Eccrine sweat glands are essential for sweating and thermoregulation in humans. Loss-of-function mutations in the Ca2+ release-activated Ca2+ (CRAC) channel genes ORAI1 and STIM1 abolish store-operated Ca2+ entry (SOCE), and patients with these CRAC channel mutations suffer from anhidrosis and hyperthermia at high ambient temperatures. Here we have shown that CRAC channel-deficient patients and mice with ectodermal tissue-specific deletion of Orai1 (Orai1K14Cre) or Stim1 and Stim2 (Stim1/2K14Cre) failed to sweat despite normal sweat gland development. SOCE was absent in agonist-stimulated sweat glands from Orai1K14Cre and Stim1/2K14Cre mice and human sweat gland cells lacking ORAI1 or STIM1 expression. In Orai1K14Cre mice, abolishment of SOCE was associated with impaired chloride secretion by primary murine sweat glands. In human sweat gland cells, SOCE mediated by ORAI1 was necessary for agonist-induced chloride secretion and activation of the Ca2+-activated chloride channel (CaCC) anoctamin 1 (ANO1, also known as TMEM16A). By contrast, expression of TMEM16A, the water channel aquaporin 5 (AQP5), and other regulators of sweat gland function was normal in the absence of SOCE. Our findings demonstrate that Ca2+ influx via store-operated CRAC channels is essential for CaCC activation, chloride secretion, and sweat production in humans and mice.

Store-operated Ca 2+ entry (SOCE) is a universal Ca 2+ influx pathway that is important for the function of many cell types. SOCE occurs upon depletion of endoplasmic reticulum (ER) Ca 2+ stores and relies on a complex molecular interplay between the plasma membrane (PM) Ca 2+ channel ORAI1 and the ER Ca 2+ sensor stromal interaction molecule (STIM) 1. Patients with null mutations in ORAI1 or STIM1 genes present with severe combined immunodeficiency (SCID)-like disease. Here, we describe the molecular mechanisms by which a loss-of-function STIM1 mutation (R429C) in human patients abolishes SOCE. R429 is located in the third coiled-coil (CC3) domain of the cytoplasmic C terminus of STIM1. Mutation of R429 destabilizes the CC3 structure and alters the conformation of the STIM1 C terminus, thereby releasing a polybasic domain that promotes STIM1 recruitment to ER-PM junctions. However, the mutation also impairs cytoplasmic STIM1 oligomerization and abolishes STIM1-ORAI1 interactions. Thus, despite its constitutive localization at ER-PM junctions, mutant STIM1 fails to activate SOCE. Our results demonstrate multifunctional roles of the CC3 domain in regulating intra-and intermolecular STIM1 interactions that control (i) transition of STIM1 from a quiescent to an active conformational state, (ii) cytoplasmic STIM1 oligomerization, and (iii) STIM1-ORAI1 binding required for ORAI1 activation.release-activated Ca 2+ (CRAC) channels in the plasma membrane (PM) that are formed by multimers of ORAI proteins, which constitute the permeation pore of the channel (1-4). The opening of ORAI channels is mediated by stromal interaction molecule (STIM) 1 and STIM2, single pass transmembrane proteins whose N and C termini are located in the endoplasmic reticulum (ER) lumen and cytoplasm, respectively (5, 6). STIM1 and STIM2 are activated after depletion of ER Ca 2+ stores in response to ligation of cell surface receptors that mediate phospholipase C (PLC) activation. The importance of SOCE is emphasized by null and loss-of-function mutations in ORAI1 and STIM1 genes, which cause a disease syndrome called CRAC channelopathy that is characterized by immunodeficiency, autoimmunity, ectodermal dysplasia, and skeletal myopathy (7). SOCE is a highly choreographed process that involves a complex conformational rearrangement of STIM1 proteins, their oligomerization in the ER lumen and in the cytoplasm, and subsequent translocation from the bulk ER to ER-PM junctions (4, 8). There, STIM1 oligomers form puncta and bind ORAI1. SOCE is initiated by dissociation of Ca 2+ from a paired EF-hand (EFh) domain in the ER luminal N terminus of STIM1 after store depletion (Fig. 1A) and association of STIM1 N termini. These events result in a conformational extension of the cytoplasmic

