Publications by authors named "Magali Savignac"

26 Publications

  • Page 1 of 1

Ca1.4 calcium channels control cytokine production by human peripheral T17 cells and psoriatic skin-infiltrating T cells.

J Allergy Clin Immunol 2021 Oct 13. Epub 2021 Oct 13.

Toulouse Institute for Infectious and Inflammatory Diseases (Infinity), INSERM UMR1291, Centre National de la Recherche Scientifique UMR5051, University Paul Sabatier Toulouse III, Toulouse, France. Electronic address:

Background: Type-17 inflammation characterizes psoriasis, a chronic skin disease. Because several inflammatory cytokines contribute to psoriasis pathogenesis, inhibiting the simultaneous production of these cytokines in T17 cells may be beneficial in psoriasis. We found that Ca1.4, encoded by CACNA1F, was the only Ca1 calcium channel expressed in T17 cells.

Objective: We sought to investigate the role of Ca1.4 expression in early T17-activation events and effector functions, as well as its association with T17 signature genes in lesional psoriatic (LP) skins.

Methods: Transcriptional gene signatures associated with CACNA1F expression were examined in LP skins by RT-PCR and in situ hybridization. Ca1 inhibitor and/or shRNA lentivectors were used to assess the contribution of Ca1.4 in T17 activation and effector functions in a 3-dimensional skin reconstruction model.

Results: CACNA1F expression correlated with inflammatory cytokine expression that characterizes LP skins and was preferentially associated with RORC expression in CD4 and CD4 cells from LP biopsies. Nicardipine, a Ca1 channel antagonist, markedly reduced inflammatory cytokine production by T17 cells from blood or LP skin. This was associated with decreased TCR-induced early calcium events at cell membrane and proximal signaling events. The knockdown of Ca1.4 in T17 cells impaired cytokine production. Finally, Ca1 inhibition reduced the expression of the keratinocyte genes characteristic of T17-mediated psoriasis inflammation in human skin equivalents.

Conclusions: Ca1.4 channels promote T17-cell functions both at the periphery and in inflammatory psoriatic skin.
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http://dx.doi.org/10.1016/j.jaci.2021.09.030DOI Listing
October 2021

Separation of the Ca 1.2-Ca 1.3 calcium channel duo prevents type 2 allergic airway inflammation.

Allergy 2021 Jun 28. Epub 2021 Jun 28.

Toulouse Institute for Infectious and Inflammatory Diseases (Infinity) INSERM UMR1291, CNRS UMR5051, Université Paul Sabatier Toulouse III, Toulouse, France.

Background: Voltage-gated calcium (Ca 1) channels contribute to T-lymphocyte activation. Ca 1.2 and Ca 1.3 channels are expressed in Th2 cells but their respective roles are unknown, which is investigated herein.

Methods: We generated mice deleted for Ca 1.2 in T cells or Ca 1.3 and analyzed TCR-driven signaling. In this line, we developed original fast calcium imaging to measure early elementary calcium events (ECE). We also tested the impact of Ca 1.2 or Ca 1.3 deletion in models of type 2 airway inflammation. Finally, we checked whether the expression of both Ca 1.2 and Ca 1.3 in T cells from asthmatic children correlates with Th2-cytokine expression.

Results: We demonstrated non-redundant and synergistic functions of Ca 1.2 and Ca 1.3 in Th2 cells. Indeed, the deficiency of only one channel in Th2 cells triggers TCR-driven hyporesponsiveness with weakened tyrosine phosphorylation profile, a strong decrease in initial ECE and subsequent reduction in the global calcium response. Moreover, Ca 1.3 has a particular role in calcium homeostasis. In accordance with the singular roles of Ca 1.2 and Ca 1.3 in Th2 cells, deficiency in either one of these channels was sufficient to inhibit cardinal features of type 2 airway inflammation. Furthermore, Ca 1.2 and Ca 1.3 must be co-expressed within the same CD4 T cell to trigger allergic airway inflammation. Accordingly with the concerted roles of Ca 1.2 and Ca 1.3, the expression of both channels by activated CD4 T cells from asthmatic children was associated with increased Th2-cytokine transcription.

Conclusions: Thus, Ca 1.2 and Ca 1.3 act as a duo, and targeting only one of these channels would be efficient in allergy treatment.
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http://dx.doi.org/10.1111/all.14993DOI Listing
June 2021

Involvement of ion channels in allergy.

Curr Opin Immunol 2018 06 25;52:60-67. Epub 2018 Apr 25.

Center of Physiopathology Toulouse Purpan, University Paul Sabatier Toulouse III, INSERM U1043, CNRS UMR 5282, 31024 Toulouse, France.

Allergic asthma is a complex disease, often characterized by an inappropriate Th2 response to normally harmless allergens. Epithelial cells damaged or activated by the allergen produce IL-33, TSLP and IL-25, activating ILC2 and dendritic cells. The latter migrate into lymph nodes where they induce Th2-cell commitment. Th2 and other type 2 innate inflammatory cells trigger inflammation and airway hyper-reactivity. The toolbox consisting of the ion channels varies from one cellular type to another and depends on its activation state, offering the possibility to design novel drugs in the field of allergy. We will discuss about some channels as calcium, nonselective cation, potassium and chloride channels that appear as good candidates in allergy.
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http://dx.doi.org/10.1016/j.coi.2018.04.006DOI Listing
June 2018

The β and α2δ auxiliary subunits of voltage-gated calcium channel 1 (Ca1) are required for T2 lymphocyte function and acute allergic airway inflammation.

