Endogenous ether lipids differentially promote tumor aggressiveness by regulating the SK3 channel.

SK3 channels are potassium channels found to promote tumor aggressiveness. We have previously demonstrated that SK3 is regulated by synthetic ether lipids, but the role of endogenous ether lipids is unknown. Here, we have studied the role of endogenous alkyl- and alkenyl-ether lipids on SK3 channels and on the biology of cancer cells. Experiments revealed that the suppression of alkylglycerone phosphate synthase or plasmanylethanolamine desaturase 1, which are key enzymes for alkyl- and alkenyl-ether-lipid synthesis, respectively, decreased SK3 expression by increasing micro RNA (miR)-499 and miR-208 expression, leading to a decrease in SK3-dependent calcium entry, cell migration, and matrix metalloproteinase 9-dependent cell adhesion and invasion. We identified several ether lipids that promoted SK3 expression and found a differential role of alkyl- and alkenyl-ether lipids on SK3 activity. The expressions of alkylglycerone phosphate synthase, SK3, and miR were associated in clinical samples emphasizing the clinical consistency of our observations. To our knowledge, this is the first report showing that ether lipids differentially control tumor aggressiveness by regulating an ion channel. This insight provides new possibilities for therapeutic interventions, offering clinicians an opportunity to manipulate ion channel dysfunction by adjusting the composition of ether lipids.

Variable effects of periprostatic adipose tissue on prostate cancer cells: Role of adipose tissue lipid composition and cancer cells related factors.

BACKGROUND: Periprostatic adipose tissue (PPAT) is likely to modulate prostate cancer (PCa) progression. We analyzed the variations in the effect of PPAT on cancer cells, according to its fatty acid (FA) composition and tumor characteristics. METHODS: The expression of markers of aggressiveness Ki67 and Zeb1, and epigenetic marks that could be modified during PCa progression, was analyzed by immunohistochemistry on a tissue-micro-array containing 59 pT3 PCa, including intra-prostatic areas and extra-prostatic foci in contact with PPAT belonging to the same tumor. In addition, we cocultivated PC3 and LNCaP cell lines with PPAT, which were then analyzed for FA composition. RESULTS: Although the contact between PPAT and cancer cells led overall to an increase in Ki67 and Zeb1, and a decrease in the epigenetic marks 5MC, 5HMC, and H3K27ac, these effects were highly heterogeneous. Increased proliferation in extra-prostatic areas was associated with the international society of uropathology score. PC3 and LNCaP cocultures with PPAT led to increased Ki67, Zeb1 and H3K27me3, but only for PPAT associated with aggressive PCa. PC3 proliferation was correlated with high 20.2 n-6 and low 20.5n-3 in PPAT. CONCLUSIONS: These results suggest that the effects of PPAT on cancer cells may depend on both PCa characteristics and PPAT composition, and could lead to propose nutritional supplementation.

Interplay between Prostate Cancer and Adipose Microenvironment: A Complex and Flexible Scenario.

Adipose tissue is part of the prostate cancer (PCa) microenvironment not only in the periprostatic area, but also in the most frequent metastatic sites, such as bone marrow and pelvic lymph nodes. The involvement of periprostatic adipose tissue (PPAT) in the aggressiveness of PCa is strongly suggested by numerous studies. Many molecules play a role in the reciprocal interaction between adipocytes and PCa cells, including adipokines, hormones, lipids, and also lipophilic pollutants stored in adipocytes. The crosstalk has consequences not only on cancer cell growth and metastatic potential, but also on adipocytes. Although most of the molecules released by PPAT are likely to promote tumor growth and the migration of cancer cells, others, such as the adipokine adiponectin and the n-6 or n-3 polyunsaturated fatty acids (PUFAs), have been shown to have anti-tumor properties. The effects of PPAT on PCa cells might therefore depend on the balance between the pro- and anti-tumor components of PPAT. In addition, genetic and environmental factors involved in the risk and/or aggressiveness of PCa, including obesity and diet, are able to modulate the interactions between PPAT and cancer cells and their consequences on the growth and the metastatic potential of PCa.

Curcumin and NCLX inhibitors share anti-tumoral mechanisms in microsatellite-instability-driven colorectal cancer.