Store-operated Ca2+ entry (SOCE) through Ca2+ release-activated Ca2+ (CRAC) channels is essential for immunity to infection. CRAC channels are formed by ORAI1 proteins in the plasma membrane and activated by stromal interaction molecules 1 (STIM1) and STIM2 in the endoplasmic reticulum (ER). Mutations in ORAI1 and STIM1 genes that abolish SOCE cause severe immunodeficiency with recurrent infections due to impaired T cell function. SOCE has also been observed in cells of the innate immune system such as macrophages and dendritic cells (DC) and may provide Ca2+ signals required for their function. The specific role of SOCE in macrophage and DC function, and its contribution to innate immunity, however, is not well defined. We found that non-selective inhibition of Ca2+ signaling strongly impairs many effector functions of bone marrow-derived macrophages (BMDMs) and dendritic cells (BMDCs) including phagocytosis, inflammasome activation, and priming of T cells. Surprisingly however, macrophages and DCs from mice with conditional deletion of Stim1 and Stim2 genes – and therefore complete inhibition of SOCE – showed no major functional defects. Their differentiation, FcR-dependent and independent phagocytosis, phagolysosome fusion, cytokine production, NLRP3 inflammasome activation and their ability to present antigens to activate T cells was preserved. Our findings demonstrate that STIM1, STIM2 and SOCE are dispensable for many critical effector functions of macrophages and DCs, which has important implications for CRAC channel inhibition as a therapeutic strategy to suppress pathogenic T cells while not interfering with myeloid cell functions required for innate immunity.

Store-operated Ca(2+) entry (SOCE) is an important Ca(2+) influx pathway, which is defined by the fact that depletion of intracellular Ca(2+) stores, mainly the endoplasmic reticulum (ER), triggers the opening of Ca(2+) channels in the plasma membrane. The best characterized SOC channel is the Ca(2+) release-activated Ca(2+) (CRAC) channel, which was first described in cells of the immune system but has since been reported in many different cell types. CRAC channels are multimers of ORAI family proteins, of which ORAI1 is the best characterized. They are activated by stromal interaction molecules (STIM) 1 and 2, which respond to the depletion of intracellular Ca(2+) stores with oligomerization and binding to ORAI proteins. The resulting SOCE is critical for the physiological function of many cell types including immune cells and platelets. Recent studies using cell lines, animal models and primary cells from human patients with defects in SOCE have highlighted the importance of this Ca(2+) entry mechanism in a variety of pathophysiological processes. This review focuses on the role of SOCE in immunity to infection, allergy, hemostasis and cancer.

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Transient receptor potential (TRP) ion channels are widely expressed in several tissues throughout the mammalian organism. Originally, TRP channel physiology was focusing on its fundamental meaning in sensory neuronal function. Today, it is known that activation of several TRP ion channels in peptidergic neurons does not only result in neuropeptide release and consecutive neurogenic inflammation. Growing evidence demonstrates functional extra-neuronal TRP channel expression in immune and epithelial cells with important implications for mucosal immunology. TRP channels maintain intracellular calcium homeostasis to regulate various functions in the respective cells such as nociception, production and release of inflammatory mediators, phagocytosis, and cell migration. In this review, we provide an overview about TRP-mediated effects in immune and epithelial cells with an emphasis on mucosal immunology of the gut. Crosstalk between neurons, epithelial cells, and immune cells induced by activation of TRP channels orchestrates the immunologic response. Understanding of its molecular mechanisms paves the way to novel clinical approaches for the treatment of various inflammatory disorders including IBD.

The forkhead box transcription factor FOXO3a has recently been identified as central mediator of the cellular response to oxidative stress inducing cell cycle arrest or apoptosis. The aim of our study was to investigate the regulation of FOXO3a in the thyroid and to determine whether alterations in FOXO3a activity occur in thyroid carcinogenesis. In vitro, we demonstrate that FOXO3a activity is negatively regulated by the PI3K/Akt cascade promoting increased phosphorylation and cytoplasmatic accumulation of FOXO3a with decreased transcription of the target genes p27kip (CDKN1B) and Bim (BCL2L11), but increased expression of GADD45A. By contrast, we show that H 2 O 2 exposure activates FOXO3a in thyrocytes with JNK (MAPK8)-mediated nuclear accumulation of FOXO3a and increased expression of the cell cycle arrest genes p27kip and GADD45A. In vivo, we observed a marked cytoplasmatic accumulation of FOXO3a in differentiated thyroid cancers versus an exclusive nuclear accumulation in follicular adenoma and normal thyroid tissue. Moreover, this cytosolic accumulation of FOXO3a correlated with an increased phospho-Akt expression in thyroid malignancies and was accompanied by decreased expression of the FOXO targets p27kip and Bim and an increase in GADD45A mRNA expression in the thyroid cancers. Our data suggest FOXO3a as a novel player of cellular stress response in the thyroid, mediating the thyrocyte's fate either to survive or to undergo apoptosis. Furthermore, PI3K-dependent FOXO3a inactivation may be a novel pathomechanism for the escape from apoptosis in thyroid cancer cells, in particular in follicular thyroid carcinoma.

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