Ángel Zarain-Herzberg | Universidad Nacional Autónoma de México (original) (raw)

Papers by Ángel Zarain-Herzberg

Canadian Journal of Physiology and Pharmacology, Aug 1, 2012

The precise control of Ca2+levels during the contraction–relaxation cycle in cardiac myocytes is ... more The precise control of Ca2+levels during the contraction–relaxation cycle in cardiac myocytes is extremely important for normal beat-to-beat contractile activity. The sarcoplasmic reticulum (SR) plays a key role controlling calcium concentration in the cytosol. The SR Ca2+-ATPase (SERCA2) transports Ca2+inside the SR lumen during relaxation of the cardiac myocyte. Calsequestrin (Casq2) is the main protein in the SR lumen, functioning as a Ca2+buffer and participating in Ca2+release by interacting with the ryanodine receptor 2 (RyR2) Ca2+-release channel. Alterations in normal Ca2+handling significantly contribute to the contractile dysfunction observed in cardiac hypertrophy and in heart failure. Transcriptional regulation of the SERCA2 gene has been extensively studied and some of the mechanisms regulating its expression have been elucidated. Overexpression of Sp1 factor in cardiac hypertrophy downregulates SERCA2 gene expression and increased levels of thyroid hormone up-regulates its transcription. Other hormones such norepinephrine, angiotensin II, endothelin-1, parathyroid hormone, prostaglandin-F2α, as well the cytokines tumor necrosis factor-α and interleukin-6 also downregulate SERCA2 expression. Calcium acting through the calcineurin–NFAT (nuclear factor of activated T cells) pathway has been suggested to regulate SERCA2 and CASQ2 gene expression. This review focuses on the current knowledge regarding transcriptional regulation of SERCA2 and CASQ2 genes in the normal and pathologic heart.

Journal of Molecular and Cellular Cardiology, Jun 1, 2007

to prevent its precipitation due to high concentrations of free Ca 2+ (2-10 mM). In mammals two C... more to prevent its precipitation due to high concentrations of free Ca 2+ (2-10 mM). In mammals two CASQ isoforms coded by two different genes have been described. The CASQ1 isoform is the only one expressed in fast-twitch skeletal muscle and constitutes the most abundant isoform in slow-twitch skeletal muscle (STSM) (∼ 75%). The CASQ2 isoform is expressed exclusively in cardiac muscle and to a lesser degree on STSM (∼ 25%). In this work we analyzed the transcription of the casq2 gene using several constructs containing the 5′-regulatory region of the human gene. The first 180 bp of promoter region shares high homology among species, which include the putative binding sites MEF-2, E-box and CArG-box for myogenic transcription factors. Functional analyses demonstrated that the proximal 288 bp has a strong transcriptional activity in neonatal rat cardiac myocytes. The data suggest that the elements present in the first 288 bp are sufficient to confer muscular specific expression, and that cis-elements contained in the upstream region (− 1.4 to − 1.7 kb) may act as a cardiac specific enhancer (i.e. MEF-2 and M-CAT). Site-directed mutagenesis of the MEF-2, E-box and CArG-box elements was performed. The MEF-2 and CArG-box, but not E-box mutations were associated with a decrease of the basal transcriptional activity by 70% and 50%, respectively. DNA/protein interaction assays revealed that MEF-2 and SRF bind to the MEF-2 and CArG elements.

The International Journal of Biochemistry & Cell Biology, Aug 1, 2019

Highlights  SERCA3 mRNA expression is decreased in hepatocelullar carcinoma cells (HCC)  NaB an... more Highlights  SERCA3 mRNA expression is decreased in hepatocelullar carcinoma cells (HCC)  NaB and TSA up-regulate SERCA3 mRNA expression in rat HCC  NaB and TSA increase H3K9 and H3K27 acetylation at the ATP2A3 promoter  TSA increases occupancy of p300 at the ATP2A3 promoter  Inhibition of p300 decreases basal and TSA-mediated SERCA3 mRNA expression  p300 mediates H3K9 acetylation at the ATP2A3 promoter

The FASEB Journal, Mar 1, 2006

Cell Calcium, Nov 1, 2020

Calcium (Ca2+) signaling controls a wide range of cellular processes, including the hallmarks of ... more Calcium (Ca2+) signaling controls a wide range of cellular processes, including the hallmarks of cancer. The Ca2+ signaling system encompasses several types of proteins, such as receptors, channels, pumps, exchangers, buffers, and sensors, of which several are mutated or with altered expression in cancer cells. Since epigenetic mechanisms are disrupted in all stages of carcinogenesis, and reversibly regulate gene expression, they have been studied by different research groups to understand their role in Ca2+ signaling remodeling in cancer cells and the carcinogenic process. In this review, we link Ca2+ signaling, cancer, and epigenetics fields to generate a comprehensive landscape of this complex group of diseases.

Cell Calcium, Nov 1, 2014

Cytosolic calcium concentration ([Ca(2+)]c) is fundamental for regulation of many cellular proces... more Cytosolic calcium concentration ([Ca(2+)]c) is fundamental for regulation of many cellular processes such metabolism, proliferation, muscle contraction, cell signaling and insulin secretion. In resting conditions, the sarco/endoplasmic reticulum (ER/SR) Ca(2+) ATPase's (SERCA) transport Ca(2+) from the cytosol to the ER or SR lumen, maintaining the resting [Ca(2+)]c about 25-100nM. A reduced activity and expression of SERCA2 protein have been described in heart failure and diabetic cardiomyopathy, resulting in an altered Ca(2+) handling and cardiac contractility. In the diabetic pancreas, there has been reported reduction in SERCA2b and SERCA3 expression in β-cells, resulting in diminished insulin secretion. Evidence obtained from different diabetes models has suggested a role for advanced glycation end products formation, oxidative stress and increased O-GlcNAcylation in the lowered SERCA2 expression observed in diabetic cardiomyopathy. However, the role of SERCA2 down-regulation in the pathophysiology of diabetes mellitus and diabetic cardiomyopathy is not yet well described. In this review, we make a comprehensive analysis of the current knowledge of the role of the SERCA pumps in the pathophysiology of insulin-dependent diabetes mellitus type 1 (TIDM) and type 2 (T2DM) in the heart and β-cells in the pancreas.

