Fig. 3.

Comparison of the function of full-length STIM1, STIM1–F394H, and STIM2.1, and the SOAR fragments derived therefrom. (A) I_CRAC measurements in HEK Orai1–CFP cells transiently expressing either STIM1, STIM–F394H, or STIM2.1. Stores were passively depleted with 10 mM BAPTA in the pipette and 20 mM Ca²⁺ bath solution, and 50 μM 2-APB was added as shown. (B) E-FRET was measured in HEK Orai1–CFP cells expressing either YFP–SOAR1, YFP–SOAR1–F394H, or YFP–SOAR2.1. Basal values and values after 50 μM 2-APB addition were obtained and summarized in C. (C) SOAR1 (basal, 0.207 ± 0.014; after 2-APB, 0.242 ± 0.015; n = 42 cells), YFP–SOAR1–F394H (basal, 0.066 ± 0.004; after 2-APB, 0.210 ± 0.007; n = 43 cells), and YFP–SOAR2.1 (basal, 0.032 ± 0.005; after 2-APB, 0.040 ± 0.006; n = 34 cells). Summary data from three separate transfections are shown (means ± SEM). (D) Ca²⁺ imaging in fura2-loaded HEK Orai1–His cells expressing YFP–SOAR1, YFP–SOAR1–F394H, or YFP–SOAR2.1. Constitutive Ca²⁺ entry was measured following addition of 1 mM extracellular Ca²⁺ and 50 μM 2-APB as shown. (E) Fluorescence images showing the distribution of YFP–SOAR1, YFP–SOAR1–F394H, or YFP–SOAR2.1 expressed in HEK Oria1–His cells, either before (Top) or after (Bottom) addition of 50 μM 2-APB. (Scale bar, 10 μm.) Representative traces and images are from three independent experiments; data are means ± SEM.