J Allergy Clin Immunol 2018 09 10;142(3):892-903.e8. Epub 2017 Nov 10.

Center of Physiopathology Toulouse Purpan, University Paul Sabatier Toulouse III, INSERM U1043, CNRS UMR 5282, Toulouse, France. Electronic address:

Background: T lymphocytes express not only cell membrane ORAI calcium release-activated calcium modulator 1 but also voltage-gated calcium channel (Ca) 1 channels. In excitable cells these channels are composed of the ion-forming pore α1 and auxiliary subunits (β and α2δ) needed for proper trafficking and activation of the channel. Previously, we disclosed the role of Ca1.2 α1 in mouse and human T2 but not T1 cell functions and showed that knocking down Ca1 α1 prevents experimental asthma.

Objective: We investigated the role of β and α2δ auxiliary subunits on Ca1 α1 function in T2 lymphocytes and on the development of acute allergic airway inflammation.

Methods: We used Caβ antisense oligonucleotides to knock down Caβ and gabapentin, a drug that binds to and inhibits α2δ1 and α2δ2, to test their effects on T2 functions and their capacity to reduce allergic airway inflammation.

Results: Mouse and human T2 cells express mainly Caβ1, β3, and α2δ2 subunits. Caβ antisense reduces T-cell receptor-driven calcium responses and cytokine production by mouse and human T2 cells with no effect on T1 cells. Caβ is mainly involved in restraining Ca1.2 α1 degradation through the proteasome because a proteasome inhibitor partially restores the α1 protein level. Gabapentin impairs the T-cell receptor-driven calcium response and cytokine production associated with the loss of α2δ2 protein in T2 cells.

Conclusions: These results stress the role of Caβ and α2δ2 auxiliary subunits in the stability and activation of Ca1.2 channels in T2 lymphocytes both in vitro and in vivo, as demonstrated by the beneficial effect of Caβ antisense and gabapentin in allergic airway inflammation.
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http://dx.doi.org/10.1016/j.jaci.2017.09.045DOI Listing
September 2018

Androgen signaling negatively controls group 2 innate lymphoid cells.

J Exp Med 2017 06 8;214(6):1581-1592. Epub 2017 May 8.

Centre de Physiopathologie de Toulouse Purpan (CPTP), Université de Toulouse, Institut National de la Santé et de la Recherche Medicale (INSERM), Centre National de la Recherche Scientifique (CNRS), UPS, 31300 Toulouse, France

Prevalence of asthma is higher in women than in men, but the mechanisms underlying this sex bias are unknown. Group 2 innate lymphoid cells (ILC2s) are key regulators of type 2 inflammatory responses. Here, we show that ILC2 development is greatly influenced by male sex hormones. Male mice have reduced numbers of ILC2 progenitors (ILC2Ps) and mature ILC2s in peripheral tissues compared with females. In consequence, males exhibit reduced susceptibility to allergic airway inflammation in response to environmental allergens and less severe IL-33-driven lung inflammation, correlating with an impaired expansion of lung ILC2s. Importantly, orchiectomy, but not ovariectomy, abolishes the sex differences in ILC2 development and restores IL-33-mediated lung inflammation. ILC2Ps express the androgen receptor (AR), and AR signaling inhibits their differentiation into mature ILC2s. Finally, we show that hematopoietic AR expression limits IL-33-driven lung inflammation through a cell-intrinsic inhibition of ILC2 expansion. Thus, androgens play a crucial protective role in type 2 airway inflammation by negatively regulating ILC2 homeostasis, thereby limiting their capacity to expand locally in response to IL-33.
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http://dx.doi.org/10.1084/jem.20161807DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5461006PMC
June 2017

SERCA2 dysfunction in Darier disease causes endoplasmic reticulum stress and impaired cell-to-cell adhesion strength: rescue by Miglustat.

J Invest Dermatol 2014 Jul 3;134(7):1961-1970. Epub 2014 Jan 3.

INSERM UMR 1163, Paris, France; Université Paris Descartes-Sorbonne Paris Cité, Paris, France; Institut Imagine, Paris, France; Department of Genetics, Necker Hospital, F-75015, France. Electronic address:

Darier disease (DD) is a severe dominant genetic skin disorder characterized by the loss of cell-to-cell adhesion and abnormal keratinization. The defective gene, ATP2A2, encodes sarco/endoplasmic reticulum (ER) Ca2+ -ATPase isoform 2 (SERCA2), a Ca2+ -ATPase pump of the ER. Here we show that Darier keratinocytes (DKs) display biochemical and morphological hallmarks of constitutive ER stress with increased sensitivity to ER stressors. Desmosome and adherens junctions (AJs) displayed features of immature adhesion complexes: expression of desmosomal cadherins (desmoglein 3 (Dsg3) and desmocollin 3 (Dsc3)) and desmoplakin was impaired at the plasma membrane, as well as E-cadherin, β-, α-, and p120-catenin staining. Dsg3, Dsc3, and E-cadherin showed perinuclear staining and co-immunostaining with ER markers, indicative of ER retention. Consistent with these abnormalities, intercellular adhesion strength was reduced as shown by a dispase mechanical dissociation assay. Exposure of normal keratinocytes to the SERCA2 inhibitor thapsigargin recapitulated these abnormalities, supporting the role of loss of SERCA2 function in impaired desmosome and AJ formation. Remarkably, treatment of DKs with the orphan drug Miglustat, a pharmacological chaperone, restored mature AJ and desmosome formation, and improved adhesion strength. These results point to an important contribution of ER stress in DD pathogenesis and provide the basis for future clinical evaluation of Miglustat in Darier patients.
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http://dx.doi.org/10.1038/jid.2014.8DOI Listing
July 2014

DREAM controls the on/off switch of specific activity-dependent transcription pathways.

Mol Cell Biol 2014 Mar 23;34(5):877-87. Epub 2013 Dec 23.

National Center of Biotechnology, CSIC and CIBERNED, Madrid, Spain.

Changes in nuclear Ca(2+) homeostasis activate specific gene expression programs and are central to the acquisition and storage of information in the brain. DREAM (downstream regulatory element antagonist modulator), also known as calsenilin/KChIP-3 (K(+) channel interacting protein 3), is a Ca(2+)-binding protein that binds DNA and represses transcription in a Ca(2+)-dependent manner. To study the function of DREAM in the brain, we used transgenic mice expressing a Ca(2+)-insensitive/CREB-independent dominant active mutant DREAM (daDREAM). Using genome-wide analysis, we show that DREAM regulates the expression of specific activity-dependent transcription factors in the hippocampus, including Npas4, Nr4a1, Mef2c, JunB, and c-Fos. Furthermore, DREAM regulates its own expression, establishing an autoinhibitory feedback loop to terminate activity-dependent transcription. Ablation of DREAM does not modify activity-dependent transcription because of gene compensation by the other KChIP family members. The expression of daDREAM in the forebrain resulted in a complex phenotype characterized by loss of recurrent inhibition and enhanced long-term potentiation (LTP) in the dentate gyrus and impaired learning and memory. Our results indicate that DREAM is a major master switch transcription factor that regulates the on/off status of specific activity-dependent gene expression programs that control synaptic plasticity, learning, and memory.
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http://dx.doi.org/10.1128/MCB.00360-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4023831PMC
March 2014

Protein kinase C-dependent activation of CaV1.2 channels selectively controls human TH2-lymphocyte functions.

J Allergy Clin Immunol 2014 Apr 22;133(4):1175-83. Epub 2013 Dec 22.

INSERM U1043, CNRS U5282, University Paul Sabatier, Center of Physiopathology from Toulouse Purpan, Toulouse, France. Electronic address:

Background: In addition to calcium release-activated calcium channel/ORAI calcium channels, the role of voltage-gated calcium (Cav1) channels in T-cell calcium signaling is emerging. Cav1 channels are formed by α1 (CaV1.1 to CaV1.4) and auxiliary subunits. We previously demonstrated that mouse TH2 cells selectively overexpressed CaV1.2 and CaV1.3 channels. Knocking down these channels with Cav1 antisense (AS) oligonucleotides inhibited TH2 functions and experimental asthma.

Objective: We investigated the expression profile and role of Cav1 channels in human T-cell subsets, with a focus on TH2 cells.

Methods: We compared the profile of CaV1 channel subunit expression in T-cell subsets isolated ex vivo from the blood of healthy donors, as well as in vitro-polarized T-cell subsets, and tested the effect of the Cav1 inhibitors nicardipine and Cav1.2AS on their functions.

Results: CaV1.4 expression was detectable in CD4(+) T cells, ex vivo TH1 cells, and TH17 cells, whereas Cav1.2 channels predominated in TH2 cells only. T-cell activation resulted in Cav1.4 downregulation, whereas Cav1.2 expression was selectively maintained in polarized TH2 cells and absent in TH1 or TH9 cells. Nicardipine and CaV1.2AS decreased Ca(2+) and cytokine responses in TH2, but not TH1, cells. Protein kinase C (PKC) α/β inhibition decreased Ca(2+) and cytokine responses, whereas both calcium and cytokine responses induced by PKC activation were inhibited by nicardipine or Cav1.2AS in TH2 cells.

Conclusion: This study highlights the selective expression of Cav1.2 channels in human TH2 cells and the role of PKC-dependent Cav1.2 channel activation in TH2 cell function. Blocking PKC or Cav1.2 channel activation in TH2 cells might represent new strategies to treat allergic diseases in human subjects.
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http://dx.doi.org/10.1016/j.jaci.2013.10.038DOI Listing
April 2014

Singularities of calcium signaling in effector T-lymphocytes.

Biochim Biophys Acta 2013 Jul 20;1833(7):1595-602. Epub 2012 Dec 20.

INSERM U1043, Toulouse, F-31024, France.