Oncology Letters, May 31, 2018

Period circadian regulator (Per)1 and Per2 genes are involved in the molecular mechanism of the c... more Period circadian regulator (Per)1 and Per2 genes are involved in the molecular mechanism of the circadian clock, and exhibit tumor suppressor properties. Several studies have reported a decreased expression of Per1, Per2 and Per3 genes in different types of cancer and cancer cell lines. Promoter methylation downregulates Per1, Per2 or Per3 expression in myeloid leukemia, breast, lung, and other cancer cells; whereas histone deacetylase inhibitors (HDACi) upregulate Per1 or Per3 expression in certain cancer cell lines. However, the transcriptional regulation of Per1 and Per2 in cancer cells by chromatin modifications is not fully understood. The present study aimed to determine whether HDACi regulate Per1 and Per2 expression in gastric cancer cell lines, and to investigate changes in chromatin modifications in response to HDACi. Treatment of KATO III and NCI-N87 human gastric cancer cells with sodium butyrate (NaB) or Trichostatin A (TSA) induced Per1 and Per2 mRNA expression in a dose-dependent manner. Chromatin immunoprecipitaion assays revealed that NaB and TSA decreased lysine 9 trimethylation on histone H3 (H3K9me3) at the Per1 promoter. TSA, but not NaB increased H3K9 acetylation at the Per2 promoter. It was also observed that binding of Sp1 and Sp3 to the Per1 promoter decreased following NaB treatment, whereas Sp1 binding increased at the Per2 promoter of NaB-and TSA-treated cells. In addition, Per1 promoter is not methylated in KATO III cells, while Per2 promoter was methylated, although NaB, TSA, and 5-Azacytidine do not change the methylated CpGs analyzed. In conclusion, HDACi induce Per1 and Per2 expression, in part, through mechanisms involving chromatin remodeling at the proximal promoter of these genes; however, other indirect mechanisms triggered by these HDACi cannot be ruled out. These findings reveal a previously unappreciated regulatory pathway between silencing of Per1 gene by H3K9me3 and upregulation of Per2 by HDACi in cancer cells.

Molecular Carcinogenesis, Jan 29, 2019

The knowledge about the role of calcium‐regulated pathways in cancer cell growth and differentiat... more The knowledge about the role of calcium‐regulated pathways in cancer cell growth and differentiation could be useful for the development of new therapeutic approaches to diminish its mortality. The ATP2A genes encode for SERCA pumps, which modulate cytosolic Ca2+ concentration, regulating various cellular processes including cell growth. ATP2A3 gene transcriptional down‐regulation has been reported in gastric and colon cancer, but there is still a lack of understanding about the epigenetic processes regulating its transcription. In this work, we report that butyrate, trichostatin A, and 5‐azacytidine treatments increase SERCA3 expression, increased apoptosis, and decreased cell viability of the KATO‐III gastric carcinoma cell line. We analyzed the methylation profile of the ATP2A3 gene promoter CpG island, finding clones with methylated status through −280 to −135 promoter region, harboring Sp1 and AP‐2 binding sites, which could have a role in transcriptional repression. Post‐translational modifications of histones show a major role in the ATP2A3 transcriptional regulation, and our results show histones marks linked to transcriptional repression associated with the −262 to −135 region, this repressive context changed to transcriptional permissive through SERCA3 re‐expressing conditions. These results suggest that the nucleotide sequence from −280 to −135 position is an ATP2A3 epigenetic regulatory CpG region in KATO‐III cells. Analyses of online‐databases show a decreased SERCA3 expression in gastric and colon tumors, as well as overall survival results, showed that high SERCA3 expression could serve as a favorable prognostic marker for colon and gastric cancer patients.

Bioscience Reports, Aug 1, 2008

The NMDA (N-methyl-D-aspartate) receptors are important in the regulation of neuronal development... more The NMDA (N-methyl-D-aspartate) receptors are important in the regulation of neuronal development, synaptic plasticity, learning and memory, and are involved in several brain pathologies. The NR1 subunit is essential for the assembly of functional receptors, as it forms the calcium-permeable ion channel and contains the obligatory co-agonist binding site. Previous studies have shown that NR1 gene (Grin1) expression is up-regulated during neuronal differentiation and its expression is widespread in the central nervous system. We have previously cloned the chicken Grin1 gene and 1.9 kb of the 5-regulatory region. In the present study, we analysed the molecular mechanisms that regulate chicken Grin1 gene transcription in undifferentiated cells and neurons. By functional analysis of chicken Grin1-luciferase gene 5-regulatory region constructs, we demonstrate that the basal promoter is delimited within 210 bp upstream from the main transcription initiation site. DNA-protein binding and functional assays revealed that the 5-UTR (untranslated region) has one consensus NRSE (neuron-restrictive silencing element) that binds NRSF (neuron-restrictive silencing factor), and one SP (stimulating protein transcription factor) element that binds SP3, both repressing Grin1 gene transcription in undifferentiated P19 cells (embryonic terato-carcinoma cells) and PC12 cells (phaeochromocytoma cells). The promoter region lacks a consensus TATA box, but contains one GSG/SP (GSG-like box near a SP-consensus site) that binds SP3 and up-regulates gene transcription in embryonic chicken cortical neurons. Taken together, these results demonstrate a dual role of SP3 in regulating the expression of the Grin1 gene, by repressing transcription in the 5-UTR in undifferentiated cells as well as acting as a transcription factor, increasing Grin1 gene transcription in neurons.

