Orai proteins interact with TRPC channels and confer responsiveness to store depletion

The TRPC (C-type transient receptor potential) class of ion channels has been hypothesized to participate in store-operated Ca²⁺ entry (SOCE). Recently, however, STIM1 and Orai1 proteins have been proposed to form SOCE channels. Whether TRPCs participate in SOCE that is dependent on or regulated by Orai has not been explored. Here we show that Orai1 physically interacts with the N and C termini of TRPC3 and TRPC6, and that in cells overexpressing either TRPC3 or TRPC6 in a store-depletion insensitive manner, these TRPCs become sensitive to store depletion upon expression of an exogenous Orai. Thus, Orai-1, -2, and -3 enhanced thapsigargin-induced calcium entry by 50-150% in cells stably overexpressing either TRPC3 or TRPC6. Orai1 expression had no significant effect on endogenous, thapsigargin-induced calcium entry in wild-type cells (HEK-293, COS1), in HEK cells expressing a thapsigargin-sensitive variant of TRPC3 (TRPC3a), or in HEK cells overexpressing another membrane protein, V1aR. Single-channel cation currents present in membrane patches of TRPC3-overexpressing cells were suppressed by expression of Orai1. We propose that Orai proteins by interacting with TRPCs act as regulatory subunits that confer STIM1-mediated store depletion sensitivity to these channels.