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Impaired PtdIns(4,5)P2 synthesis in nerve terminals produces defects in synaptic vesicle trafficking

Abstract

Phosphatidylinositol-4,5-bisphosphate (PtdIns(4,5)P2) has an important function in cell regulation both as a precursor of second messenger molecules and by means of its direct interactions with cytosolic and membrane proteins. Biochemical studies have suggested a role for PtdIns(4,5)P2 in clathrin coat dynamics, and defects in its dephosphorylation at the synapse produce an accumulation of coated endocytic intermediates. However, the involvement of PtdIns(4,5)P2 in synaptic vesicle exocytosis remains unclear. Here, we show that decreased levels of PtdIns(4,5)P2 in the brain and an impairment of its depolarization-dependent synthesis in nerve terminals lead to early postnatal lethality and synaptic defects in mice. These include decreased frequency of miniature currents, enhanced synaptic depression, a smaller readily releasable pool of vesicles, delayed endocytosis and slower recycling kinetics. Our results demonstrate a critical role for PtdIns(4,5)P2 synthesis in the regulation of multiple steps of the synaptic vesicle cycle.

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Figure 1: Early postnatal lethality in PI PK1γ-deficient mice.
Figure 2: Alteration of PtdInsP2 metabolism in PIPK1γ-/- mice.
Figure 3: Synaptic transmission in PIPK1γ-/- neurons grown in culture.
Figure 4: Slower rates of endocytosis and recycling in PIPK1γ-/- neurons.
Figure 5: Defects in membrane recycling in PIPK1γ-deficient nerve terminals as revealed by electron microscopy.

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Acknowledgements

We would like to thank L. Liu, L. Lucast, L. Daniell and W. Yan for technical assistance; J. Kunz and R. Jahn for the antibodies; and J. Morgan and O. Cremona for critical reading of the manuscript. This work was supported in part by National Institutes of Health grants to P.D.C, M.W., T.A.R. and R.M.F., and by pilot grants from the Yale Center for Genomic and Proteomics to P.D.C and M.W.

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Correspondence to Pietro De Camilli.

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Supplementary information

Supplementary Figure 1

Absence of PIPK1g in knockout mice. (JPG 43 kb)

Supplementary Figure 2

Glutamate release defects in cortical synaptosomes from PIPK1g heterozygotes. (JPG 41 kb)

Supplementary Figure 3

The recycling pool size is significantly smaller in nerve terminals from PIPK1g KO neurons. (JPG 27 kb)

Supplementary Figure 4

Vesicle reacidification is not delayed in PIPK1g KO nerve terminals. (JPG 41 kb)

Supplementary Legends

Legends for Supplementary Figures 1–4. (DOC 43 kb)

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Paolo, G., Moskowitz, H., Gipson, K. et al. Impaired PtdIns(4,5)P2 synthesis in nerve terminals produces defects in synaptic vesicle trafficking. Nature 431, 415–422 (2004). https://doi.org/10.1038/nature02896

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