BACKGROUND AND AIMS: Recent evidences highlight a role of the mitochondria calcium homeostasis in the development of colorectal cancer (CRC). To overcome treatment resistance, we aimed to evaluate the role of the mitochondrial sodium-calcium-lithium exchanger (NCLX) and its targeting in CRC. We also identified curcumin as a new inhibitor of NCLX. METHODS: We examined whether curcumin and pharmacological compounds induced the inhibition of NCLX-mediated mitochondrial calcium (mtCa2+) extrusion, the role of redox metabolism in this process. We evaluated their anti-tumorigenic activity in vitro and in a xenograft mouse model. We analyzed NCLX expression and associations with survival in The Cancer Genome Atlas (TCGA) dataset and in tissue microarrays from 381 patients with microsatellite instability (MSI)-driven CRC. RESULTS: In vitro, curcumin exerted strong anti-tumoral activity through its action on NCLX with mtCa2+ and reactive oxygen species overload associated with a mitochondrial membrane depolarization, leading to reduced ATP production and apoptosis. NCLX inhibition with pharmacological and molecular approaches reproduced the effects of curcumin. NCLX inhibitors decreased CRC tumor growth in vivo. Both transcriptomic analysis of TCGA dataset and immunohistochemical analysis of tissue microarrays demonstrated that higher NCLX expression was associated with MSI status, and for the first time, NCLX expression was significantly associated with recurrence-free survival. CONCLUSIONS: Our findings highlight a novel anti-tumoral mechanism of curcumin through its action on NCLX and mitochondria calcium overload that could benefit for therapeutic schedule of patients with MSI CRC.

K2P channel C-type gating involves asymmetric selectivity filter order-disorder transitions.

K2P potassium channels regulate cellular excitability using their selectivity filter (C-type) gate. C-type gating mechanisms, best characterized in homotetrameric potassium channels, remain controversial and are attributed to selectivity filter pinching, dilation, or subtle structural changes. The extent to which such mechanisms control C-type gating of innately heterodimeric K2Ps is unknown. Here, combining K2P2.1 (TREK-1) x-ray crystallography in different potassium concentrations, potassium anomalous scattering, molecular dynamics, and electrophysiology, we uncover unprecedented, asymmetric, potassium-dependent conformational changes that underlie K2P C-type gating. These asymmetric order-disorder transitions, enabled by the K2P heterodimeric architecture, encompass pinching and dilation, disrupt the S1 and S2 ion binding sites, require the uniquely long K2P SF2-M4 loop and conserved "M3 glutamate network," and are suppressed by the K2P C-type gate activator ML335. These findings demonstrate that two distinct C-type gating mechanisms can operate in one channel and underscore the SF2-M4 loop as a target for K2P channel modulator development.

The multifaceted role of TMEM16A in cancer.

The calcium-activated chloride channel TMEM16A is intimately linked to cancers. Over decades, TMEM16A over-expression and contribution to prognosis have been widely studied for multiple cancers strengthening the idea that TMEM16A could be a valuable biomarker and a promising therapeutic target. Surprisingly, from the survey of the literature, it appears that TMEM16A has been involved in multiple cancer-related functions and a large number of molecular targets of TMEM16A have been proposed. Thus, TMEM16A appears to be an ion channel with a multifaceted role in cancers. In this review, we summarize the latest development regarding TMEM16A contribution to cancers. We will survey TMEM16A contribution in cancer prognosis, the origins of its over-expression in cancer cells, the multiple biological functions and molecular pathways regulated by TMEM16A. Then, we will consider the question regarding the molecular mechanism of TMEM16A in cancers and the possible basis for the multifaceted role of TMEM16A in cancers.

TMEM16A controls EGF-induced calcium signaling implicated in pancreatic cancer prognosis.

Pancreatic cancer typically spreads rapidly and has poor survival rates. Here, we report that the calcium-activated chloride channel TMEM16A is a biomarker for pancreatic cancer with a poor prognosis. TMEM16A is up-regulated in 75% of cases of pancreatic cancer and high levels of TMEM16A expression are correlated with low patient survival probability. TMEM16A up-regulation is associated with the ligand-dependent EGFR signaling pathway. In vitro, TMEM16A is required for EGF-induced store-operated calcium entry essential for pancreatic cancer cell migration. TMEM16A also has a profound impact on phosphoproteome remodeling upon EGF stimulation. Moreover, molecular actors identified in this TMEM16A-dependent EGFR-induced calcium signaling pathway form a gene set that makes it possible not only to distinguish neuro-endocrine tumors from other forms of pancreatic cancer, but also to subdivide the latter into three clusters with distinct genetic profiles that could reflect their molecular underpinning.

Store-operated Ca2+ entry regulates Ca2+-activated chloride channels and eccrine sweat gland function.

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.

SK3/TRPC1/Orai1 complex regulates SOCE-dependent colon cancer cell migration: a novel opportunity to modulate anti-EGFR mAb action by the alkyl-lipid Ohmline.

