Article

Ca2+-regulated pool of phosphatidylinositol-3-phosphate produced by phosphatidylinositol 3-kinase C2α on neurosecretory vesicles

Molecular Dynamics of Synaptic Function Laboratory, Queensland Brain Institute and School of Biomedical Sciences, The University of Queensland, St. Lucia, 4072 Queensland, Australia.
Molecular biology of the cell (Impact Factor: 5.98). 11/2008; 19(12):5593-603. DOI: 10.1091/mbc.E08-06-0595
Source: PubMed

ABSTRACT Phosphatidylinositol-3-phosphate [PtdIns(3)P] is a key player in early endosomal trafficking and is mainly produced by class III phosphatidylinositol 3-kinase (PI3K). In neurosecretory cells, class II PI3K-C2alpha and its lipid product PtdIns(3)P have recently been shown to play a critical role during neuroexocytosis, suggesting that two distinct pools of PtdIns(3)P might coexist in these cells. However, the precise characterization of this additional pool of PtdIns(3)P remains to be established. Using a selective PtdIns(3)P probe, we have identified a novel PtdIns(3)P-positive pool localized on secretory vesicles, sensitive to PI3K-C2alpha knockdown and relatively resistant to wortmannin treatment. In neurosecretory cells, stimulation of exocytosis promoted a transient albeit large increase in PtdIns(3)P production localized on secretory vesicles sensitive to PI3K-C2alpha knockdown and expression of PI3K-C2alpha catalytically inactive mutant. Using purified chromaffin granules, we found that PtdIns(3)P production is controlled by Ca(2+). We confirmed that PtdIns(3)P production from recombinantly expressed PI3K-C2alpha is indeed regulated by Ca(2+). We provide evidence that a dynamic pool of PtdIns(3)P synthesized by PI3K-C2alpha occurs on secretory vesicles in neurosecretory cells, demonstrating that the activity of a member of the PI3K family is regulated by Ca(2+) in vitro and in living neurosecretory cells.

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    • "Interestingly, the ATP-dependent priming stage of exocytosis was originally found to be regulated by Ca 2+ [86]. In this view it is interesting to note that the kinetics of the PtdIns3P increase detected on secretory vesicles closely followed that of the Ca 2+ signal in PC12 cells [85]. Although the functional significance of this PtdIns3P increase is still unknown, it is tempting to speculate that the Ca 2+ signal elicited during the stimulation not only triggers the fusion of LDCVs but also prepares the next round of fusion by promoting the priming of new LDCVs through this and perhaps other pathways. "
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    • "Interestingly, the ATP-dependent priming stage of exocytosis was originally found to be regulated by Ca 2+ [86]. In this view it is interesting to note that the kinetics of the PtdIns3P increase detected on secretory vesicles closely followed that of the Ca 2+ signal in PC12 cells [85]. Although the functional significance of this PtdIns3P increase is still unknown, it is tempting to speculate that the Ca 2+ signal elicited during the stimulation not only triggers the fusion of LDCVs but also prepares the next round of fusion by promoting the priming of new LDCVs through this and perhaps other pathways. "
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    • "This effect was sensitive to PI3K-C2a knockdown. Furthermore, PI3K-C2a activity appears to be directly up regulated by Ca 2? (Wen et al. 2008). These results provide the first direct evidence that a member of the PI3K family can be regulated by Ca 2? signaling. "
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