Biochimica Et Biophysica Acta: Molecular Basis Of Disease, 2021

et al., A systematic review of post-translational modifications in the mitochondrial permeability... more et al., A systematic review of post-translational modifications in the mitochondrial permeability transition pore complex associated with cardiac diseases, BBA-Molecular Basis of Disease (2020),

<p><b>A)</b> Chromatin Immunoprecipitation (ChIP) assay of the rat <i>CAS... more <p><b>A)</b> Chromatin Immunoprecipitation (ChIP) assay of the rat <i>CASQ2</i> gene promoter; fragmented chromatin was incubated with antibodies against MEF-2c, NFATc3, Sp1 and IgG. PCR was performed to amplify the region between -259 to -21 bp of <i>CASQ2</i> gene promoter. A picture of a representative gel is shown. <b>B)</b> Densitometry analysis of representative experiments (<i>n</i> = 2) was made with the Image-Studio Lite 5.2.5 software, (LI-COR, NE, USA). <b>C)</b> Total protein extracts were immunoprecipitated (IP) with anti-NFAT or anti-MEF-2 antibodies as indicated. Western blot analysis of immunoprecipitated proteins was performed using antibody against MEF-2 or NFAT as indicated. The identity of IP proteins is as follows: Lane 1, Mock (Protein A Sepharose antibody-binding beads without antibody); Lane 2, NFAT IP; Lane 3 MEF-2 IP. (The data showed in the graphs can be found as supporting information in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0184724#pone.0184724.s001&quot; target="_blank">S1 Dataset</a> file. The complete image of the blot can be found in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0184724#pone.0184724.s004&quot; target="_blank">S3 Fig</a> file).</p

<p>Lane 1: labeled TATA-box pAT5 dsDNA-biotin probe without <i>T</i>. <i>... more <p>Lane 1: labeled TATA-box pAT5 dsDNA-biotin probe without <i>T</i>. <i>solium</i> nuclear extract; lane 2: TATA-box pAT5 interaction with <i>T</i>. <i>solium</i> nuclear extract; lane 3: TATA-box pAT5 plus <i>T</i>. <i>solium</i> nuclear extract and anti-pTsTBP1-C antibodies; lane 4: <i>T</i>. <i>solium</i> nuclear extract plus anti-pTsTBP1-C with TATA-box pAT5; lane 5: TATA-box pAT5 plus <i>T</i>. <i>solium</i> nuclear extract and normal rabbit IgG, lane 6: <i>T</i>. <i>solium</i> nuclear extract plus normal rabbit IgG and TATA box pAT5, lane 7: TATA-box pAT5 plus <i>T</i>. <i>solium</i> nuclear extract and anti-pTsTBP1-N antibodies, and lane 8: <i>T</i>. <i>solium</i> nuclear extract plus anti-pTsTBP1-N antibodies and TATA-box pAT5.</p

<p>A) 10% SDS-PAGE of cysticerci <i>T</i>. <i>solium</i> nuclear ex... more <p>A) 10% SDS-PAGE of cysticerci <i>T</i>. <i>solium</i> nuclear extract patterns stained with Coomassie blue (lane 1). B) Western blot of TsTBP1 on <i>T</i>. <i>solium</i> nuclear extract with: normal serum IgG (lane 1), anti-pTsTBP1-N (lane 2), and anti-pTsTBP1-C antibodies (lane 3). C) Localization of TBP1 on <i>Taenia crassiceps</i> cysticerci sections by confocal microscopy with DAPI (blue), anti-histone H1 (green), anti-pTsTBP1-N antibodies (red) and merging of previous images (yellow signal). Negative control for primary and secondary antibodies, were normal mouse IgG plus anti-mouse IgG-Alexa-568 and normal rabbit IgG plus anti-rabbit IgG-Alexa-488. D) Digital amplification of a single nucleus to observe the localization of DNA (blue), histone H1 (green), and TBP1 (red), and their co-localization (yellow signal).</p

<p>A) Lane 1: Labeled dsDNA-³²P probe without nuclear extract; lane ... more <p>A) Lane 1: Labeled dsDNA-³²P probe without nuclear extract; lane 2: TsTBP1-pAT5 TATA-box interaction with <i>T</i>. <i>solium</i> nuclear extract; lanes 3, 4, and 5: competence with pAT5 TATA-box cold probe in a molar excess of 25X, 50X, and 100X, respectively; lane 6: super-shift interaction using anti-pTsTBP1-N; lane 7: consensus TATA-box probe interaction with <i>T</i>. <i>solium</i> nuclear extract (used as positive control); lane 8: consensus mutated TATA-box probe interaction with nuclear extract (used as negative control); lane 9, 10 and 11: cross-competence with Ts2-CysPrx TATA-box cold probe in a molar excess of 25X, 50X, and 100X, respectively; lane 12: anti-TsTBP1-N antibody without <i>T</i>. <i>solium</i> nuclear extract (negative control). Shifted, super-shifted bands and the free-labeled dsDNA probe, are indicated by arrows. B) Densitometric analysis shows a decrease on the intensity of shifted bands in homologous and heterologous competition. Results are present as percentage mean ± SD of the shifted band in lane 2 (P < 0.005).</p

