Endosomal Fusion upon SNARE Knockdown is Maintained by Residual SNARE Activity and Enhanced Docking |
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Authors: | Ioanna Bethani Achim Werner Chandini Kadian Ulf Geumann Reinhard Jahn Silvio O. Rizzoli |
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Affiliation: | Department of Neurobiology, Max-Planck-Institute for Biophysical Chemistry, Am Fassberg 11, 37077 Göttingen, Germany; International Max Planck Research School for Neurosciences, Göttingen, Germany; International Max Planck Research School for Molecular Biology, Göttingen, Germany; Current address: Department of Biochemistry I, University of Göttingen, 37073, Göttingen, Germany; STED Microscopy of Synaptic Function, European Neuroscience Institute, 37077 Göttingen, Germany; DFG Research Center Molecular Physiology of the Brain and Excellence Cluster 171, Göttingen, Germany |
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Abstract: | SNARE proteins mediate membrane fusion in the secretory pathway of eukaryotic cells. Genetic deletion and siRNA-based knockdown have been instrumental in assigning given SNAREs to defined intracellular transport steps. However, SNARE depletion occasionally results in barely detectable phenotypes. To understand how cells cope with SNARE loss, we have knocked down several SNAREs functioning in early endosome fusion. Surprisingly, knockdown of syntaxin 13, syntaxin 6 and vti1a, alone or in combinations, did not result in measurable changes of endosomal trafficking or fusion. We found that the residual SNARE levels (typically ∼10%) were sufficient for a substantial amount of SNARE–SNARE interactions. Conversely, in wild-type cells, most SNARE molecules were concentrated in clusters, constituting a spare pool not readily available for interactions. Additionally, the knockdown organelles exhibited enhanced docking. We conclude that SNAREs are expressed at much higher levels than needed for maintenance of organelle fusion, and that loss of SNAREs is compensated for by the co-regulation of the docking machinery. |
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Keywords: | docking endosomes fusion genetic depletion SNARE complex SNARE |
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