There is now compelling evidence that subcellular signalling microdomains are crucial for encoding and relaying calcium (Ca2+) signals. The STIM-Orai system constitutes a critical illustration of this concept. Upon depletion of the endoplasmic reticulum (ER), both proteins move to so-called ER-PM junctions, where the plasma membrane (PM) closely apposes the ER membrane with a separation of approximately 15nm. I will present the first three-dimensional model of such ER-PM junctions and will show how the spatial organisation of Orai channels in the PM and Ca2+ pumps in the ER membrane shape the local Ca2+ signature in a non-trivial manner, which has direct consequences for downstream Ca2+ signalling. The coordination of Ca2+ increases in such microdomains can lead to whole cell Ca2+ oscillations. Given their intrinsic stochasticity, we have pursued a statistical analysis of Ca2+ spikes using concepts from stochastic point processes and Bayesian inference. I will show how we can quantify Ca2+ spiking at the single cell level in the presence of dynamic stimulation. This will not only move us closer to characterizing the role of cellular heterogeneity in Ca2+ signalling, but will also highlight how we can use whole-cell Ca2+ signals to infer properties of the subcellular Ca2+ signalling toolkit.