<p><b>A)</b> To verify the NFAT overexpression, the NFATc1 or NFATc3 expression... more <p><b>A)</b> To verify the NFAT overexpression, the NFATc1 or NFATc3 expression vectors were transfected into neonatal rat cardiomyocytes. After 48 h, the cells were harvested and NFATc1 and NFATc3 protein levels were evaluated by Western blot. A picture of a representative gel is shown. Densitometry analysis of the blot was made using the Image-Studio Lite 5.2.5 software, LI-COR, NE, USA. <b>B)</b> Luciferase activity of neonatal cardiomyocytes co-transfected with pGL3-<i>hCASQ2</i>prom/Luc -3102/+176 bp (black bars) or pGL3-<i>hCASQ2</i>prom/Luc -288/+176 bp (gray bars) and the NFATc1 or NFATc3 expression vectors. After 24 h, the cells were harvested and luciferase activity was measured. <b>C)</b> Luciferase activity of neonatal cardiomyocytes co-transfected with pGL3-<i>hCASQ2</i>prom/Luc -3102/+176 bp construct and NFATc1 or NFATc3 expression vectors as above. Then, cells were treated with vehicle (black bars) or CsA (gray bars). After 24 h, the cells were harvested, and luciferase activity was measured. Results for functional assays (Fig 3B and 3C) are expressed as the mean of three independent experiments +/- SEM. The luciferase activity of control condition is given the arbitrary value of 1. *p values <0.05 were considered statistically significant. (The data showed in the graphs can be found as supporting information in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0184724#pone.0184724.s001&quot; target="_blank">S1 Dataset</a> file. The complete image of the blot can be found in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0184724#pone.0184724.s003&quot; target="_blank">S2 Fig</a> file).</p

<p>In bold letters are represented the putative TATA-box for each gene. Underlined bases ar... more <p>In bold letters are represented the putative TATA-box for each gene. Underlined bases are the mutated bases in the TATA-box consensus.</p

<p><b>A)</b> Luciferase activity of neonatal cardiomyocytes co-transfected with... more <p><b>A)</b> Luciferase activity of neonatal cardiomyocytes co-transfected with the pGL3-<i>hCASQ2</i>prom/Luc -288/+176 bp wild-type construct (black bars) or the -230 bp NFAT site mutated construct (gray bars) and the NFATc3 expression vector. <b>B)</b> Luciferase activity of neonatal cardiomyocytes co-transfected with pGL3-<i>hCASQ2</i>prom/Luc -288/+176 bp wild-type construct (black bars), the -103 bp SRF site mutated construct (gray bars) or the -133 bp MEF-2 site mutated construct (white bars) and the NFATc3 expression vector. After 24 h, the cells were harvested, and luciferase activity was determined. Results are expressed as the mean of three independent experiments +/- SEM. The -288 bp construct luciferase activity is given the arbitrary value of 1. *p values <0.05 were considered statistically significant. (The data showed in the graphs can be found as supporting information in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0184724#pone.0184724.s001&quot; target="_blank">S1 Dataset</a> file).</p

<p>TsTBP1 was aligned with <i>Sus scrofa</i> TBP1 (SsTBP1, GeneBank ID: XP_0033... more <p>TsTBP1 was aligned with <i>Sus scrofa</i> TBP1 (SsTBP1, GeneBank ID: XP_003361466.1), <i>Homo sapiens</i> TBP1 (HsTBP1, GeneBank ID: NP_003185.1), <i>E</i>. <i>granulosus</i> TBP1 (EgTBP1, GeneBank ID: CDS17003.1), <i>E</i>. <i>multilocularis</i> TBP1 (EmTBP1, GeneBank ID: CDJ04746.1). The NH₂-ter is enclosed in a box, and the remaining amino acid sequence corresponds to the COOH-terminal domain (COOH-ter). Identical amino acids are highlighted in gray background. Important residues that bind TATA-box are in red letters; transcription factor II A (TFIIA) in white; transcription factor II B (TFIIB) in blue; negative cofactor 2 (NC2) in orange and TBP1-associated factor 1 (TAF1) in yellow. Amino acid sequences used to produce the TsTBP probe and the synthetic peptides pTsTBP1-N and pTsTBP1-C are in small boxes and underlined, respectively. Letter X on <i>S</i>. <i>scrofa</i> TBP1 sequence means amino acids not identified. The symbols under the amino acids indicate: (-) absence and (:) homology of amino acids.</p

<p>A) Lane 1: Labeled dsDNA-³²P probe without nuclear extract; lane ... more <p>A) Lane 1: Labeled dsDNA-³²P probe without nuclear extract; lane 2: TsTBP1-Ts2-CysPrx TATA-box interaction with <i>T</i>. <i>solium</i> nuclear extract; lanes 3, 4, and 5: competence with Ts2-CysPrx TATA-box cold probe in a molar excess of 25X, 50X, and 100X, respectively; lane 6: super-shift interaction using anti-pTsTBP1-N; lane 7: consensus TATA-box probe interaction with <i>T</i>. <i>solium</i> nuclear extract (used as positive control); lane 8: consensus mutated TATA-box probe interaction with nuclear extract (used as negative control); lane 9, 10, and 11: cross-competence with pAT5 TATA-box cold probe in a molar excess of 25X, 50X, and 100X, respectively; lane 12: anti-TsTBP1-N antibody without <i>T</i>. <i>solium</i> nuclear extract (negative control). Shifted, super-shifted bands, and the free-labeled dsDNA probe are indicated by arrows. B) The densitometric analysis shows a decrease on the intensity of shifted bands in homologous and heterologous competition. Results are present as percentage mean ± SD of the shifted band in lane 2 (P < 0.005).</p

