**6. Future directions**

Another important molecular interactor of STIM1 is the hypoxia-inducible factor-1 alpha (HIF-1α), which is upregulated during hepatocarcinoma growth [85]. Li et al. found that HIF-1α directly controls STIM1 transcription, but also that STIM1-mediated SOCE is required for HIF-1α accumulation in hepatocarcinoma cells via activation of Ca2+/CaM-dependent protein kinase II, revealing a mutual dependence of STIM1 and HIF-1α in the regulation of Ca2+

High levels of ORAI1 and STIM1 are found in many types of cancer cells. In gastric cancer tumor progression, this higher expression is associated with a negative impact on survival rates of patients, an effect that was partially due to targeting expression of metastasis-associated in colon cancer-1 (MACC1) [86], an essential regulator of the transcription for the gene coding for the hepatocyte growth factor receptor, MET. Similarly, a recent report described that STIM1 promotes cell migration and the epithelial-to-mesenchymal transition (EMT) by acti-

Finally, an excellent report from Stephan Feske laboratory [54], described how SOCE is crucial for mitochondrial fatty acid oxidation, and that Ca2+ entry through ORAI1 was essential to activate adenylyl cyclase, cyclic AMP production, the transcriptional regulator peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC-1α) and peroxisome proliferator-activated receptor alpha (PPARα), which is mediated by the activation

Shortly after the molecular description of ORAI1 as the CRAC channel, it was revealed the essential role of other ORAI proteins in cell signaling. The involvement of ORAI3, together with ORAI1, in arachidonic acid-regulated Ca2+ (ARC) channels was early proposed [88, 89]. In contrast to SOC channels, the activation of ARC channels depends on the pool of STIM1 resident in the plasma membrane [90]. More interestingly, ORAI1 and ORAI3 show a differential sensitivity to reactive oxygen species, due to the extracellularly located Cys195 residue which is found in ORAI1, but not in ORAI3. The differential redox sensitivity underlies the differential responses between naïve and T helper lymphocytes, an event that lets T(H) cells

ORAI3, but not ORAI1, was also involved in the activation of PLCδ in response to arachidonic acid, an activation that controls oscillation frequency of Ca2+ spikes triggered by carbachol [92]. ORAI3 channels are overexpressed in estrogen receptor-positive breast cancer cells [93], and it was later demonstrated, using the MCF-7 cancer cell line, that silencing ORAI3 slows down cell cycle and triggers arrest at G1 phase [94]. EGF triggers Ca2+ entry through ORAI3, and the channel is transcriptionally upregulated by the estrogen receptor alpha (ERα) [95]. It is now accepted that cancer cells show a remodeling of ORAI proteins, with an enhanced participation of ORAI3 compared to noncancerous cells, suggesting that heteromerization of

vating TGF-β, Snail and Wnt/β-catenin pathways in prostate cancer cells [87].

transport and tumor growth [85].

12 Calcium and Signal Transduction

**5. ORAI3 and calcium signaling**

proliferate and secrete cytokines in oxidative environments [91].

ORAI3 and ORAI1 is a common feature in malignant transformation [96].

of CREB.

During the past decade, a significant progress was made regarding the molecular description of the proteins involved in store-operated Ca2+ entry. Although some details remain unclear,

**Figure 2.** Activation of SOCE by STIM1/ORAI1 and pathways involved in SOCE-dependent signaling. Panel A: diverse stimuli that triggers the activation of the phospholipase C pathway, such as activation of EGF receptor (EGFR), stimulate the production of inositol 1,4,5-trisphosphate (IP3) which binds and activates IP3 receptor (IP3R) at the endoplasmic reticulum (ER). This activation leads to the release of Ca2+ from the ER, with the subsequent transient depletion of intraluminal [Ca2+] and the activation of STIM1. Ca2+-unbound STIM1 aggregates in oligomers and translocates to plasma membrane (PM)-ER junctions where it binds and activates ORAI1. Extracellular Ca2+ entry through ORAI1 activates multiple Ca2+-dependent targets, as shown in panel B, but also provides a Ca2+ source to replenish intraluminal Ca2+ levels. This replenishment is accomplished by the ER-Ca2+-ATPase which pumps Ca2+ into the ER lumen. Panel B: schematic illustration of the most important pathways regulated by STIM1/ORAI1. AC, adenylyl cyclase; GPCR, G protein-coupled receptor; PM-STIM1, plasma membrane-resident STIM1; p-STIM1, phosphorylated STIM1.

