Coding variants identified in patients with diabetes alter PICK1 BAR domain function in insulin granule biogenesis

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  • Joscha Rombach
  • Matthew D. Lycas
  • Nikolaj R. Christensen
  • Donnie S. Stapleton
  • Mathias A. Olsen
  • Mikkel Stoklund
  • Tommas T.E. Nielsen
  • Mark P. Keller
  • Anna M. Jansen
  • Massimo Pietropaolo
  • Jens B. Simonsen
  • Alan D. Attie

Bin/amphiphysin/Rvs (BAR) domains are positively charged crescent-shaped modules that mediate curvature of negatively charged lipid membranes during remodeling processes. The BAR domain proteins PICK1, ICA69, and the arfaptins have recently been demonstrated to coordinate the budding and formation of immature secretory granules (ISGs) at the trans- Golgi network. Here, we identify 4 coding variants in the PICK1 gene from a whole-exome screening of Danish patients with diabetes that each involve a change in positively charged residues in the PICK1 BAR domain. All 4 coding variants failed to rescue insulin content in INS-1E cells upon knock down of endogenous PICK1. Moreover, 2 variants showed dominantnegative properties. In vitro assays addressing BAR domain function suggested that the coding variants compromised BAR domain function but increased the capacity to cause fission of liposomes. Live confocal microscopy and super-resolution microscopy further revealed that PICK1 resides transiently on ISGs before egress via vesicular budding events. Interestingly, this egress of PICK1 was accelerated in the coding variants. We propose that PICK1 assists in or complements the removal of excess membrane and generic membrane trafficking proteins, and possibly also insulin, from ISGs during the maturation process; and that the coding variants may cause premature budding, possibly explaining their dominant-negative function.

TidsskriftJournal of Clinical Investigation
Udgave nummer5
StatusUdgivet - 2022

Bibliografisk note

Funding Information:
We thank Torben Hansen, University of Copenhagen, and the Lundbeck Foundation Centre for Applied Medical Genomics in Personalised Disease Prediction, Prevention, and Care (LuCamp) (Denmark) for access to the WES data. We are grateful to Nabeela Khadim and Anders Bohl Pedersens for excellent technical assistance. We thank Nils Billestrup, University of Copenhagen, for expertise and guidance with the primary β cell study. We thank Thomas Hartig Braunstein and Pablo Hernandez-Varas of the Core Facility for Integrated Microscopy, Department of Biomedical Sciences, University of Copenhagen, for help with microscopy; and Elle Kielar Grevstad of the Biochemistry Optical Core, University of Madison–Wisconsin, for help with live-cell microscopy. We also thank Robert C. Malenka for the shRNA constructs, Richard Huganir for the pHsSynXW vector, and Peter Arvan for the GRINCH cells. The work was supported by the Danish Research Council, Health and Disease, grant 6110-00625B; and the Novo Nordisk Foundation, grant NNF18OCT0034152.

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© 2022 American Society for Clinical Investigation. All rights reserved.

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