Canadian Journal of Physiology and Pharmacology, Aug 1, 2012

The precise control of Ca2+levels during the contraction–relaxation cycle in cardiac myocytes is ... more The precise control of Ca2+levels during the contraction–relaxation cycle in cardiac myocytes is extremely important for normal beat-to-beat contractile activity. The sarcoplasmic reticulum (SR) plays a key role controlling calcium concentration in the cytosol. The SR Ca2+-ATPase (SERCA2) transports Ca2+inside the SR lumen during relaxation of the cardiac myocyte. Calsequestrin (Casq2) is the main protein in the SR lumen, functioning as a Ca2+buffer and participating in Ca2+release by interacting with the ryanodine receptor 2 (RyR2) Ca2+-release channel. Alterations in normal Ca2+handling significantly contribute to the contractile dysfunction observed in cardiac hypertrophy and in heart failure. Transcriptional regulation of the SERCA2 gene has been extensively studied and some of the mechanisms regulating its expression have been elucidated. Overexpression of Sp1 factor in cardiac hypertrophy downregulates SERCA2 gene expression and increased levels of thyroid hormone up-regulates its transcription. Other hormones such norepinephrine, angiotensin II, endothelin-1, parathyroid hormone, prostaglandin-F2α, as well the cytokines tumor necrosis factor-α and interleukin-6 also downregulate SERCA2 expression. Calcium acting through the calcineurin–NFAT (nuclear factor of activated T cells) pathway has been suggested to regulate SERCA2 and CASQ2 gene expression. This review focuses on the current knowledge regarding transcriptional regulation of SERCA2 and CASQ2 genes in the normal and pathologic heart.

Journal of Molecular and Cellular Cardiology, Jun 1, 2007

to prevent its precipitation due to high concentrations of free Ca 2+ (2-10 mM). In mammals two C... more to prevent its precipitation due to high concentrations of free Ca 2+ (2-10 mM). In mammals two CASQ isoforms coded by two different genes have been described. The CASQ1 isoform is the only one expressed in fast-twitch skeletal muscle and constitutes the most abundant isoform in slow-twitch skeletal muscle (STSM) (∼ 75%). The CASQ2 isoform is expressed exclusively in cardiac muscle and to a lesser degree on STSM (∼ 25%). In this work we analyzed the transcription of the casq2 gene using several constructs containing the 5′-regulatory region of the human gene. The first 180 bp of promoter region shares high homology among species, which include the putative binding sites MEF-2, E-box and CArG-box for myogenic transcription factors. Functional analyses demonstrated that the proximal 288 bp has a strong transcriptional activity in neonatal rat cardiac myocytes. The data suggest that the elements present in the first 288 bp are sufficient to confer muscular specific expression, and that cis-elements contained in the upstream region (− 1.4 to − 1.7 kb) may act as a cardiac specific enhancer (i.e. MEF-2 and M-CAT). Site-directed mutagenesis of the MEF-2, E-box and CArG-box elements was performed. The MEF-2 and CArG-box, but not E-box mutations were associated with a decrease of the basal transcriptional activity by 70% and 50%, respectively. DNA/protein interaction assays revealed that MEF-2 and SRF bind to the MEF-2 and CArG elements.

The International Journal of Biochemistry & Cell Biology, Aug 1, 2019

Highlights  SERCA3 mRNA expression is decreased in hepatocelullar carcinoma cells (HCC)  NaB an... more Highlights  SERCA3 mRNA expression is decreased in hepatocelullar carcinoma cells (HCC)  NaB and TSA up-regulate SERCA3 mRNA expression in rat HCC  NaB and TSA increase H3K9 and H3K27 acetylation at the ATP2A3 promoter  TSA increases occupancy of p300 at the ATP2A3 promoter  Inhibition of p300 decreases basal and TSA-mediated SERCA3 mRNA expression  p300 mediates H3K9 acetylation at the ATP2A3 promoter

The FASEB Journal, Mar 1, 2006

Cell Calcium, Nov 1, 2020

Calcium (Ca2+) signaling controls a wide range of cellular processes, including the hallmarks of ... more Calcium (Ca2+) signaling controls a wide range of cellular processes, including the hallmarks of cancer. The Ca2+ signaling system encompasses several types of proteins, such as receptors, channels, pumps, exchangers, buffers, and sensors, of which several are mutated or with altered expression in cancer cells. Since epigenetic mechanisms are disrupted in all stages of carcinogenesis, and reversibly regulate gene expression, they have been studied by different research groups to understand their role in Ca2+ signaling remodeling in cancer cells and the carcinogenic process. In this review, we link Ca2+ signaling, cancer, and epigenetics fields to generate a comprehensive landscape of this complex group of diseases.

Cell Calcium, Nov 1, 2014

Cytosolic calcium concentration ([Ca(2+)]c) is fundamental for regulation of many cellular proces... more Cytosolic calcium concentration ([Ca(2+)]c) is fundamental for regulation of many cellular processes such metabolism, proliferation, muscle contraction, cell signaling and insulin secretion. In resting conditions, the sarco/endoplasmic reticulum (ER/SR) Ca(2+) ATPase&amp;amp;amp;amp;amp;amp;#39;s (SERCA) transport Ca(2+) from the cytosol to the ER or SR lumen, maintaining the resting [Ca(2+)]c about 25-100nM. A reduced activity and expression of SERCA2 protein have been described in heart failure and diabetic cardiomyopathy, resulting in an altered Ca(2+) handling and cardiac contractility. In the diabetic pancreas, there has been reported reduction in SERCA2b and SERCA3 expression in β-cells, resulting in diminished insulin secretion. Evidence obtained from different diabetes models has suggested a role for advanced glycation end products formation, oxidative stress and increased O-GlcNAcylation in the lowered SERCA2 expression observed in diabetic cardiomyopathy. However, the role of SERCA2 down-regulation in the pathophysiology of diabetes mellitus and diabetic cardiomyopathy is not yet well described. In this review, we make a comprehensive analysis of the current knowledge of the role of the SERCA pumps in the pathophysiology of insulin-dependent diabetes mellitus type 1 (TIDM) and type 2 (T2DM) in the heart and β-cells in the pancreas.

