The Effects of Ca$^{2+}$ on the Dynamics of PIP$_2$ containing Lipid Bilayers

Changes of intercellular Ca$^{2+}$ concentrations are one of the most ubiquitous signaling events that accompany or precede large scale cellular responses. We are in particular interested in the direct modulation of phosphatidylinositol 4,5-bisphosphate (PIP$_2$) organization in the membrane due to Ca $^{+2}$. At physiological conditions, PIP$_2$'s headgroup is multiply negatively charged (\textgreater\ 3 effective charges) and interacts with the cationic Ca$^{2+}$. By coordinating several PIP$_2$ head-groups, calcium ions can induce condensation and aggregation of PIP$_2$. A series of experiments were conducted on supported lipid bilayers containing physiological quantities of PIP$_2$. Fluorescence correlation spectroscopy (FCS) was used to study the response of the PIP$_2$ to changes in the concentration of Ca$^{2+}$ ions. As Ca$^{2+}$ concentration increases, the FCS indicates that PIP$_2$ goes from a freely diffusing single species to a multiple species system. The diffusion rates of the additional species decrease with increasing [Ca$^{2+}$], thus indicating increasing aggregate sizes with increasing, but physiological relevant Ca$^{2+}$ concentrations. An intriguing effect was observed at very low Ca$^{2+}$ levels. The diffusion rate was consistently measured to increase upon addition of small concentrations of Ca$^{2+}$ before decreasing as the concentrations increased. A series of polymer cushioned bilayers were used to attempt to gain greater insight into the nature of the membrane/support interaction and the nature of this effect.