Plasma Membrane Orai1 and Septin Organization during Calcium Signalling

The STIM-ORAI-mediated calcium release-activated calcium channel, or CRAC channel, is a key source of calcium influx that maintains cellular calcium balance and mediates calcium signaling. STIM in the ER membrane senses depletion of ER calcium stores and moves to ER-plasma membrane junctions, then recruits plasma membrane ORAI channels to the junctions and directly gates the channels. We have shown previously that septins are crucial for efficient STIM1-ORAI1 cluster formation following store depletion. Septins are known to specify diffusive barriers in the plasma membrane and to serve as scaffolds to recruit signalling proteins, but their detailed role in calcium signaling remains to be characterized. Here we utilize live-cell super-resolution microscopy and single-molecule tracking to map ORAI1 relative to STIM1 and membrane-localized septins. These observations at high spatial and temporal resolution enable precise analyses of ORAI1 movements before and after store depletion, and of the relation of septins to the membrane nanodomains where STIM and ORAI interact.