Oncology Letters, May 31, 2018

Period circadian regulator (Per)1 and Per2 genes are involved in the molecular mechanism of the c... more Period circadian regulator (Per)1 and Per2 genes are involved in the molecular mechanism of the circadian clock, and exhibit tumor suppressor properties. Several studies have reported a decreased expression of Per1, Per2 and Per3 genes in different types of cancer and cancer cell lines. Promoter methylation downregulates Per1, Per2 or Per3 expression in myeloid leukemia, breast, lung, and other cancer cells; whereas histone deacetylase inhibitors (HDACi) upregulate Per1 or Per3 expression in certain cancer cell lines. However, the transcriptional regulation of Per1 and Per2 in cancer cells by chromatin modifications is not fully understood. The present study aimed to determine whether HDACi regulate Per1 and Per2 expression in gastric cancer cell lines, and to investigate changes in chromatin modifications in response to HDACi. Treatment of KATO III and NCI-N87 human gastric cancer cells with sodium butyrate (NaB) or Trichostatin A (TSA) induced Per1 and Per2 mRNA expression in a dose-dependent manner. Chromatin immunoprecipitaion assays revealed that NaB and TSA decreased lysine 9 trimethylation on histone H3 (H3K9me3) at the Per1 promoter. TSA, but not NaB increased H3K9 acetylation at the Per2 promoter. It was also observed that binding of Sp1 and Sp3 to the Per1 promoter decreased following NaB treatment, whereas Sp1 binding increased at the Per2 promoter of NaB-and TSA-treated cells. In addition, Per1 promoter is not methylated in KATO III cells, while Per2 promoter was methylated, although NaB, TSA, and 5-Azacytidine do not change the methylated CpGs analyzed. In conclusion, HDACi induce Per1 and Per2 expression, in part, through mechanisms involving chromatin remodeling at the proximal promoter of these genes; however, other indirect mechanisms triggered by these HDACi cannot be ruled out. These findings reveal a previously unappreciated regulatory pathway between silencing of Per1 gene by H3K9me3 and upregulation of Per2 by HDACi in cancer cells.

Molecular Carcinogenesis, Jan 29, 2019

The knowledge about the role of calcium‐regulated pathways in cancer cell growth and differentiat... more The knowledge about the role of calcium‐regulated pathways in cancer cell growth and differentiation could be useful for the development of new therapeutic approaches to diminish its mortality. The ATP2A genes encode for SERCA pumps, which modulate cytosolic Ca2+ concentration, regulating various cellular processes including cell growth. ATP2A3 gene transcriptional down‐regulation has been reported in gastric and colon cancer, but there is still a lack of understanding about the epigenetic processes regulating its transcription. In this work, we report that butyrate, trichostatin A, and 5‐azacytidine treatments increase SERCA3 expression, increased apoptosis, and decreased cell viability of the KATO‐III gastric carcinoma cell line. We analyzed the methylation profile of the ATP2A3 gene promoter CpG island, finding clones with methylated status through −280 to −135 promoter region, harboring Sp1 and AP‐2 binding sites, which could have a role in transcriptional repression. Post‐translational modifications of histones show a major role in the ATP2A3 transcriptional regulation, and our results show histones marks linked to transcriptional repression associated with the −262 to −135 region, this repressive context changed to transcriptional permissive through SERCA3 re‐expressing conditions. These results suggest that the nucleotide sequence from −280 to −135 position is an ATP2A3 epigenetic regulatory CpG region in KATO‐III cells. Analyses of online‐databases show a decreased SERCA3 expression in gastric and colon tumors, as well as overall survival results, showed that high SERCA3 expression could serve as a favorable prognostic marker for colon and gastric cancer patients.

Bioscience Reports, Aug 1, 2008

The NMDA (N-methyl-D-aspartate) receptors are important in the regulation of neuronal development... more The NMDA (N-methyl-D-aspartate) receptors are important in the regulation of neuronal development, synaptic plasticity, learning and memory, and are involved in several brain pathologies. The NR1 subunit is essential for the assembly of functional receptors, as it forms the calcium-permeable ion channel and contains the obligatory co-agonist binding site. Previous studies have shown that NR1 gene (Grin1) expression is up-regulated during neuronal differentiation and its expression is widespread in the central nervous system. We have previously cloned the chicken Grin1 gene and 1.9 kb of the 5-regulatory region. In the present study, we analysed the molecular mechanisms that regulate chicken Grin1 gene transcription in undifferentiated cells and neurons. By functional analysis of chicken Grin1-luciferase gene 5-regulatory region constructs, we demonstrate that the basal promoter is delimited within 210 bp upstream from the main transcription initiation site. DNA-protein binding and functional assays revealed that the 5-UTR (untranslated region) has one consensus NRSE (neuron-restrictive silencing element) that binds NRSF (neuron-restrictive silencing factor), and one SP (stimulating protein transcription factor) element that binds SP3, both repressing Grin1 gene transcription in undifferentiated P19 cells (embryonic terato-carcinoma cells) and PC12 cells (phaeochromocytoma cells). The promoter region lacks a consensus TATA box, but contains one GSG/SP (GSG-like box near a SP-consensus site) that binds SP3 and up-regulates gene transcription in embryonic chicken cortical neurons. Taken together, these results demonstrate a dual role of SP3 in regulating the expression of the Grin1 gene, by repressing transcription in the 5-UTR in undifferentiated cells as well as acting as a transcription factor, increasing Grin1 gene transcription in neurons.

