Figure 2.

All subunits of the Orai1 hexamer contribute equally to channel gating. Whole-cell currents were recorded in the presence of 20 mM extracellular Ca²⁺ (Ca²⁺_o) from HEK293 cells transfected with mCh-STIM1 and 6xOrai1-GFP. (A) Wild-type 6xOrai1 current at −100 mV developed slowly after break-in with 10 mM EGTA in the recording pipette and was blocked by addition of 10 μM La³⁺. (B) Reduction of channel activity by L273D mutations introduced into the indicated subunit (SU) of the hexamer. Channel activity is plotted as the peak CRAC current density at −100 mV (circles, individual cells; lines, mean ± SE; n = 7–11 cells per hexamer variant). (C) Effects of single L273D mutations on the extent of CDI. CDI was evoked by 200-ms steps from the holding potential of +30 mV to the indicated test potentials. The L273D mutation in each of three indicated subunits reduced the extent of CDI relative to the WT hexamer. (D) The extent of CDI plotted as a function of voltage for WT and L273D mutant hexamers, and quantified as current remaining at the end of the step relative to the peak current (see Materials and Methods). The L273D mutation reduced CDI similarly regardless of its location in subunit 1, 3, or 6, across the entire voltage range (mean ± SE, n = 4–6 cells per hexamer variant).