carus and Faculty of Medicine of the TU Dresden, Dresden, Germany
3German Center for Diabetes Research (DZD), Neuherberg, Germany
4Center of Membrane Biochemistry and Lipid Research, University Hospital and Faculty of Medicine Carl Gustav Carus, TU Dresden, Dresden, Germany
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Michal Grzybek
2Paul Langerhans Institute Dresden (PLID) of the Helmholtz Center Munich at the University Hospital Carl Gustav Carus and Faculty of Medicine of the TU Dresden, Dresden, Germany
3German Center for Diabetes Research (DZD), Neuherberg, Germany
4Center of Membrane Biochemistry and Lipid Research, University Hospital and Faculty of Medicine Carl Gustav Carus, TU Dresden, Dresden, Germany
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Jovana Vasiljević
1Molecular Diabetology, University Hospital and Faculty of Medicine Carl Gustav Carus, TU Dresden, Dresden, Germany
2Paul Langerhans Institute Dresden (PLID) of the Helmholtz Center Munich at the University Hospital Carl Gustav Carus and Faculty of Medicine of the TU Dresden, Dresden, Germany
3German Center for Diabetes Research (DZD), Neuherberg, Germany
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Johannes Broichhagen
6Max Planck Institute for Medical Research, Department of Chemical Biology, Heidelberg, Germany
7Leibniz-Forschungsinstitut für Molekulare Pharmakologie, Berlin, Germany
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Kai Johnsson
6Max Planck Institute for Medical Research, Department of Chemical Biology, Heidelberg, Germany
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Thomas Kurth
8TU Dresden, Center for Molecular and Cellular Bioengineering (CMCB), Technology Platform, Electron Microscopy and Histology Facility, Dresden, Germany
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Matthias Mann
5Max Planck Institute of Biochemistry, Martinsried, Germany
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Ünal Coskun
2Paul Langerhans Institute Dresden (PLID) of the Helmholtz Center Munich at the University Hospital Carl Gustav Carus and Faculty of Medicine of the TU Dresden, Dresden, Germany
3German Center for Diabetes Research (DZD), Neuherberg, Germany
4Center of Membrane Biochemistry and Lipid Research, University Hospital and Faculty of Medicine Carl Gustav Carus, TU Dresden, Dresden, Germany
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Michele Solimena
1Molecular Diabetology, University Hospital and Faculty of Medicine Carl Gustav Carus, TU Dresden, Dresden, Germany
2Paul Langerhans Institute Dresden (PLID) of the Helmholtz Center Munich at the University Hospital Carl Gustav Carus and Faculty of Medicine of the TU Dresden, Dresden, Germany
3German Center for Diabetes Research (DZD), Neuherberg, Germany
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  • ORCID record for Michele Solimena
  • For correspondence: martin.neukam@tu-dresden.de michele.solimena@uniklinikum-dresden.de
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Summary

Endocrine cells employ regulated exocytosis of secretory granules to secrete hormones and neurotransmitters. The competence of secretory granules for exocytosis depends on spatio-temporal variables such as proximity to the plasma membrane and age, with newly-generated granules being preferentially released. Despite recent advances, we still lack a comprehensive view about the molecular composition of insulin granules and how this may change over the lifetime of the organelles. Here we report a strategy for the purification of insulin secretory granules of distinct age from insulinoma INS-1 cells. Tagging the granule-resident protein phogrin with a cleavable CLIP tag, we obtained intact fractions of age-distinct granules for proteomic and lipidomic analyses. We find that the lipid composition changes over time along with the physical properties of the membrane.

Competing Interest Statement

The authors have declared no competing interest.

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The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. All rights reserved. No reuse allowed without permission.
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Posted April 29, 2022.
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Purification of age-distinct insulin secretory granules through antigen restriction
Martin Neukam, Pia Sala, Andreas-David Brunner, Katharina Ganß, Alessandra Palladini, Michal Grzybek, Jovana Vasiljević, Johannes Broichhagen, Kai Johnsson, Thomas Kurth, Matthias Mann, Ünal Coskun, Michele Solimena
bioRxiv 2020.06.03.103770; doi: https://doi.org/10.1101/2020.06.03.103770
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Purification of age-distinct insulin secretory granules through antigen restriction
Martin Neukam, Pia Sala, Andreas-David Brunner, Katharina Ganß, Alessandra Palladini, Michal Grzybek, Jovana Vasiljević, Johannes Broichhagen, Kai Johnsson, Thomas Kurth, Matthias Mann, Ünal Coskun, Michele Solimena
bioRxiv 2020.06.03.103770; doi: https://doi.org/10.1101/2020.06.03.103770

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