CD4(+) helper T (Th) lymphocytes orchestrate the immune response and include several types of effectors such as Th1, Th17 and Th2 cells. They fight against intracellular, extracellular pathogens and parasites respectively. They may also cause distinct immunopathological disorders. Th1 and Th17 are implicated in the development of autoimmune diseases while Th2 cells can initiate allergic diseases. These subsets differ by their TCR-associated signaling. In addition, the regulation of intracellular calcium concentration is not the same in Th1, Th2 and 17 cells. Our group showed that Th2 cells selectively overexpressed voltage-activated calcium (Cav1)-related channels. An increasing number of groups report the presence of Cav1-related products in T-lymphocyte subsets. This is a matter of debate since these calcium channels are classically defined as activated by high cell membrane depolarization in excitable cells. However, the use of mice with ablation of some Cav1 subunits shows undoubtedly an immune phenotype raising the question of how Cav1 channels are regulated in lymphocytes. We showed that knocking down Cav1.2 and/or Cav1.3 subunits impairs the functions of Th2 lymphocytes and is beneficial in experimental models of asthma, while it has no effect on Th1 cell functions. Beyond the role of Cav1 channels in T-lymphocytes, the identification of key components selectively implicated in one or the other T cell subset paves the way for the design of new selective therapeutic targets in the treatment of immune disorders while preserving the other T-cell subsets. This article is part of a Special Issue entitled: 12th European Symposium on Calcium.
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http://dx.doi.org/10.1016/j.bbamcr.2012.12.001DOI Listing
July 2013

[Calcium signaling in T lymphocytes].

Med Sci (Paris) 2012 Aug-Sep;28(8-9):773-9. Epub 2012 Aug 22.

Inserm U1043, centre de physiopathologie de Toulouse Purpan, place du Docteur Baylac, BP 3028, 31024 Toulouse Cedex 3, France.

Calcium signaling is essential for all the functions of T lymphocytes, including those of Th2 cells. Th2 lymphocytes producing interleukins 4, 5 and 13 orchestrate allergic diseases including asthma. T-cell activation induces an influx of Ca(2+) from the external medium through ORAI calcium channels although other calcium channels are likely to be involved. Among them, voltage-gated calcium (Ca(v)1) channels have been reported in some T-cell subsets including Th2 cells. The inhibition of Ca(v)1 channels abrogates T-cell receptor-driven calcium influx and interleukin production by Th2 cells. From a therapeutic point of view, the inhibition of Ca(v)1 channels prevents Th2-dependent experimental allergic asthma. In this review, we will discuss the singularities of calcium responses depending upon the T-cell subset and its state of activation.
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http://dx.doi.org/10.1051/medsci/2012288020DOI Listing
October 2012

Darier disease : a disease model of impaired calcium homeostasis in the skin.

Biochim Biophys Acta 2011 May 15;1813(5):1111-7. Epub 2010 Dec 15.

INSERM, U563, Toulouse, F-31300, France.

The importance of extracellular calcium in epidermal differentiation and intra-epidermal cohesion has been recognized for many years. Darier disease (DD) was the first genetic skin disease caused by abnormal epidermal calcium homeostasis to be identified. DD is characterized by loss of cell-to-cell adhesion and abnormal keratinization. DD is caused by genetic defects in ATP2A2 encoding the sarco/endoplasmic reticulum Ca(2+)-ATPase isoform 2 (SERCA2). SERCA2 is a calcium pump of the endoplasmic reticulum (ER) transporting Ca(2+) from the cytosol to the lumen of ER. ATP2A2 mutations lead to loss of Ca(2+) transport by SERCA2 resulting in decreased ER Ca(2+) concentration in Darier keratinocytes. Here, we review the role of SERCA2 pumps and calcium in normal epidermis, and we discuss the consequences of ATP2A2 mutations on Ca(2+) signaling in DD. This article is part of a Special Issue entitled: 11th European Symposium on Calcium.
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http://dx.doi.org/10.1016/j.bbamcr.2010.12.006DOI Listing
May 2011

Increased B cell proliferation and reduced Ig production in DREAM transgenic mice.

J Immunol 2010 Dec 8;185(12):7527-36. Epub 2010 Nov 8.

Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas, Madrid, Spain.

DREAM/KChIP-3 is a calcium-dependent transcriptional repressor highly expressed in immune cells. Transgenic mice expressing a dominant active DREAM mutant show reduced serum Ig levels. In vitro assays show that reduced Ig secretion is an intrinsic defect of transgenic B cells that occurs without impairment in plasma cell differentiation, class switch recombination, or Ig transcription. Surprisingly, transgenic B cells show an accelerated entry in cell division. Transcriptomic analysis of transgenic B cells revealed that hyperproliferative B cell response could be correlated with a reduced expression of Klf9, a cell-cycle regulator. Pulse-chase experiments demonstrated that the defect in Ig production is associated with reduced translation rather than with increased protein degradation. Importantly, transgenic B cells showed reduced expression of the Eif4g3 gene, which encodes a protein related to protein translation. Our results disclose, to our knowledge, a novel function of DREAM in proliferation and Ig synthesis in B lymphocytes.
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http://dx.doi.org/10.4049/jimmunol.1000152DOI Listing
December 2010

Knocking down Cav1 calcium channels implicated in Th2 cell activation prevents experimental asthma.

