Rab3 Proteins Involved in Vesicle Biogenesis and Priming in Embryonic Mouse Chromaffin Cells |
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| Authors: | Jean‐Sébastien Schonn Jan R. T. Van Weering Ralf Mohrmann Oliver M. Schlüter Thomas C. Südhof Heidi De Wit Matthijs Verhage Jakob B. Sørensen |
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| Affiliation: | 1. Membrane biophysics, Max‐Planck‐Institute for Biophysical Chemistry, Am Fassberg 11, D‐37077 G?ttingen, Germany;2. These three authors contributed equally.;3. Department of Functional Genomics, Center for Neurogenomics and Cognitive Research, Neuroscience Campus Amsterdam, Vrije Universiteit (VU) Amsterdam and VU Medical Center, 1081 HV Amsterdam, The Netherlands;4. Present address: Henry Wellcome Integrated Signalling Laboratories, School of Biochemistry, Medical Sciences Building, University of Bristol, Bristol BS8 1TD, UK.;5. Department of Physiology, University of Saarland, 66421 Homburg, Germany;6. Department of Molecular Neurobiology, European Neuroscience Institute, 37077 G?ttingen, Germany;7. Department of Molecular Neurobiology, Max‐Planck‐Institute for Experimental Medicine, Hermann‐Rein Street 3, D‐37075 G?ttingen, Germany;8. Department of Molecular and Cellular Physiology, Howard Hughes Medical Institute, Stanford University, Palo Alto, CA 94304‐5543, USA;9. Department of Neuroscience and Pharmacology, Faculty of Health Sciences, University of Copenhagen, 2200 Copenhagen, Denmark;10. Lundbeck Foundation Center for Biomembranes in Nanomedicine, University of Copenhagen, 2200 Copenhagen, Denmark |
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| Abstract: | The four Rab3 paralogs A–D are involved in exocytosis, but their mechanisms of action are hard to study due to functional redundancy. Here, we used a quadruple Rab3 knockout (KO) (rab3a, rab3b, rab3c, rab3d null, here denoted as ABCD?/?) mouse line to investigate Rab3 function in embryonic mouse adrenal chromaffin cells by electron microscopy and electrophysiological measurements. We show that in cells from ABCD?/? animals large dense‐core vesicles (LDCVs) are less abundant, while the number of morphologically docked granules is normal. By capacitance measurements, we show that deletion of Rab3s reduces the size of the releasable vesicle pools but does not alter their fusion kinetics, consistent with an altered function in vesicle priming. The sustained release component has a sigmoid shape in ABCD?/? cells when normalized to the releasable pool size, indicating that vesicle priming follows at a higher rate after an initial delay. Rescue experiments showed that short‐term (4–6 h) overexpression of Rab3A or Rab3C suffices to rescue vesicle priming and secretion, but it does not restore the number of secretory vesicles. We conclude that Rab3 proteins play two distinct stimulating roles for LDCV fusion in embryonic chromaffin cells, by facilitating vesicle biogenesis and stabilizing the primed vesicle state. |
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| Keywords: | amperometry chromaffin cell exocytosis flash photolysis granule biogenesis GTP‐binding proteins membrane capacitance priming Rab3 |
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