Regulation of Voltage-Gated Calcium Channel Activity by Palmitoylation: A Fatty Acid Tale

Abstract

The phospholipid phosphatidylinositol 4,5-bisphospate (PIP2) interacts with voltage-gated Ca2+ channels to facilitate their opening. Conversely, inhibition of channel activity following activation of Gq-protein coupled receptors is associated with PIP2 breakdown. PIP2's freed fatty acid tails appear to remain associated with channels, stabilizing closed conformations. My lab is testing a model where an accessory β-subunit CaVβ2a acts as a phospholipid mimic resistant to metabolism; its two palmitoyl groups bind to the channel at a site normally occupied by PIP2's two fatty acid tails. To test this model, we have concentrated on examining N-type Ca2+ channel modulation in a recombinant system where CaV2.2 is coexpressed with α2δ-1 and one of four CaVβ subunits. M1 muscarinic or neurokinin-1 receptor stimulation inhibits N-current from CaVβ1b-, CaVβ3-, or CaVβ4-containing channels, but enhances N-current from CaVβ2a-containing channels. Exogenously applied arachidonic acid produces the same pattern of modulation. Further studies with mutated, chimeric CaVβ subunits and free palmitic acid revealed palmitoylation of CaVβ2a is essential for loss of inhibition. Loss of inhibition appears independent of kinetic changes that occur with different channel complexes. In contrast, channel mutations that reorient CaVβ2a's relative position to CaV2.2 rescue inhibition suggesting that in these experiments, the palmitoyl groups become sufficiently displaced that endogenously released arachidonic acid can once again bind to the inhibitory site. These findings suggest a new dynamic function for palmitoylation and predict that other doubly palmitoylated proteins reach up into the membrane with their lipid fingers to interact with and change the functioning of transmembrane proteins.(Funded by the NIH, AHA and the UMass Medical School).