Am J Respir Crit Care Med 2010 Jun 18;181(12):1310-7. Epub 2010 Feb 18.

INSERM U563, Centre de Physiopathologie de Toulouse Purpan, Place du Dr Baylac, Toulouse Cedex 3, France.

Rationale: Th2 cells orchestrate allergic asthma and the cytokines they produce (IL-4, IL-5, and IL-13) are deleterious in allergy. Therefore, it is important to identify key signaling molecules expressed by Th2 cells that are essential for their function. We have previously shown that dihydropyridines selectively modulate Th2 cell functions.

Objectives: Because dihydropyridines bind to and modulate voltage-dependent calcium (Ca(v)1) channel in excitable cells, we aimed at showing that Th2 cells selectively express functional Ca(v)1-related channels, the inhibition of which may prevent asthma.

Methods: We looked for Ca(v)1 channel expression in Th2 and Th1 cells by real-time polymerase chain reaction and Western blotting. We sequenced the isoforms expressed by Th2 cells and tested whether Ca(v)1 antisense oligodeoxynucleotides (Ca(v)1AS) affected Ca(2+) signaling and cytokine production. Finally, we tested the effect of Ca(v)1AS in the passive asthma model by injection of ovalbumin-specific Th2 cells transfected with Ca(v)1AS into BALB/c mice challenged with intranasal ovalbumin and in the active model of asthma by intranasal delivery of Ca(v)1AS together with soluble ovalbumin in BALB/c mice previously immunized with ovalbumin in alum.

Measurements And Main Results: We show that mouse Th2 but not Th1 cells expressed Ca(v)1.2 and Ca(v)1.3 channels. Th2 cells transfected with Ca(v)1AS had impaired Ca(2+) signaling and cytokine production, and lost their ability to induce airway inflammation on adoptive transfer. Furthermore, intranasal administration of Ca(v)1AS suppressed airway inflammation and hyperreactivity in an active model of asthma.

Conclusions: These results indicate that Th2 cells selectively express Ca(v)1 channels that may be efficiently targeted in T lymphocytes to prevent experimental asthma.
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http://dx.doi.org/10.1164/rccm.200907-1166OCDOI Listing
June 2010

Ca2+-operated transcriptional networks: molecular mechanisms and in vivo models.

Physiol Rev 2008 Apr;88(2):421-49

Centro Nacional de Biotecnología, Consejo Superior Investigaciones Científicas, Madrid, Spain.

Calcium is the most universal signal used by living organisms to convey information to many different cellular processes. In this review we present well-known and recently identified proteins that sense and decode the calcium signal and are key elements in the nucleus to regulate the activity of various transcriptional networks. When possible, the review also presents in vivo models in which the genes encoding these calcium sensors-transducers have been modified, to emphasize the critical role of these Ca(2+)-operated mechanisms in many physiological functions.
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http://dx.doi.org/10.1152/physrev.00041.2005DOI Listing
April 2008

Calcium channel blocker prevents T helper type 2 cell-mediated airway inflammation.

Am J Respir Crit Care Med 2007 Jun 8;175(11):1117-24. Epub 2007 Mar 8.

INSERM, U563, CHU Purpan, Place du Dr Baylac, 31024 Toulouse Cedex 3, France.

Rationale: Ca(2+) signaling controls the production of T helper (Th) type 2 cytokines known to be deleterious in asthma. Recently, we showed that Ca(2+) signaling was dihydropyridine (DHP)-sensitive in Th2 lymphocytes and that the DHP derivate, nicardipine, used in the treatment of cardiovascular pathologies, prevents Th2-dependent B cell polyclonal activation.

Objectives: We tested the effect of nicardipine in experimental allergic asthma.

Methods: BALB/c mice immunized with ovalbumin (OVA) in alum and challenged with intranasal OVA were treated with nicardipine once the Th2 response, or even airway inflammation, was induced. We also tested the effect of nicardipine in asthma induced by transferring OVA-specific Th2 cells in BALB/c mice exposed to intranasal OVA. We checked the impact of nicardipine on T-cell responses and airway inflammation.

Measurements And Main Results: Nicardipine inhibited in vitro Ca(2+) response in Th2 cells. In vivo, it impeded the development of Th2-mediated airway inflammation and reduced the capacity of lymphocytes from lung-draining lymph nodes to secrete Th2, but not Th1, cytokines. Nicardipine did not affect antigen presentation to CD4(+) T lymphocytes, nor the initial localization of Th2 cells into the lungs of mice exposed to intranasal OVA; however, it reduced the production of type 2 cytokines and the amplification of the Th2 response in mice with asthma. Conversely, nicardipine had no effect on Th1-mediated airway inflammation.

Conclusions: Nicardipine improves experimental asthma by impairing Th2-dependent inflammation. This study could provide a rationale for developing drugs selectively targeting DHP receptors of Th2 lymphocytes, potentially beneficial in the treatment of asthma.
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http://dx.doi.org/10.1164/rccm.200607-1026OCDOI Listing
June 2007

Calcium-dependent transcription of cytokine genes in T lymphocytes.

