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Epigenomic changes are an important feature of malignant tumors. How tumor aggressiveness is affected by DNA methylation of specific loci is largely unexplored. In genome-wide DNA methylation analyses, we identified the KCa3.1 channel gene (KCNN4) promoter to be hypomethylated in an aggressive non-small-cell lung carcinoma (NSCLC) cell line and in patient samples. Accordingly, KCa3.1 expression was increased in more aggressive NSCLC cells. Both findings were strong predictors for poor prognosis in lung adenocarcinoma. Increased KCa3.1 expression was associated with aggressive features of NSCLC cells. Proliferation and migration of pro-metastatic NSCLC cells depended on KCa3.1 activity. Mechanistically, elevated KCa3.1 expression hyperpolarized the membrane potential, thereby augmenting the driving force for Ca2+ influx. KCa3.1 blockade strongly reduced the growth of xenografted NSCLC cells in mice as measured by positron emission tomography-computed tomography. Thus, loss of DNA methylation of the KCNN4 promoter and increased KCa3.1 channel expression and function are mechanistically linked to poor survival of NSCLC patients. [ABSTRACT FROM AUTHOR]

The endoplasmic reticulum (ER) is involved in many cellular functions, including protein folding and Ca2+ homeostasis. The ability of cells to respond to the ER stress is critical for cell survival, and disruption in such regulation can lead to apoptosis. ER stress is accompanied by alterations in Ca2+ homeostasis, and the ER Ca2+ store depletion by itself can induce ER stress and apoptosis. Despite that, the ER Ca2+ leak channels activated in response to the ER stress remain poorly characterized. Here we demonstrate that ER Ca2+ depletion during the ER stress occurs via translocon, the ER protein complex involved in translation. Numerous ER stress inducers stimulate the ER Ca2+ leak that can be prevented by translocon inhibitor, anisomycin. Expression of GRP78, an ER stress marker, increased following treatment with puromycin (a translocon opener) and was suppressed by anisomycin, confirming a primary role of translocon in ER stress induction. Inhibition of ER store depletion by anisomycin significantly reduces apoptosis stimulated by the ER stress inducers. We suggest that translocon opening is physiologically modulated by GRP78, particularly during the ER stress. The ability to modulate the ER Ca2+ permeability and subsequent ER stress can lead to development of a novel therapeutic approach. [ABSTRACT FROM AUTHOR]

Breast cancer (BC) has a poor prognosis due to its strong metastatic ability. Accumulating data present ether à go-go (hEag1) K+ channels as relevant player in controlling cell cycle and proliferation of non-invasive BC cells. However, the role of hEag1 in invasive BC cells migration is still unknown. In this study, we studied both the functional expression and the involvement in cell migration of hEag1 in the highly metastatic MDA-MB-231 human BC cells. We showed that hEag1 mRNA and proteins were expressed in human invasive ductal carcinoma tissues and BC cell lines. Functional activity of hEag1 channels in MDA-MB-231 cells was confirmed using astemizole, a hEag1 blocker, or siRNA. Blocking or silencing hEag1 depolarized the membrane potential and reduced both Ca2+ entry and MDA-MB-231 cell migration without affecting cell proliferation. Recent studies have reported that Ca2+ entry through Orai1 channels is required for MDA-MB-231 cell migration. Down-regulation of hEag1 or Orai1 reduced Ca2+ influx and cell migration with similar efficiency. Interestingly, no additive effects on Ca2+ influx or cell migration were observed in cells co-transfected with sihEag1 and siOrai1. Finally, both Orai1 and hEag1 are expressed in invasive breast adenocarcinoma tissues and invaded metastatic lymph node samples (LNM+). In conclusion, this study is the first to demonstrate that hEag1 channels are involved in the serum-induced migration of BC cells by controlling the Ca2+ entry through Orai1 channels. hEag1 may therefore represent a potential target for the suppression of BC cell migration, and thus prevention of metastasis development. J. Cell. Physiol. 227: 3837-3846, 2012. © 2012 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

Selective Serotonin Reuptake Inhibitor antidepressants, such as fluoxetine (Prozac), have been shown to induce cell death in cancer cells, paving the way for their potential use as cancer therapy. These compounds are able to increase cytosolic calcium concentration ([Ca2+]cyt), but the involved mechanisms and their physiological consequences are still not well understood. Here, we show that fluoxetine induces an increase in [Ca2+]cyt by emptying the endoplasmic reticulum (ER) through the translocon, an ER Ca2+ leakage structure. Our data also show that fluoxetine inhibits oxygen consumption and lowers mitochondrial ATP. This latter is essential for Ca2+ reuptake into the ER, and we postulated therefore that the fluoxetine-induced decrease in mitochondrial ATP production results in the emptying of the ER, leading to capacitative calcium entry. Furthermore, Ca2+ quickly accumulated in the mitochondria, leading to mitochondrial Ca2+ overload and cell death. We found that fluoxetine could induce an early necrosis in human peripheral blood lymphocytes and Jurkat cells, and could also induce late apoptosis, especially in the tumor cell line. These results shed light on fluoxetine-induced cell death and its potential use in cancer treatment.

Intracellular calcium signals regulate cell function and cell survival by controlling many processes. CD95 engagement results in distinct intracellular calcium signals that control the cell fate, apoptosis, or survival, depending on the ligand (membrane or soluble). Intracellular calcium determination is an exquisite readout to explore the molecular mechanisms elicited by CD95 engagement. The most widely applied methods for studying calcium signaling pathways use fluorescent indicators and imaging methods with fluorescence microscopy. This technical approach, however, requires many precautions that we discuss in this chapter.

Epigenomic changes are an important feature of malignant tumors. How tumor aggressiveness is affected by DNA methylation of specific loci is largely unexplored. In genome-wide DNA methylation analyses, we identified the KCa 3.1 channel gene (KCNN4) promoter to be hypomethylated in an aggressive non-small-cell lung carcinoma (NSCLC) cell line and in patient samples. Accordingly, KCa 3.1 expression was increased in more aggressive NSCLC cells. Both findings were strong predictors for poor prognosis in lung adenocarcinoma. Increased KCa 3.1 expression was associated with aggressive features of NSCLC cells. Proliferation and migration of pro-metastatic NSCLC cells depended on KCa 3.1 activity. Mechanistically, elevated KCa 3.1 expression hyperpolarized the membrane potential, thereby augmenting the driving force for Ca(2+) influx. KCa 3.1 blockade strongly reduced the growth of xenografted NSCLC cells in mice as measured by positron emission tomography-computed tomography. Thus, loss of DNA methylation of the KCNN4 promoter and increased KCa 3.1 channel expression and function are mechanistically linked to poor survival of NSCLC patients., (© 2015 UICC.)