Ca2+ store depletion causes STIM1 to accumulate in ER regions closely associated with the plasma membrane

Abstract

Stromal interacting molecule 1 (STIM1), reported to be an endoplasmic reticulum (ER) Ca2+ sensor controlling store-operated Ca2+ entry, redistributes from a diffuse ER localization into puncta at the cell periphery after store depletion. STIM1 redistribution is proposed to be necessary for Ca2+ release–activated Ca2+ (CRAC) channel activation, but it is unclear whether redistribution is rapid enough to play a causal role. Furthermore, the location of STIM1 puncta is uncertain, with recent reports supporting retention in the ER as well as insertion into the plasma membrane (PM). Using total internal reflection fluorescence (TIRF) microscopy and patch-clamp recording from single Jurkat cells, we show that STIM1 puncta form several seconds before CRAC channels open, supporting a causal role in channel activation. Fluorescence quenching and electron microscopy analysis reveal that puncta correspond to STIM1 accumulation in discrete subregions of junctional ER located 10–25 nm from the PM, without detectable insertion of STIM1 into the PM. Roughly one third of these ER–PM contacts form in response to store depletion. These studies identify an ER structure underlying store-operated Ca2+ entry, whose extreme proximity to the PM may enable STIM1 to interact with CRAC channels or associated proteins.

Thapsigargin activates Ca2+ entry both by store-dependent, STIM1/Orai1-mediated, and store-independent, TRPC3/PLC/PKC-mediated pathways in human endothelial cells

“…Upon Ca 2+ store depletion, STIM1 oligomerizes, translocates close to the plasma membrane and aggregates into punctate structures [2,21,26]. The interaction between STIM1 and Orai1 leads to channel opening and Ca 2+ entry [23,25,27].…”

Dynamic Movement of the Calcium Sensor STIM1 and the Calcium Channel Orai1 in Activated T-Cells: Puncta and Distal Caps

“…Recent work has shown that STIM1 and Orai1 are in membrane contact points between the ER and PM and that movement of a complex containing the two proteins requires at least 10 nm between the two membranes (Varnai et al, 2007). The current consensus is that STIM1 senses the depletion of Ca 2ϩ from ER stores and then moves to sites of ER-PM apposition where interactions between STIM1 and Orai1 cause the opening of CRAC channels composed of Orai1 protein subunits Vig et al, 2006a;Wu et al, 2006;Feske, 2007;Hogan and Rao, 2007;Lewis, 2007;Putney, 2007a;Varnai et al, 2007).We used confocal imaging techniques to visualize the dynamic movement of STIM1 and Orai1 after T-cell activation. We were particularly interested in demonstrating close interactions between fluorescently tagged versions of STIM1 and Orai1 using the technique of Fö rster resonance energy transfer (FRET).…”

“…Some STIM1 resides in the PM in unstimulated cells (Manji et al, 2000;Williams et al, 2002), and externally applied anti-STIM1 antibodies have been reported to inhibit channel activation , but the relationship between PM STIM1 and the STIM1 that moves to puncta after store depletion is not clear. Many studies on STIM1 localization were performed with YFP-tagged fusion proteins that could not insert into the PM because of the YFP moiety (Hauser and Tsien, 2007) yet these fusion proteins were clearly capable of activating CRAC channels (Liou et al, 2005;Baba et al, 2006;Mercer et al, 2006;Soboloff et al, 2006;Wu et al, 2006). Recent work has shown that STIM1 and Orai1 are in membrane contact points between the ER and PM and that movement of a complex containing the two proteins requires at least 10 nm between the two membranes (Varnai et al, 2007).…”

New factors contributing to dynamic calcium regulation in the skeletal muscle triad—a crowded place

“…Stimulating help came from the nonexcitable tissue area, where the molecular players for SOCE were identified as the ER Ca 2+ sensor stromal interaction molecule 1 (Stim1; Roos et al 2005;Wu et al 2006;Liou et al 2005) and the Ca 2+ selective CRAC channel Orai1 (Feske et al 2006). Orai1 was identified as a key regulator of the CRAC channel complex that was mutated in compromised T cells from patients with severe combined immune deficiency (SCID).…”

“…When Ca 2+ dissociates from Stim1 during SR Ca 2+ depletion, a conformational unfolding of the EF and the sterile-α-motif permits aggregation of Stim1 molecules into clusters (Stathopulos et al 2008). In Jurkat cells, ER Ca 2+ depletion was associated with local accumulations of Stim1 clusters as "puncta" near the plasma membrane within tens of seconds that coincided with the development of CRAC currents (Wu et al 2006). This suggested a slow signalling event during which ER Ca 2+ depletion leads to collection and relocation of Stim1 molecules from regions distant from the plasma membrane towards the membrane to interact with the CRAC channels.…”