Biochimica Et Biophysica Acta: Molecular Basis Of Disease, 2021

et al., A systematic review of post-translational modifications in the mitochondrial permeability... more et al., A systematic review of post-translational modifications in the mitochondrial permeability transition pore complex associated with cardiac diseases, BBA-Molecular Basis of Disease (2020),

<p><b>A)</b> Chromatin Immunoprecipitation (ChIP) assay of the rat <i>CAS... more <p><b>A)</b> Chromatin Immunoprecipitation (ChIP) assay of the rat <i>CASQ2</i> gene promoter; fragmented chromatin was incubated with antibodies against MEF-2c, NFATc3, Sp1 and IgG. PCR was performed to amplify the region between -259 to -21 bp of <i>CASQ2</i> gene promoter. A picture of a representative gel is shown. <b>B)</b> Densitometry analysis of representative experiments (<i>n</i> = 2) was made with the Image-Studio Lite 5.2.5 software, (LI-COR, NE, USA). <b>C)</b> Total protein extracts were immunoprecipitated (IP) with anti-NFAT or anti-MEF-2 antibodies as indicated. Western blot analysis of immunoprecipitated proteins was performed using antibody against MEF-2 or NFAT as indicated. The identity of IP proteins is as follows: Lane 1, Mock (Protein A Sepharose antibody-binding beads without antibody); Lane 2, NFAT IP; Lane 3 MEF-2 IP. (The data showed in the graphs can be found as supporting information in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0184724#pone.0184724.s001&quot; target="_blank">S1 Dataset</a> file. The complete image of the blot can be found in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0184724#pone.0184724.s004&quot; target="_blank">S3 Fig</a> file).</p

<p>Lane 1: labeled TATA-box pAT5 dsDNA-biotin probe without <i>T</i>. <i>... more <p>Lane 1: labeled TATA-box pAT5 dsDNA-biotin probe without <i>T</i>. <i>solium</i> nuclear extract; lane 2: TATA-box pAT5 interaction with <i>T</i>. <i>solium</i> nuclear extract; lane 3: TATA-box pAT5 plus <i>T</i>. <i>solium</i> nuclear extract and anti-pTsTBP1-C antibodies; lane 4: <i>T</i>. <i>solium</i> nuclear extract plus anti-pTsTBP1-C with TATA-box pAT5; lane 5: TATA-box pAT5 plus <i>T</i>. <i>solium</i> nuclear extract and normal rabbit IgG, lane 6: <i>T</i>. <i>solium</i> nuclear extract plus normal rabbit IgG and TATA box pAT5, lane 7: TATA-box pAT5 plus <i>T</i>. <i>solium</i> nuclear extract and anti-pTsTBP1-N antibodies, and lane 8: <i>T</i>. <i>solium</i> nuclear extract plus anti-pTsTBP1-N antibodies and TATA-box pAT5.</p

<p>A) 10% SDS-PAGE of cysticerci <i>T</i>. <i>solium</i> nuclear ex... more <p>A) 10% SDS-PAGE of cysticerci <i>T</i>. <i>solium</i> nuclear extract patterns stained with Coomassie blue (lane 1). B) Western blot of TsTBP1 on <i>T</i>. <i>solium</i> nuclear extract with: normal serum IgG (lane 1), anti-pTsTBP1-N (lane 2), and anti-pTsTBP1-C antibodies (lane 3). C) Localization of TBP1 on <i>Taenia crassiceps</i> cysticerci sections by confocal microscopy with DAPI (blue), anti-histone H1 (green), anti-pTsTBP1-N antibodies (red) and merging of previous images (yellow signal). Negative control for primary and secondary antibodies, were normal mouse IgG plus anti-mouse IgG-Alexa-568 and normal rabbit IgG plus anti-rabbit IgG-Alexa-488. D) Digital amplification of a single nucleus to observe the localization of DNA (blue), histone H1 (green), and TBP1 (red), and their co-localization (yellow signal).</p

<p>A) Lane 1: Labeled dsDNA-³²P probe without nuclear extract; lane ... more <p>A) Lane 1: Labeled dsDNA-³²P probe without nuclear extract; lane 2: TsTBP1-pAT5 TATA-box interaction with <i>T</i>. <i>solium</i> nuclear extract; lanes 3, 4, and 5: competence with pAT5 TATA-box cold probe in a molar excess of 25X, 50X, and 100X, respectively; lane 6: super-shift interaction using anti-pTsTBP1-N; lane 7: consensus TATA-box probe interaction with <i>T</i>. <i>solium</i> nuclear extract (used as positive control); lane 8: consensus mutated TATA-box probe interaction with nuclear extract (used as negative control); lane 9, 10 and 11: cross-competence with Ts2-CysPrx TATA-box cold probe in a molar excess of 25X, 50X, and 100X, respectively; lane 12: anti-TsTBP1-N antibody without <i>T</i>. <i>solium</i> nuclear extract (negative control). Shifted, super-shifted bands and the free-labeled dsDNA probe, are indicated by arrows. B) Densitometric analysis shows a decrease on the intensity of shifted bands in homologous and heterologous competition. Results are present as percentage mean ± SD of the shifted band in lane 2 (P < 0.005).</p