Pflugers Arch 2007 Jul 2;454(4):523-33. Epub 2007 Mar 2.

Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Cientificas, Madrid, Spain.

The increase in intracellular calcium ion concentration is a general signaling mechanism used in many biological systems. In T lymphocytes, calcium is essential for activation, differentiation, and effector functions. In this study, we will summarize recent developments of how intracellular calcium concentrations are modified in T cells to affect the activity of three major calcium-dependent transcriptional effectors, i.e., NFAT, MEF2, and DREAM, involved in cytokine gene expression.
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http://dx.doi.org/10.1007/s00424-007-0238-yDOI Listing
July 2007

[A role for voltage-dependent related Ca2+ channels in calcium signaling of T lymphocytes].

Med Sci (Paris) 2007 Feb;23(2):136-8

Inserm U563, CPTP IFR30, CHU Purpan, 1, place du Docteur Baylac, 31059 Toulouse Cedex 9, France.

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http://dx.doi.org/10.1051/medsci/2007232136DOI Listing
February 2007

G protein-coupled receptor kinase 2-mediated phosphorylation of downstream regulatory element antagonist modulator regulates membrane trafficking of Kv4.2 potassium channel.

J Biol Chem 2007 Jan 13;282(2):1205-15. Epub 2006 Nov 13.

Departamento de Biología Molecular and Centro de Biología Molecular Severo Ochoa Universidad Autonoma de Madrid-Consejo Superior de Investigaciones Científicas, Campus de Cantoblanco, 28049 Madrid, Spain.

Downstream regulatory element antagonist modulator (DREAM)/potassium channel interacting protein (KChIP3) is a multifunctional protein of the neuronal calcium sensor subfamily of Ca2+-binding proteins with specific roles in different cell compartments. In the nucleus, DREAM acts as a Ca2+-dependent transcriptional repressor, and outside the nucleus DREAM interacts with Kv4 potassium channels, regulating their trafficking to the cell membrane and their gating properties. In this study we characterized the interaction of DREAM with GRK6 and GRK2, members of the G protein-coupled receptor kinase family of proteins, and their phosphorylation of DREAM. Ser-95 was identified as the site phosphorylated by GRK2. This phosphorylation did not modify the repressor activity of DREAM. Mutation of Ser-95 to aspartic acid, however, blocked DREAM-mediated membrane expression of the Kv4.2 potassium channel without affecting channel tetramerization. Treatment with the calcineurin inhibitors FK506 and cyclosporin A also blocked DREAM-mediated Kv4.2 channel trafficking and calcineurin de-phosphorylated GRK2-phosphorylated DREAM in vitro. Our results indicate that these two Ca2+-dependent posttranslational events regulate the activity of DREAM on Kv4.2 channel function.
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http://dx.doi.org/10.1074/jbc.M607166200DOI Listing
January 2007

The cGMP/protein kinase G pathway contributes to dihydropyridine-sensitive calcium response and cytokine production in TH2 lymphocytes.

J Biol Chem 2006 May 13;281(18):12421-7. Epub 2006 Mar 13.

INSERM, 1U563, Centre de Physiopathologie de Toulouse Purpan, F-31024 Toulouse Cedex 3, France.

Th2 lymphocytes differ from other CD4+ T lymphocytes not only by their effector tasks but also by their T cell receptor (TCR)-dependent signaling pathways. We previously showed that dihydropyridine receptors (DHPR) involved in TCR-induced calcium inflow were selectively expressed in Th2 cells. In this report, we studied whether cGMP-dependent protein kinase G (PKG) activation was implicated in the regulation of DHPR-dependent calcium response and cytokine production in Th2 lymphocytes. The contribution of cGMP in Th2 signaling was supported by the following results: 1) TCR activation elicited cGMP production, which triggered calcium increase responsible for nuclear factor of activated T cell translocation and Il4 gene expression; 2) guanylate cyclase activation by nitric oxide donors increased intracellular cGMP concentration and induced calcium inflow and IL-4 production; 3) reciprocally, guanylate cyclase inhibition reduced calcium response and Th2 cytokine production associated with TCR activation. In addition, DHPR blockade abolished cGMP-induced [Ca2+]i increase, indicating that TCR-induced DHP-sensitive calcium inflow is dependent on cGMP in Th2 cells. Th2 lymphocytes from PKG1-deficient mice displayed impaired calcium signaling and IL-4 production, as did wild-type Th2 cells treated with PKG inhibitors. Altogether, our data indicate that, in Th2 cells, cGMP is produced upon TCR engagement and activates PKG, which controls DHP-sensitive calcium inflow and Th2 cytokine production.
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http://dx.doi.org/10.1074/jbc.M510653200DOI Listing
May 2006

Downstream regulatory element antagonist modulator regulates Ca2+ homeostasis and viability in cerebellar neurons.

J Neurosci 2005 Nov;25(47):10822-30

Department of Molecular and Cellular Biology, National Centre of Biotechnology, Consejo Superior de Investigaciones Científicas, E-28049 Madrid, Spain.