BACKGROUND: Barely 10-20% of patients with metastatic colorectal cancer (mCRC) receive a clinical benefit from the use of anti-EGFR monoclonal antibodies (mAbs). We hypothesized that this could depends on their efficiency to reduce Store Operated Calcium Entry (SOCE) that are known to enhance cancer cells. RESULTS: In the present study, we demonstrate that SOCE promotes migration of colon cancer cell following the formation of a lipid raft ion channel complex composed of TRPC1/Orai1 and SK3 channels. Formation of this complex is stimulated by the phosphorylation of the reticular protein STIM1 by EGF and activation of the Akt pathway. Our data show that, in a positive feedback loop SOCE activates both Akt pathway and SK3 channel activity which lead to SOCE amplification. This amplification occurs through the activation of Rac1/Calpain mediated by Akt. We also show that Anti-EGFR mAbs can modulate SOCE and cancer cell migration through the Akt pathway. Interestingly, the alkyl-lipid Ohmline, which we previously showed to be an inhibitor of SK3 channel, can dissociated the lipid raft ion channel complex through decreased phosphorylation of Akt and modulation of mAbs action. CONCLUSIONS: This study demonstrates that the inhibition of the SOCE-dependent colon cancer cell migration trough SK3/TRPC1/Orai1 channel complex by the alkyl-lipid Ohmline may be a novel strategy to modulate Anti-EGFR mAb action in mCRC.

Immature human dendritic cells enhance their migration through KCa3.1 channel activation.

Migration capacity is essential for dendritic cells (DCs) to present antigen to T cells for the induction of immune response. The DC migration is supposed to be a calcium-dependent process, while not fully understood. Here, we report a role of the KCa3.1/IK1/SK4 channels in the migration capacity of both immature (iDC) and mature (mDC) human CD14(+)-derived DCs. KCa3.1 channels were shown to control the membrane potential of human DC and the Ca(2+) entry, which is directly related to migration capacities. The expression of migration marker such as CCR5 and CCR7 was modified in both types of DCs by TRAM-34 (100nM). But, only the migration of iDC was decreased by use of both TRAM-34 and KCa3.1 siRNA. Confocal analyses showed a close localization of CCR5 with KCa3.1 in the steady state of iDC. Finally, the implication of KCa3.1 seems to be limited to the migration capacities as T cell activation of DCs appeared unchanged. Altogether, these results demonstrated that KCa3.1 channels have a pro-migratory effect on iDC migration. Our findings suggest that KCa3.1 in human iDC play a major role in their migration and constitute an attractive target for the cell therapy optimization.

SIGMAR1 Regulates Membrane Electrical Activity in Response to Extracellular Matrix Stimulation to Drive Cancer Cell Invasiveness.

The sigma 1 receptor (Sig1R) is a stress-activated chaperone that regulates ion channels and is associated with pathologic conditions, such as stroke, neurodegenerative diseases, and addiction. Aberrant expression levels of ion channels and Sig1R have been detected in tumors and cancer cells, such as myeloid leukemia and colorectal cancer, but the link between ion channel regulation and Sig1R overexpression during malignancy has not been established. In this study, we found that Sig1R dynamically controls the membrane expression of the human voltage-dependent K(+) channel human ether-à-go-go-related gene (hERG) in myeloid leukemia and colorectal cancer cell lines. Sig1R promoted the formation of hERG/ß1-integrin signaling complexes upon extracellular matrix stimulation, triggering the activation of the PI3K/AKT pathway. Consequently, the presence of Sig1R in cancer cells increased motility and VEGF secretion. In vivo, Sig1R expression enhanced the aggressiveness of tumor cells by potentiating invasion and angiogenesis, leading to poor survival. Collectively, our findings highlight a novel function for Sig1R in mediating cross-talk between cancer cells and their microenvironment, thus driving oncogenesis by shaping cellular electrical activity in response to extracellular signals. Given the involvement of ion channels in promoting several hallmarks of cancer, our study also offers a potential strategy to therapeutically target ion channel function through Sig1R inhibition.

The sigma-1 receptor: a regulator of cancer cell electrical plasticity?

Originally mistaken as an opioid receptor, the sigma-1 receptor (Sig1R) is a ubiquitous membrane protein that has been involved in many cellular processes. While the precise function of Sig1R has long remained mysterious, recent studies have shed light on its role and the molecular mechanisms triggered. Sig1R is in fact a stress-activated chaperone mainly associated with the ER-mitochondria interface that can regulate cell survival through the control of calcium homeostasis. Sig1R functionally regulates ion channels belonging to various molecular families and it has thus been involved in neuronal plasticity and central nervous system diseases. Interestingly, Sig1R is frequently expressed in tumors but its function in cancer has not been yet clarified. In this review, we discuss the current understanding of Sig1R. We suggest herein that Sig1R shapes cancer cell electrical signature upon environmental conditions. Thus, Sig1R may be used as a novel therapeutic target to specifically abrogate pro-invasive functions of ion channels in cancer tissue.

