Calcium microdomains at the immunological synapse: how ORAI channels, mitochondria and calcium pumps generate local calcium signals for efficient T-cell activation

Abstract

Cell polarization enables restriction of signalling into microdomains. Polarization of lymphocytes following formation of a mature immunological synapse (IS) is essential for calcium-dependent T-cell activation. Here, we analyse calcium microdomains at the IS with total internal reflection fluorescence microscopy. We find that the subplasmalemmal calcium signal following IS formation is sufficiently low to prevent calcium-dependent inactivation of ORAI channels. This is achieved by localizing mitochondria close to ORAI channels. Furthermore, we find that plasma membrane calcium ATPases (PMCAs) are re-distributed into areas beneath mitochondria, which prevented PMCA up-modulation and decreased calcium export locally. This nano-scale distribution-only induced following IS formation-maximizes the efficiency of calcium influx through ORAI channels while it decreases calcium clearance by PMCA, resulting in a more sustained NFAT activity and subsequent activation of T cells.

Ca2+ homeostasis in the endoplasmic reticulum measured with a new low-Ca2+-affinity targeted aequorin

“…In HeLa cells, the data obtained in non-Ca 2+ We have also studied the effect of mitochondrial membrane depolarization with FCCP on the rate of ER Ca 2+ refilling. Mitochondria have been reported to control the activity of store operated Ca 2+ channels by avoiding their Ca 2+ -inactivation [14,15]. Under this view, mitochondrial membrane depolarization should lead to rapid channel inactivation during refilling and thus, to a significant reduction in the rate of refilling.…”

The mitochondrial dynamics in cancer and immune-surveillance

“…An optimal T cell activation requires Drp1-dependent mitochondrial fragmentation to accumulate mitochondria beneath the TCR clusters on the plasma membrane, during the immune synapse (IS) formation [197]. In addition, Drp1-dependent mitochondria fragmentation is required at the IS for local calcium buffering and for the correct activation of several T cell transcription factors, such as NF-AT and NF-κB, thus driving cytokine production [198][199][200] (Fig. 2C).…”

Mutations of the Ca2+-sensing Stromal Interaction Molecule STIM1 Regulate Ca2+ Influx by Altered Oligomerization of STIM1 and by Destabilization of the Ca2+ Channel Orai1

“…The thereby exposed interaction domain (named either CRAC activation domain (CAD), STIM1 Orai1-activating region (SOAR) or ORAI1-activating small fragment (OASF)) directly interact with and trap Orai1 channels (8 -10), which are initially evenly distributed within the PM (11). The domains of clustered STIM1/Orai1 molecules are the sites of Ca 2ϩ influx (12)(13)(14). The minimal requirement for activation are two STIM1 molecules bound per Orai1 tetramer; however, this stoichiometry results in channels with very low open probability (P o ), whereas maximal P o is achieved with eight bound STIM1 proteins per Orai1 tetramer (15,16).…”