Figure S3.

Effects of SARAF and the STIM1 ID on FCDI and SCDI. (A) Time course of mean ± SEM whole-cell currents in HEK293 cells expressing EYFP-STIM1 and mCherry-Orai1 and infused with 20 mM BAPTA under 20 mM external calcium are shown for the indicated number of SARAF KO (red) or SARAF KO rescue (black) cells. (B) Expression levels of STIM1 and Orai1 are shown for each cell. F, fluorescence. (C) Values of t_1/2 of current activation are shown for each cell. (D and E) Average current densities (D) and time course of the mean ± SEM normalized currents (E) recorded from WT or SARAF KO HEK293 cells expressing Orai1-SS-GFP or coexpressing mCherry-Orai1 and STIM1-EYFP D76A, as indicated. Pipette solution contained 5 mM EGTA. (F) Representative traces showing currents during hyperpolarizing steps (shown on top) from 0 to −80 mV in WT (black) or SARAF KO (red) cells expressing STIM1 and Orai1. (G) The fast and slow τ values of current decay were extracted by double-exponential fitting, and the summary of averaged values is shown. Note that SARAF does not contribute to FCDI. (H) Time course of average whole-cell currents (± SEM) recorded from WT HEK293 cells coexpressing STIM1 and Orai1 with or without SARAF, as indicated. Pipette solution contained 5 mM EGTA and 1 µM TG. (I) The averaged time course (±SEM) of normalized currents recorded from WT HEK293 cells coexpressing Orai1 and STIM1 with or without SARAF, as indicated. Intracellular pipette solution contained 5 mM EGTA and 50 µM IP₃. (J) Time course of normalized currents recorded from individual HEK293 cells expressing Orai1 and WT STIM1 (left) or STIM1 7A (right). Intracellular pipette solution contained either 5 mM EGTA and 50 µM IP₃ or 20 mM BAPTA, as indicated. Note that although currents induced by WT STIM1 undergo slow inactivation, those induced by STIM1 7A do not. All numbers in parentheses or within bars for quantification represent the number of cells.