The Orai-1 and STIM-1 complex controls human dendritic cell maturation.

Ca(2+) signaling plays an important role in the function of dendritic cells (DC), the professional antigen presenting cells. Here, we described the role of Calcium released activated (CRAC) channels in the maturation and cytokine secretion of human DC. Recent works identified STIM1 and Orai1 in human T lymphocytes as essential for CRAC channel activation. We investigated Ca(2+) signaling in human DC maturation by imaging intracellular calcium signaling and pharmalogical inhibitors. The DC response to inflammatory mediators or PAMPs (Pathogen-associated molecular patterns) is due to a depletion of intracellular Ca(2+) stores that results in a store-operated Ca(2+) entry (SOCE). This Ca(2+) influx was inhibited by 2-APB and exhibited a Ca(2+)permeability similar to the CRAC (Calcium-Released Activated Calcium), found in T lymphocytes. Depending on the PAMPs used, SOCE profiles and amplitudes appeared different, suggesting the involvement of different CRAC channels. Using siRNAi, we identified the STIM1 and Orai1 protein complex as one of the main pathways for Ca(2+) entry for LPS- and TNF-α-induced maturation in DC. Cytokine secretions also seemed to be SOCE-dependent with profile differences depending on the maturating agents since IL-12 and IL10 secretions appeared highly sensitive to 2-APB whereas IFN-γ was less affected. Altogether, these results clearly demonstrate that human DC maturation and cytokine secretions depend on SOCE signaling involving STIM1 and Orai1 proteins.

The sigma-1 receptor binds to the Nav1.5 voltage-gated Na+ channel with 4-fold symmetry.

The sigma-1 receptor (Sig1R) is up-regulated in many human tumors and plays a role in the control of cancer cell proliferation and invasiveness. At the molecular level, the Sig1R modulates the activity of various ion channels, apparently through a direct interaction. We have previously shown using atomic force microscopy imaging that the Sig1R binds to the trimeric acid-sensing ion channel 1A with 3-fold symmetry. Here, we investigated the interaction between the Sig1R and the Nav1.5 voltage-gated Na(+) channel, which has also been implicated in promoting the invasiveness of cancer cells. We show that the Sig1R and Nav1.5 can be co-isolated from co-transfected cells, consistent with an intimate association between the two proteins. Atomic force microscopy imaging of the co-isolated proteins revealed complexes in which Nav1.5 was decorated by Sig1Rs. Frequency distributions of angles between pairs of bound Sig1Rs had two peaks, at ∼90° and ∼180°, and the 90° peak was about twice the size of the 180° peak. These results demonstrate that the Sig1R binds to Nav1.5 with 4-fold symmetry. Hence, each set of six transmembrane regions in Nav1.5 likely constitutes a Sig1R binding site, suggesting that the Sig1R interacts with the transmembrane regions of its partners. Interestingly, two known Sig1R ligands, haloperidol and (+)-pentazocine, disrupted the Nav1.5/Sig1R interaction both in vitro and in living cells. Finally, we show that endogenously expressed Sig1R and Nav1.5 also functionally interact.

Sig1R protein regulates hERG channel expression through a post-translational mechanism in leukemic cells.

Sig1R (Sigma-1receptor) is a 25-kDa protein structurally unrelated to other mammalian proteins. Sig1R is present in brain, liver, and heart and is overexpressed in cancer cells. Studies using exogenous sigma ligands have shown that Sig1R interacts with a variety of ion channels, but its intrinsic function and mechanism of action remain unclear. The human ether-à-gogo related gene (hERG) encodes a cardiac channel that is also abnormally expressed in many primary human cancers, potentiating tumor progression through the modulation of extracellular matrix adhesive interactions. We show herein that sigma ligands inhibit hERG current density and cell adhesion to fibronectin in K562 myeloid leukemia cells. Heterologous expression in Xenopus oocytes demonstrates that Sig1R potentiates hERG current by stimulating channel subunit biosynthesis. Silencing Sig1R in leukemic K562 cells depresses hERG current density and cell adhesion to fibronectin by reducing hERG membrane expression. In K562 cells, Sig1R silencing does not modify hERG mRNA contents but reduces hERG mature form densities. In HEK cells expressing hERG and Sig1R, both proteins co-immunoprecipitate, demonstrating a physical association. Finally, Sig1R expression enhances both channel protein maturation and stability. Altogether, these results demonstrate for the first time that Sig1R controls ion channel expression through the regulation of subunit trafficking activity.