Figure 5: Simultaneous imaging of STIM1 localization and ER Ca2+ concentration.
From: Imaging intraorganellar Ca2+ at subcellular resolution using CEPIA
(a) The images of GEM-CEPIA1er (upper) and mCherry-STIM1 (middle) at three time points indicated in b as grey dotted lines (T1, T2 and T3). The expanded images of mCherry-STIM1 within the white boxes were shown in the lower panels. Scale bars, 20 μm (upper and middle) and 5 μm (lower). (b) Time courses of ER Ca2+ concentration (blue) and the number of mCherry-STIM1 puncta normalized with the minimum and maximum (magenta). As [Ca2+]ER was depleted with histamine stimulation in the Ca2+-free solution, mCherry-STIM1 formed puncta (T2). After Ca2+ addback in the external solution, [Ca2+]ER gradually recovered and mCherry-STIM1 puncta disappeared (T3). (c) The normalized number of mCherry-STIM1 puncta was plotted against ER Ca2+ concentration during puncta formation (black) and dissociation (magenta). The plots obtained from three independent experiments were overlaid. The relationship between [Ca2+]ER and puncta formation can be fitted by Hill plot with a Hill coefficient of 7.9 and a K1/2 of 350 μM for puncta formation, and 9.7 and 530 μM for puncta dissociation.
