Phosphoinositides Alter Lipid Bilayer Properties

Abstract

Phosphoinositides are involved in cell-signaling pathways that regulate vital cell functions such as membrane excitability and trafficking, and cell metabolism, motility and proliferation. At the plasma membrane, phosphatidylinositol-4,5-bisphosphate (PIP2), which constitutes approximately 0.25% of cell phospholipid, is a key messenger in membrane-delimited signaling. PIP2 regulates structurally and functionally diverse membrane proteins including voltage- and ligand-gated ion channels, inwardly rectifying ion channels, transporters and receptors. The mechanism(s) by which PIP2 regulates many of its various “receptors” remain to be elucidated. Here we explore the notion that the amphiphilic phosphoinositides, by adsorbing to the bilayer/solution interface, alter bilayer properties such as curvature and elasticity. Such changes in bilayer properties can alter the equilibrium between membrane protein conformational states and thereby alter function. Taking advantage of the gramicidin channels’ sensitivity to changes in the lipid bilayer properties, we used fluorescence-based and single-channel gA assays to examine the effects of (diC8) phosphoinositides -PI, PI(4,5)P2, PI(3,5)P2, PI(3,4)P2 PI(3,4,5)P3 as well as long-chain PI(4,5)P2 on the lipid bilayer. The diC8 phosphoinositides, except for PI(3,5)P2, alter lipid bilayer properties with potency that decreases with increasing charge. Among the long-chain PI(4,5)P2s, the naturally occurring 1-stearyl-2-arachidonyl-PI(4,5)P2 is a more potent bilayer modifier than di-oleoyl-PI(4,5)P2. The diC8 and the naturally occurring PI(4,5)P2 have similar effects on short and long gA channels, indicating that changes in bilayer curvature dominate over those on bilayer elasticity. In contrast, diC8PI, which was more bilayer-active than diC8PIP2 altered bilayer elasticity. Our results show that application of exogenous PIP2 and its structural analogues (with changes in acyl chain length or phosphorylation state) alters lipid bilayer properties. These PIP2 lipid bilayer effects may be important for some of the many different effects on membrane protein function.