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Liposomes, Exosomes, and Virosomes: From Modeling Complex
Membrane Processes to Medical Diagnostics and Drug Delivery
Tuesday Speaker Abstracts
23
Plasma Membrane PI(4,5)P2 Is Critical for Secretory Granule Exocytosis
Muhmmad Omar Hmeadi
, Sebastian Barg.Nikhil Gandasi.
Uppsala University, Uppsala, Sweden.
Phosphoinositides, which include PI(4)P (phosphatidylinositol- 4-phosphate), PI(4,5)P2
(phosphatidylinositol- 4,5-bisphosphate) and PI(3,4,5)P3 (phosphatidylinositol-3,4,5-
trisphosphate), are involved in multiple signalling cascades but their role in the regulation of
exocytosis is not fully understood. Recent studies indicate that phosphoinositides, in particular
PI(4,5)P2 and PI(3,4,5)P3, are enriched near docked secretory granules in cell free systems. Here
we have studied phosphoinositide dynamics near secretory granules in live pancreatic β-cells
using TIRF microscopy. Phosphoinositides were detected with EGFP-labelled PH domains of
phospholipase C (PLC) δ1 (high affinity PI(4,5)P2 sensor), PLC δ4 (low affinity PI(4,5)P2
sensor), Grp1 (PI(3,4,5)P3), and EGFP-labelled P4M domains (PI(4)P). Secretory granules were
labelled with NPY-mCherry or NPY-td-Orange2. In live rat insulin secreting (INS1) cells we
found all tested phosphoinositide markers to be evenly distributed across the cell membrane,
without discernible clustering or accumulation at granule docking sites. However, when the cells
were permeabilized with alpha-toxin, the phosphoinositide markers formed a punctate pattern
with partial co-localization with docked secretory granules. In live cells, exposure to insulin
promoted synthesis of PI(3,4,5)P3, but did not influence secretory granule distribution or
exocytosis. Chemically-induced recruitment of a 5’-phosphatase to the plasma membrane
decreased PI(4,5)P2 levels, and resulted in an 85% decrease in secretory granule exocytosis. Our
data indicate that plasma membrane PI(4,5)P2 does not influence secretory granule recruitment
or distribution. However, since PI(4,5)P2 is the predominant phosphoinositide, we hypothesize
that it plays a role in the priming of secretory granules prior to exocytosis.