The Na+/Ca2+ exchangers NCX1, NCX2, and NCX3 are vital for the control of cellular Ca2+ homeostasis. Here, we show that a doublet of downstream regulatory element sites in the promoter of the NCX3 gene mediates transcriptional repression of NCX3 by the Ca2+-modulated transcriptional repressor downstream regulatory element antagonist modulator (DREAM). Overexpression of a DREAM EF-hand mutant insensitive to Ca2+ (EFmDREAM) in hippocampus and cerebellum of transgenic mice significantly reduced NCX3 mRNA and protein levels without modifying NCX1 and NCX2 expression. Cerebellar granules from EFmDREAM transgenic mice showed increased levels of cytosolic Ca2+ and were more vulnerable to increased Ca2+ influx after partial opening of voltage-gated plasma membrane Ca2+ channels induced by increasing K+ in the culture medium but survived better in the conditions of reduced Ca2+ influx prevailing in low extracellular K+. Overexpression of NCX3 in EFmDREAM transgenic granules using a lentiviral vector restored the normal survival response to high K+ observed in wild-type granules. Thus, the downregulation of the regulator of Ca2+ homeostasis NCX3 by Ca2+-regulated DREAM is a striking example of the autoregulatory property of the Ca2+ signal in neurons.
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http://dx.doi.org/10.1523/JNEUROSCI.3912-05.2005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6725879PMC
November 2005

Transcriptional repressor DREAM regulates T-lymphocyte proliferation and cytokine gene expression.

EMBO J 2005 Oct 22;24(20):3555-64. Epub 2005 Sep 22.

Departamento de Biología Molecular y Celular, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Cientificas, Madrid, Spain.

Downstream Regulatory Element Antagonist Modulator (DREAM) is a Ca2+-dependent transcriptional repressor expressed in the brain, thyroid gland and thymus. Here, we analyzed the function of DREAM and the related protein KChIP-2 in the immune system using transgenic (tg) mice expressing a cross-dominant active mutant (EFmDREAM) for DREAM and KChIPs Ca2+-dependent transcriptional derepression. EFmDREAM tg mice showed reduced T-cell proliferation. Tg T cells exhibited decreased interleukin (IL)-2, -4 and interferon (IFN)gamma production after polyclonal activation and following antigen-specific response. Chromatin immunoprecipitation and transfection assays showed that DREAM binds to and represses transcription from these cytokine promoters. Importantly, specific transient knockdown of DREAM or KChIP-2 induced basal expression of IL-2 and IFNgamma in wild-type splenocytes. These data propose DREAM and KChIP-2 as Ca2+-dependent repressors of the immune response.
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http://dx.doi.org/10.1038/sj.emboj.7600810DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1276700PMC
October 2005

The repressor DREAM acts as a transcriptional activator on Vitamin D and retinoic acid response elements.

Nucleic Acids Res 2005 22;33(7):2269-79. Epub 2005 Apr 22.

Instituto de Investigaciones Biomédicas Alberto Sols, Consejo Superior de Investigaciones Científicas and Universidad Autónoma de Madrid Arturo Duperier 4, 28029 Madrid, Spain.

DREAM (downstream regulatory element antagonist modulator) is a transcriptional repressor, which binds DREs (downstream response elements) in a Ca2+-regulated manner. The DREs consist of core GTCA motifs, very similar to binding motifs for non-steroid nuclear receptors. In this work, we find that DREAM stimulates basal and ligand-dependent activation of promoters containing vitamin D and retinoic acid response elements (VDREs and RAREs), consisting of direct repeats of the sequence AGT/GTCA spaced by 3 or 5 nt, respectively. Stimulation occurs when the element is located upstream, but not downstream, the transcription initiation site. Activation requires both Ca2+ binding to the EF-hands and the leucine-charged domains (LCDs), analogous to those responsible for the interaction of the nuclear receptors with coregulators. Further more, DREAM can bind both 'in vitro' and in chromatin immunoprecipitation assays to these elements. Importantly, 'in vivo' binding is only observed in vitamin D- or RA-treated cells. These results show that DREAM can function as an activator of transcription on certain promoters and demonstrate a novel role for DREAM acting as a potential modulator of genes containing binding sites for nuclear receptors.
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http://dx.doi.org/10.1093/nar/gki503DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1084319PMC
May 2005

Lymphocyte calcium signaling involves dihydropyridine-sensitive L-type calcium channels: facts and controversies.

Crit Rev Immunol 2004 ;24(6):425-47

INSERM U563, CHU Purpan, IFR 30, Place du Dr Baylac, 31024 Toulouse Cedex 3, France.

Calcium influx into lymphocytes is essential for activation, differentiation, and effector functions. While several channel- and receptor-types contribute to calcium influx, voltage-gated calcium channels (VGCC) mediate a well-characterized calcium influx pathway that is most exclusively identified in excitable cells. The role of L-type VGCCs, which belong to high-voltage activated calcium channels and are defined as dihydropyridine (DHP) receptors in excitable cells, is well documented. Interestingly, while lymphocytes do not range in the excitable cell category, the modulatory role of DHP agonists and antagonists and the identification of L-type VGCC-related molecules in B and T lymphocytes, mainly in Th2 cells, suggest these proteins are involved in the calcium response of these cells. Because the identity and the regulation of DHP receptors/channels in lymphocytes is far from being solved, we will discuss the challenging issues of demonstrating a role of L-type VGCCs in nonexcitable cells and the arguments supporting their role in lymphocytes. We will comment on the limitation of the use of DHP agonists and antagonists to ascertain a specific involvement of L-type VGCCs in lymphocyte calcium signaling. Finally, we will provide new clues on the interest of a potential use of DHP antagonists in Th2-cell-mediated pathology.
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http://dx.doi.org/10.1615/critrevimmunol.v24.i6.30DOI Listing
July 2005

Day-night changes in downstream regulatory element antagonist modulator/potassium channel interacting protein activity contribute to circadian gene expression in pineal gland.

