Store-Operated Calcium Channels: New Perspectives on Mechanism and Function

Calcium is a remarkably multifunctional signaling ion, at the heart of diverse biological processes that direct the birth, development, function, and eventual death of cells, tissues, and organisms. Cells use a diverse array of transporters and channels to regulate intracellular Ca2+ concentration ([Ca2+]i). A major pathway present in nearly all metazoan cells is the store-operated Ca2+ channel (SOC). The defining feature of SOCs, the one that distinguishes them from all other classes of Ca2+ channels discussed in this volume, is their activation by the reduction of Ca2+ concentration in the lumen of the ER ([Ca2+]ER). Though they were originally described in nonexcitable cells (cells lacking the ability to fire action potentials), they are now known to be present in virtually all cells, including excitable cells like skeletal muscle and neurons (Parekh and Putney 2005). Physiologically, SOCs are most commonly activated by stimuli that release Ca2+ from the ER. This generally involves receptors that activate phospholipase C to produce inositol 1,4,5-trisphosphate (IP3) and activate IP3 receptors in the ER, but can also result from Ca2+-induced Ca2+ release through ER/SR ryanodine receptors. The notion that ER Ca2+ depletion can control Ca2+ entry was first formulated by Jim Putney 25 years ago as the “capacitative calcium entry” hypothesis based on observations that Ca2+ entry triggered by muscarinic agonists was more closely linked to the emptiness of the ER store than to IP3 elevation or occupation of the muscarinic receptor (Putney 1986). The introduction of thapsigargin (TG) (Thastrup et al. 1989), a sarcoendoplasmic reticulum Ca2+-ATPase (SERCA) inhibitor that depletes ER Ca2+ independently of receptors and IP3, and methods for measuring [Ca2+]i in single cells (fura-2, indo-1, etc.) (Grynkiewicz et al. 1985) provided powerful tools that greatly accelerated progress in establishing store-operated Ca2+ entry (SOCE, as it was later renamed) as a ubiquitous Ca2+ entry pathway. TG-induced Ca2+ entry was soon shown to occur in dozens of cell types (Putney and Bird 1993; Parekh and Penner 1997), though nothing was known about the diversity of pathways that might be involved, let alone their molecular basis. A major step forward was the identification of store-operated Ca2+ currents in mast cells and T cells. This achievement arose initially from attempts to identify Ca2+ conductances triggered by secretory agonists in mast cells and antigen receptors in T cells (Penner et al. 1988; Lewis and Cahalan 1989). In both cases, extremely small currents were detected and linked to large [Ca2+]i rises, suggesting a high Ca2+ selectivity. In T cells, the current was shown to activate spontaneously during whole-cell recordings and in response to the T-cell mitogen phytohemagglutinin in perforated-patch recordings (Lewis and Cahalan 1989). Soon after, fluorescence-based studies showed that Ca2+ influx triggered through T cell mitogens shared several features with TG-induced influx, suggesting that T cell receptor agonists activated the store-dependent pathway (Mason et al. 1991; Sarkadi et al. 1991). These two paths of research converged when Hoth and Penner described a highly Ca2+-selective current in mast cells that was activated in whole-cell recordings spontaneously (by Ca2+ chelators), by IP3, and by ionomycin, and called it the Ca2+ release-activated Ca2+ (CRAC) current (Hoth and Penner 1992). In Jurkat T cells, Zweifach and Lewis showed that TG activated a similar current, which appeared to be identical to the mitogen-stimulated current described earlier, and made the first estimate of its characteristically tiny conductance (∼20 femtosiemens, far too small to resolve single-channel currents) (Zweifach and Lewis 1993). These initial studies defined a membrane conductance that over the next decade would be described biophysically and pharmacologically in detail, providing a characteristic “fingerprint” culled from its ion selectivity, unitary conductance, and regulation by intra- and extracellular Ca2+ and pharmacological inhibitors (Parekh and Penner 1997; Prakriya et al. 2004). Among several currents that were described as store-operated in different cells, the CRAC current emerged as the prototype because of its extensive characterization and the weight of evidence showing that it could be activated by ER Ca2+ depletion independently of surface receptors or changes in cytosolic [Ca2+]. This included activation by intracellular Ca2+ chelators, SERCA inhibitors or ionomycin at constant intracellular [Ca2+]i, and by the Ca2+ chelator TPEN loaded into the ER (Prakriya et al. 2004; Parekh and Putney 2005). In fact, the CRAC channel is the only store-operated channel whose input–output relation is known. This relation, first examined by Hofer et al. (1998) and later quantified by Luik et al. (2008) using an ER-targeted cameleon protein, shows that ICRAC is a highly nonlinear function of [Ca2+]ER, with a Hill coefficient of ∼4 and a K1/2 of 170 µM. Given a resting [Ca2+]ER of ∼400 µM, these results suggest that the ER must be depleted by ∼25% before ICRAC begins to activate significantly. Over the two decades after Putney formalized the capacitative Ca2+ entry hypothesis, many mechanisms were proposed as the link between Ca2+ store depletion and SOCE. Among these, diffusible messengers released from the ER, insertion of CRAC channels into the plasma membrane, and conformational coupling of CRAC channels with IP3 receptors in the ER were the most extensively studied, but in the absence of molecular substrates were difficult to establish (Prakriya et al. 2004; Parekh and Putney 2005). The CRAC channel fingerprint proved useful in ruling out a number of candidate genes for the CRAC channel itself, particularly members of the transient receptor potential (TRP) protein family, but its identity remained a mystery (Prakriya et al. 2004; Parekh and Putney 2005). The discoveries of the ER Ca2+ sensor STIM1 in 2005 and the CRAC channel protein Orai1 a year later marked an unmistakable turning point in the field, as they provided the first and most essential molecular tools with which to dissect the SOCE mechanism. The history of these discoveries and the early revelations they afforded have been reviewed extensively (Cahalan et al. 2007; Wu et al. 2007; Fahrner et al. 2009; Putney 2009; Varnai et al. 2009; Hogan et al. 2010). In this review, I will summarize our current understanding of how Ca2+ store depletion leads to Ca2+ entry at a molecular level, and the role of STIM oligomerization and additional proteins in this process. I will also describe how these discoveries and the ensuing studies have increased awareness of SOCE roles in physiology and disease, and have created entirely new directions for research. Throughout I will emphasize work on STIM1 and Orai1 mainly because they have been the most extensively studied isoforms, but will discuss other STIM and Orai isoforms to highlight important functional differences. For more information on these other isoforms, the reader is referred to the reviews cited above.