<p><b>A)</b> To verify the NFAT overexpression, the NFATc1 or NFATc3 expression... more <p><b>A)</b> To verify the NFAT overexpression, the NFATc1 or NFATc3 expression vectors were transfected into neonatal rat cardiomyocytes. After 48 h, the cells were harvested and NFATc1 and NFATc3 protein levels were evaluated by Western blot. A picture of a representative gel is shown. Densitometry analysis of the blot was made using the Image-Studio Lite 5.2.5 software, LI-COR, NE, USA. <b>B)</b> Luciferase activity of neonatal cardiomyocytes co-transfected with pGL3-<i>hCASQ2</i>prom/Luc -3102/+176 bp (black bars) or pGL3-<i>hCASQ2</i>prom/Luc -288/+176 bp (gray bars) and the NFATc1 or NFATc3 expression vectors. After 24 h, the cells were harvested and luciferase activity was measured. <b>C)</b> Luciferase activity of neonatal cardiomyocytes co-transfected with pGL3-<i>hCASQ2</i>prom/Luc -3102/+176 bp construct and NFATc1 or NFATc3 expression vectors as above. Then, cells were treated with vehicle (black bars) or CsA (gray bars). After 24 h, the cells were harvested, and luciferase activity was measured. Results for functional assays (Fig 3B and 3C) are expressed as the mean of three independent experiments +/- SEM. The luciferase activity of control condition is given the arbitrary value of 1. *p values <0.05 were considered statistically significant. (The data showed in the graphs can be found as supporting information in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0184724#pone.0184724.s001&quot; target="_blank">S1 Dataset</a> file. The complete image of the blot can be found in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0184724#pone.0184724.s003&quot; target="_blank">S2 Fig</a> file).</p

<p>In bold letters are represented the putative TATA-box for each gene. Underlined bases ar... more <p>In bold letters are represented the putative TATA-box for each gene. Underlined bases are the mutated bases in the TATA-box consensus.</p

<p><b>A)</b> Luciferase activity of neonatal cardiomyocytes co-transfected with... more <p><b>A)</b> Luciferase activity of neonatal cardiomyocytes co-transfected with the pGL3-<i>hCASQ2</i>prom/Luc -288/+176 bp wild-type construct (black bars) or the -230 bp NFAT site mutated construct (gray bars) and the NFATc3 expression vector. <b>B)</b> Luciferase activity of neonatal cardiomyocytes co-transfected with pGL3-<i>hCASQ2</i>prom/Luc -288/+176 bp wild-type construct (black bars), the -103 bp SRF site mutated construct (gray bars) or the -133 bp MEF-2 site mutated construct (white bars) and the NFATc3 expression vector. After 24 h, the cells were harvested, and luciferase activity was determined. Results are expressed as the mean of three independent experiments +/- SEM. The -288 bp construct luciferase activity is given the arbitrary value of 1. *p values <0.05 were considered statistically significant. (The data showed in the graphs can be found as supporting information in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0184724#pone.0184724.s001&quot; target="_blank">S1 Dataset</a> file).</p

<p>TsTBP1 was aligned with <i>Sus scrofa</i> TBP1 (SsTBP1, GeneBank ID: XP_0033... more <p>TsTBP1 was aligned with <i>Sus scrofa</i> TBP1 (SsTBP1, GeneBank ID: XP_003361466.1), <i>Homo sapiens</i> TBP1 (HsTBP1, GeneBank ID: NP_003185.1), <i>E</i>. <i>granulosus</i> TBP1 (EgTBP1, GeneBank ID: CDS17003.1), <i>E</i>. <i>multilocularis</i> TBP1 (EmTBP1, GeneBank ID: CDJ04746.1). The NH₂-ter is enclosed in a box, and the remaining amino acid sequence corresponds to the COOH-terminal domain (COOH-ter). Identical amino acids are highlighted in gray background. Important residues that bind TATA-box are in red letters; transcription factor II A (TFIIA) in white; transcription factor II B (TFIIB) in blue; negative cofactor 2 (NC2) in orange and TBP1-associated factor 1 (TAF1) in yellow. Amino acid sequences used to produce the TsTBP probe and the synthetic peptides pTsTBP1-N and pTsTBP1-C are in small boxes and underlined, respectively. Letter X on <i>S</i>. <i>scrofa</i> TBP1 sequence means amino acids not identified. The symbols under the amino acids indicate: (-) absence and (:) homology of amino acids.</p

<p>A) Lane 1: Labeled dsDNA-³²P probe without nuclear extract; lane ... more <p>A) Lane 1: Labeled dsDNA-³²P probe without nuclear extract; lane 2: TsTBP1-Ts2-CysPrx TATA-box interaction with <i>T</i>. <i>solium</i> nuclear extract; lanes 3, 4, and 5: competence with Ts2-CysPrx TATA-box cold probe in a molar excess of 25X, 50X, and 100X, respectively; lane 6: super-shift interaction using anti-pTsTBP1-N; lane 7: consensus TATA-box probe interaction with <i>T</i>. <i>solium</i> nuclear extract (used as positive control); lane 8: consensus mutated TATA-box probe interaction with nuclear extract (used as negative control); lane 9, 10, and 11: cross-competence with pAT5 TATA-box cold probe in a molar excess of 25X, 50X, and 100X, respectively; lane 12: anti-TsTBP1-N antibody without <i>T</i>. <i>solium</i> nuclear extract (negative control). Shifted, super-shifted bands, and the free-labeled dsDNA probe are indicated by arrows. B) The densitometric analysis shows a decrease on the intensity of shifted bands in homologous and heterologous competition. Results are present as percentage mean ± SD of the shifted band in lane 2 (P < 0.005).</p