Orai3 and the Selective Activation of the Arc Channel by Arachidonic Acid

Abstract

The store-operated CRAC channels and the store-independent, arachidonic acid-activated ARC channels represent the founding members of a new family of biophysically similar, highly Ca2+-selective, Ca2+ entry channels - the “Orai channels”. Both of these channels are dependent on STIM1 for their activation, but they differ in the pool of STIM1 responsible. Thus, whereas STIM1 in the ER regulates the CRAC channels on store-depletion, ARC channels are exclusively regulated by the pool of STIM1 that constitutively resides in the PM.Recent studies have shown that the functional CRAC channel pore is formed by a tetrameric arrangement of Orai1 units. In contrast, a heteropentameric assembly of three Orai1 subunits and two Orai3 subunits forms the functional ARC channel pore (Mignen et al. J. Physiol. 587: 4181). Importantly, this inclusion of Orai3 subunits in the channel structure has been shown to play a specific, and unique, role in determining the selectivity of the ARC channels for activation by arachidonic acid. Using an approach based on the generation and expression of various concatenated constructs, we examined the basis for this Orai3-dependent effect on selectivity for arachidonic acid. These studies revealed that, whilst heteropentamers containing only one Orai3 subunit are sensitive to arachidonic acid, specific selectivity for activation by this fatty acid is only achieved on inclusion of the second Orai3 subunit in the pentamer. Further studies identified the cytosolic N-terminal domain as the region of the Orai3 molecule that is specifically responsible for this switch in selectivity. Substitution of just this domain into an otherwise complete Orai1 subunit within a concatenated 31111 pentamer is sufficient to change the resulting channel from one that is predominantly store-operated, to one that is essentially exclusively activated by arachidonic acid.