PtdIns(4,5)P2 is not required for secretory granule docking

Phosphoinositides (PtdIns) play important roles in exocytosis and are thought to regulate secretory granule docking by co‐clustering with the SNARE protein syntaxin to form a docking receptor in the plasma membrane. Here we tested this idea by high‐resolution TIRF imaging of EGFP‐labeled PtdIns markers or syntaxin‐1 at secretory granule release sites in live insulin‐secreting cells. In intact cells, PtdIns markers distributed evenly across the plasma membrane with no preference for granule docking sites. In contrast, syntaxin‐1 was found clustered in the plasma membrane, mostly beneath docked granules. We also observed rapid accumulation of syntaxin‐1 at sites were granules arrived to dock. Acute depletion of plasma membrane PtdIns(4,5)P2 by recruitment of a 5’­phosphatase strongly inhibited Ca2+‐dependent exocytosis, but had no effect on docked granules or the distribution and clustering of syntaxin‐1. Cell permeabilization by α‐toxin or formaldehyde‐fixation caused PtdIns marker to slowly cluster, in part near docked granules. In summary, our data indicate that PtdIns(4,5)P2 accelerates granule priming, but challenge a role of PtdIns in secretory granule docking or clustering of syntaxin‐1 at the release site.

The SNARE protein syntaxin forms small clusters in the plasma membrane that are important for docking and exocytosis of secretory granules. In cell‐free model systems, clustering of syntaxin depends on co‐aggregation with phosphoinosit...

Source: Traffic - March 1, 2018 Category: Research Authors: Muhmmad Omar ‐Hmeadi, Nikhil Gandasi, Sebastian Barg Tags: ORIGINAL ARTICLE Source Type: research