Fig. 1. STIM1/Orai1-mediated Ca²⁺ influx in response to heating.

(a) Maximal fluorescence intensity changes of HEK293T cells transfected with various DNA constructs evoked by the temperature stimulation (n=5, ***p<0.001, one way ANOVA). (b,c) Fura-2 ratiometric single cell calcium imaging of HeLa cells transfected with various DNA constructs (b), and co-transfected with GFP-STIM1 and the indicated siRNAs (c) in response to a heating pulse to ~50°C. 100 µM ATP was applied at the end as a control for cell viability. (d) Temperature responses to a heating pulse to 41°C of Jurkat cells assayed on FLIPR with or without extracellular Ca²⁺. The black dashed line represents fit with a mono-exponential function and the τ value of this fit was given to illustrate the overall kinetics of the Ca²⁺ signal decay of the heat off-response. (e) Temperature-dependent off-responses of Jurkat cells assayed on FLIPR with or without 20 µM Synta1 and fitted with Boltzmann equations. Cells were heated from 25°C to the indicated various temperatures and then cooled back to 25°C for assaying the heat off-response. (data shown as mean ± SEM.)