STIM proteins promote a change in the IP3R mode of Ca ²⁺ release. Analysis of global and puff Ca²⁺ release in the indicated MDA-MB-231 (WT, n = 15; S1KO, n = 16; S2KO, n = 12; dKO, n = 15) cells or LM2-4 (WT, n = 14; S1KO, n = 8; S2KO, n = 11; dKO, n = 12) cells. (A and H) Representative traces of Cal-520 fluorescence ratios (ΔF/F0) recorded from the center of an individual puff site before and after ci-IP3 uncaging by a pulse (1 s) of 405-nm light in the indicated MDA-MB-231 (A) or LM2-4 (H) cells. (B, C, I, and J) Boxplot analysis shows the number of Ca²⁺ puffs (B and I) or puff sites (C and J) per cell for the indicated type of MDA-MB-231 (B and C) or LM2-4 (I and J) cells. (D and K) Average global Ca²⁺ responses recorded after UV-induced IP3 uncaging for the indicated type of MDA-MB-231 (D) or LM2-4 (K) cells. (E and L) Time course of Ca²⁺ release via Ca²⁺ puff in the indicated MDA-MB-231 (E) or LM2-4 (L) cells was quantified by multiplying the number of Ca²⁺ puffs by the average puff amplitude within 6-s time intervals. Note that in S1KO or S2KO cells, the rise in cell-wide Ca²⁺ (D and K) occurs, while localized Ca²⁺ release decreases (E and L). (F, G, M, and N) Amplitude distributions of local Ca²⁺ puffs in the indicated MDA-MB-231 (F) or LM2-4 (M). (G and N) Curves show the normalized Gaussian fit to the data shown in F or M for the indicated type of cells. Statistics: one-way ANOVA with Tukey’s post hoc test; *P < 0.05.