Figure 3.

IP₃-induced Ca²⁺ release is suppressed in TRP1-deficient cells. (A) Intact BCR-induced IP₃ production in TRP1-deficient cells. Cells were stimulated with anti-BCR antibody M4 (1 μg/ml) for the indicated time. Data points are the mean ±SE from four experiments. (B) Western blot analysis demonstrating the IP₃R-1 or IP₃R-2 expression indistinguishable in WT and mutant cells using a polyclonal antibody against either IP₃R-1 or IP₃R-2. (C) The ER luminal Ca²⁺ concentration increased with activation of the Ca²⁺ pump, and declined upon application of IP₃. (D) The level of activation of the IP₃R can be quantitatively compared by the initial rate of Ca²⁺ release, which we estimated by fitting an exponential curve (continuous line) to the initial part of the Ca²⁺ decay signal (black circles). (E) IP₃-concentration dependence of Ca²⁺ release. Release rates were obtained by fitting a single exponential to the initial part of Ca²⁺ decay signal measured in luminal Ca²⁺ monitoring (reference 27). The continuous curve and dotted curve represent the best fit hyperbolic equations, r_max/(1 + EC₅₀/[IP₃]), where r_max is the extrapolated values of the maximal rate of Ca²⁺ release, for Ca²⁺ release rates in WT and TRP1-deficient cells, respectively. r_max and EC₅₀ were 0.153 s^-1 and 0.66 μM in WT cells, and 0.118 s⁻¹ and 0.83 μM in mutant cells. Data obtained from TRP1⁻-14 and TRP1⁻-16 were combined. Data points are the mean ±SE from six to seven experiments. **P < 0.01.