J Neurosci 2004 Jun;24(23):5346-55

Departamento Biologia Molecular y Celular, Centro Nacional de Biotecnologia, Consejo Superior de Investigaciones Cientificas, 28049 Madrid, Spain.

The molecular mechanisms controlling the oscillatory synthesis of melatonin in rat pineal gland involve the rhythmic expression of several genes including arylalkylamine N-acetyltransferase (AA-NAT), inducible cAMP early repressor (ICER), and Fos-related antigen-2 (fra-2). Here we show that the calcium sensors downstream regulatory element antagonist modulator/potassium channel interacting protein (DREAM/KChIP)-3 and KChIP-1, -2 and -4 bind to downstream regulatory element (DRE) sites located in the regulatory regions of these genes and repress basal and induced transcription from ICER, fra-2 or AA-NAT promoters. Importantly, we demonstrate that the endogenous binding activity to DRE sites shows day-night oscillations in rat pineal gland and retina but not in the cerebellum. The peak of DRE binding activity occurs during the day period of the circadian cycle, coinciding with the lowest levels of fra-2, ICER, and AA-NAT transcripts. We show that a rapid clearance of DRE binding activity during the entry in the night period is related to changes at the posttranscriptional level of DREAM/KChIP. The circadian pattern of DREAM/KChIP activity is maintained under constant darkness, indicating that an endogenous clock controls DREAM/KChIP function. Our data suggest involvement of the family of DREAM repressors in the regulation of rhythmically expressed genes engaged in circadian rhythms.
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http://dx.doi.org/10.1523/JNEUROSCI.1460-04.2004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6729300PMC
June 2004

Dihydropyridine receptors are selective markers of Th2 cells and can be targeted to prevent Th2-dependent immunopathological disorders.

J Immunol 2004 May;172(9):5206-12

Institut National de la Santé et de la Recherche Médicale Unité 563, Institut Federatif de Recherche 30, Hôpital Purpan, Toulouse, France.

Th1 cells that produce IFN-gamma are essential in the elimination of intracellular pathogens, and Th2 cells that synthetize IL-4 control the eradication of helminths. However, highly polarized Th1 or Th2 responses may be harmful and even lethal. Thus, the development of strategies to selectively down-modulate Th1 or Th2 responses is of therapeutic importance. Herein, we demonstrate that dihydropyridine receptors (DHPR) are expressed on Th2 and not on Th1 murine cells. By using selective agonists and antagonists of DHPR, we show that DHPR are involved in TCR-dependent calcium response in Th2 cells as well as in IL-4, IL-5, and IL-10 synthesis. Nicardipine, an inhibitor of DHPR, is beneficial in experimental models of Th2-dependent pathologies in rats. It strongly inhibits the Th2-mediated autoimmune glomerulonephritis induced by injecting Brown Norway (BN) rats with heavy metals. This drug also prevents the chronic graft vs host reaction induced by injecting CD4(+) T cells from BN rats into (LEW x BN)F(1) hybrids. By contrast, treatment with nicardipine has no effect on the Th1-dependent experimental autoimmune encephalomyelitis triggered in LEW rats immunized with myelin. These data indicate that 1) DHPR are a selective marker of Th2 cells, 2) these calcium channels contribute to calcium signaling in Th2 cells, and 3) blockers of these channels are beneficial in the treatment of Th2-mediated pathologies.
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http://dx.doi.org/10.4049/jimmunol.172.9.5206DOI Listing
May 2004

[Role of L-type calcium channels in the calcium response and interleukin 4 (IL-4) synthesis by Th2 lymphocytes]].

J Soc Biol 2003 ;197(3):259-65

INSERM U563, CHU Purpan, Place du Dr Baylac, Toulouse 31049.

CD4+ T lymphocytes are divided in Th1 cells that produce interferon (IFN) gamma and Th2 cells that synthesize IL-4. These subsets may arise from a common precursor: a combination of IL-12 plus anti-IL-4 monoclonal antibody (mAb) drives Th1 cell differentiation while IL-4 plus anti-IFN gamma mAb favor Th2 cell development. TCR stimulation activates protein kinase C that controls a calcium entry through L type calcium channels in Th2 cells. L type calcium channels are induced during Th2 but not Th1 cell differentiation. In addition, L type calcium channel inhibitors may be successfully used in the treatment of an experimental model of Th2 cell-mediated immunopathology. Thus, this signaling pathway that characterizes Th2 cells can be a target for the treatment of Th2 diseases.
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March 2004
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