a topology of STIM1-ORAI1 contact sites, selectivity filters in ORAI1, and the role of posttranslational modifications have been reported for both proteins. The involvement of STIM and ORAI proteins in different pathways is now much clearer, and they are now considered master regulators of Ca2+-dependent signaling pathways. However, in many cases pathways were studied in cancer cell lines in vitro, so physiological models are required to evaluate the importance of STIM1 and ORAI1 in the pathophysiology of cells in vivo. Nevertheless, primary cell cultures and established cell lines constitute a widely accepted experimental approach for basic studies in cell signaling and understanding the role of STIM1/ORAI1 in cell biology and cell signaling. With these tools, we have reached the conclusion that STIM1 and ORAI1 are involved in the control of Ca2+ refilling within the ER. More important, STIM1 and ORAI1 directly modulate Ca2+ signaling in a wide variety of pathways, with a significant role in gene expression, cell migration, and tumor cell metastasis (**Figure 2**). Because the expression of STIM1/ORAI1 is deregulated in cancer cells, it is required to evaluate the relative importance of STIM1/ORAI1 as pharmacological targets for the treatment of disease, not only with the use of in vitro cell cultures, but also in animal models for the study of human disease.

[2] Zhang SL, Yu Y, Roos J, Kozak JA, Deerinck TJ, Ellisman MH, Stauderman KA, Cahalan MD. STIM1 is a Ca2+ sensor that activates CRAC channels and migrates from the Ca2+

Regulation of Calcium Signaling by STIM1 and ORAI1 http://dx.doi.org/10.5772/intechopen.78587 15

[3] Stathopulos PB, Li GY, Plevin MJ, Ames JB, Ikura M. Stored Ca2+ depletion-induced oligomerization of stromal interaction molecule 1 (STIM1) via the EF-SAM region: An initiation mechanism for capacitive Ca2+ entry. Journal of Biological Chemistry.

[4] Zheng L, Stathopulos PB, Schindl R, Li GY, Romanin C, Ikura M. Auto-inhibitory role of the EF-SAM domain of STIM proteins in store-operated calcium entry. Proceedings of the National Academy of Sciences of the United States of America. 2011;**108**(4):1337-1342

[5] Huang GN, Zeng W, Kim JY, Yuan JP, Han L, Muallem S, Worley PF. STIM1 carboxylterminus activates native SOC, I(crac) and TRPC1 channels. Nature Cell Biology. 2006;

[6] Grigoriev I, Gouveia SM, van der Vaart B, Demmers J, Smyth JT, Honnappa S, Splinter D, Steinmetz MO, Putney JW Jr, Hoogenraad CC, Akhmanova A. STIM1 is a MT-plus-endtracking protein involved in remodeling of the ER. Current Biology. 2008;**18**(3):177-182 [7] Zeng W, Yuan JP, Kim MS, Choi YJ, Huang GN, Worley PF, Muallem S. STIM1 gates TRPC channels, but not Orai1, by electrostatic interaction. Molecular Cell. 2008;**32**(3):439-448 [8] Zheng L, Stathopulos PB, Li GY, Ikura M. Biophysical characterization of the EF-hand and SAM domain containing Ca2+ sensory region of STIM1 and STIM2. Biochemical and

[9] Brandman O, Liou J, Park WS, Meyer T. STIM2 is a feedback regulator that stabilizes basal cytosolic and endoplasmic reticulum Ca2+ levels. Cell. 2007;**131**(7):1327-1339 [10] Hou X, Pedi L, Diver MM, Long SB. Crystal structure of the calcium release-activated

[11] Yen M, Lokteva LA, Lewis RS. Functional analysis of Orai1 concatemers supports a hexameric stoichiometry for the CRAC channel. Biophysical Journal. 2016;**111**(9):1897-1907

[12] McNally BA, Somasundaram A, Jairaman A, Yamashita M, Prakriya M. The C- and N-terminal STIM1 binding sites on Orai1 are required for both trapping and gating

[13] Muik M, Fahrner M, Schindl R, Stathopulos P, Frischauf I, Derler I, Plenk P, Lackner B, Groschner K, Ikura M, Romanin C. STIM1 couples to ORAI1 via an intramolecular transition into an extended conformation. The EMBO Journal. 2011;**30**(9):1678-1689 [14] Muik M, Fahrner M, Derler I, Schindl R, Bergsmann J, Frischauf I, Groschner K, Romanin C. A cytosolic homomerization and a modulatory domain within STIM1 C terminus determine coupling to ORAI1 channels. The Journal of Biological Chemistry.

[15] Liou J, Kim ML, Heo WD, Jones JT, Myers JW, Ferrell JE, Meyer T. STIM is a Ca2+ sensor essential for Ca2+-store-depletion-triggered Ca2+ influx. Current Biology. 2005;

store to the plasma membrane. Nature. 2005;**437**(7060):902-905

Biophysical Research Communications. 2008;**369**(1):240-246

calcium channel Orai. Science. 2012;**338**(6112):1308-1313

CRAC channels. The Journal of Physiology. 2013;**591**(11):2833-2850

2006;**281**(47):35855-35862

**8**(9):1003-1010

2009;**284**(13):8421-8426

**15**(13):1235-1241
