Physiological Calcium Concentrations Induce PI(4,5)P2 Clustering: PI(4,5)P2 as a Lipidic Calcium Sensor

Abstract

Phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) is a minor component of the plasma membrane of eukaryotic cells that is essential for several cellular mechanisms, including organization and remodeling of the actin cytoskeleton, membrane trafficking, endocytosis and exocytosis. The regulation of these processes is thought to involve localized enrichment of PI(4,5)P2 in the plasma membrane at particular timings. Particularly in calcium-triggered SNARE-dependent exocytosis, PI(4,5)P2 is known to interact and co-segregate with certain synaptic proteins. However, the mechanisms responsible for PI(4,5)P2 clustering in particular sites of the plasma membrane are still not completely understood.Using membrane model systems and fluorescently labeled PI(4,5)P2, we were able to demonstrate that PI(4,5)P2 lateral distribution is sensitive to physiologic calcium concentrations, in the low micromolar range. Calcium concentrations of 0-100 μM were used to mimic neuron intracellular calcium concentrations before and after a stimulus. Fluorescence self-quenching and energy migration data obtained from steady-state and time-resolved fluorescence methods clearly showed that Ca2+ ions promoted the clustering of PI(4,5)P2 molecules in both liquid disordered and liquid ordered membranes. Sensitivity of PI(4,5)P2 clustering to [Ca2+] is shown to be highly dependent on both PI(4,5)P2 density and membrane lipid phase.In this way, the lateral distribution of PI(4,5)P2 presents significant sensitivity to changes in calcium concentration of only a few micromolar, allowing this lipid to function as a lipidic calcium sensor. In the particular case of SNARE-dependent exocytosis, it is expected that localized increases in calcium concentration will dramatically change the pattern of PI(4,5)P2 interaction with synaptic proteins, possibly enhancing fusion rates and promoting exocytosis.Acknowledgements: FCT Portugal is acknowledged for financial support (PTDC/QUI-BIQ/112067/2009; PTDC/QUI-BIQ/099947/2008; SFRH/BPD/64320/2009; SFRH/BD/80575/2011).