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Sample records for 2b subunit gene

  1. Localization of eight additional genes in the human major histocompatibility complex, including the gene encoding the casein kinase II {beta} subunit (CSNK2B)

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    Albertella, M.R.; Jones, H.; Thomson, W. [Oxford Univ. (United Kingdom)] [and others

    1996-09-01

    A wide range of autoimmune and other diseases are known to be associated with the major histocompatibility complex. Many of these diseases are linked to the genes encoding the polymorphic histocompatibility complex. Many of these diseases are linked to the genes encoding the polymorphic histocompatibility antigens in the class I and class II regions, but some appear to be more strongly associated with genes in the central 1100-kb class III region, making it important to characterize this region fully for the presence of novel genes. An {approximately}220-kb segment of DNA in the class III region separating the Hsp70 (HSPA1L) and BAT1 (D6S8IE) genes, which was previously known to contain 14 genes. Genomic DNA fragments spanning the gaps between the known genes were used as probes to isolate cDNAs corresponding to five new genes within this region. Evidence from Northern blot analysis and exon trapping experiments that suggested the presence of at least two more new genes was also obtained. Partial cDNA and complete exonic genomic sequencing of one of the new genes has identified it as the casein kinase II{beta} subunit (CSNK2B). Two of the other novel genes lie within a region syntenic to that implicated in susceptibility to experimental allergic orchitis in the mouse, an autoimmune disease of the testis, and represent additional candidates for the Orch-1 locus associated with this disease. In addition, characterization of the 13-kb intergenic gap separating the RD (D6545) and G11 (D6S60E) genes has revealed the presence of a gene encoding a 1246-amino-acid polypeptide that shows significant sequence similarity to the yeast anti-viral Ski2p gene product. 49 refs., 8 figs.

  2. 霍乱毒素B亚单位与志贺毒素2B亚单位融合表达及抗原性检测%Clone and express ctb-stx2 b fusion gene in Enterohemrrhagic escherichia coli O157:H7 Shigeai toxin 2B subunit and V cholera toxin B subunit and the detection of their immunogenicity

    Institute of Scientific and Technical Information of China (English)

    李振军; 孙强正; 景怀琦; 徐建国

    2008-01-01

    Objective To clone and express the fusion gene encoding Enterohemrrhagic escherichia coli O157:H7(EHEC O157:H7)Shigela toxin 2B subunit(Stx2B)and vibrio cholera toxin B subunit (CTB)as well as to detect the immunogenicity and GM1-binding ability of fusion protein.Methods To design a primer to amplify stx2b gene and ctb-stx2b fusion gene encoding Stx2B and CTB-Stx2B respectively and to clone the genes into express plasmid pET30a(+)C in order to construct pET30a-ctb-stx2b after T-A sequencing was varified,then to transform constructed plasmid into E.coliBL21(DE3)induced by IPTG and purified by a purify kit and to detect molecular weight and immunogenicity by SDSPAGE and Western-blot.Results The amplified ctb-stx2b fragments appeared to he 750 bp and gene sequence was identical to designed sequence.The prokaryotic expression system pET30a-ctb-stx2b/BL21 could express protein weight about Mr20×103and the expressed protein could react to CTB monoclone anti-body.The fusion protein CTB-Stx2B could bind GM1.Conclusion CTB-Stx2B had successfully been expressed in prokaryotic while the expressed protein had good immunogenicity and GM1-Binding ability.This study provided information on further EHEC O157:H7 vaccine research.%目的 克隆表达出血性大肠埃希菌(EHEC)O157:H7志贺毒素2B亚单位(Stx2B)与霍乱毒素B亚单位(CTB)的融合蛋白(CTB-Stx2B),并检测其抗原性和与神经节苷脂(GM1)结合能力.方法 设计引物扩增融合蛋白CTB-Stx2B的编码基因ctb-stx2b,T-A克隆测序验证后克隆入原核表达质粒pET30a(+)C,构建表达质粒pET30a(+)-ctb-stx2b,转化E.coliBL21(DE3),IPTG诱导表达、纯化,获得目的蛋白CTB-Stx2B,SDS-PAGE和Western-blot检测其抗原性和形成五聚体的能力;GM1-ELISA法检测其与GM1结合能力.结果 扩增出约750 bp的目的片段,测序鉴定与设计序列一致;原核表达质粒pET30a(+)-ctb-stx2b转化E.coliBL21(DE3)后,经酶切和PCR扩增鉴定正确;IPTG

  3. Identification of a Novel Rat NR2B Subunit Gene Promoter Region Variant and Its Association with Microwave-Induced Neuron Impairment.

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    Wang, Li-Feng; Tian, Da-Wei; Li, Hai-Juan; Gao, Ya-Bing; Wang, Chang-Zhen; Zhao, Li; Zuo, Hong-Yan; Dong, Ji; Qiao, Si-Mo; Zou, Yong; Xiong, Lu; Zhou, Hong-Mei; Yang, Yue-Feng; Peng, Rui-Yun; Hu, Xiang-Jun

    2016-05-01

    Microwave radiation has been implicated in cognitive dysfunction and neuronal injury in animal models and in human investigations; however, the mechanism of these effects is unclear. In this study, single nucleotide polymorphism (SNP) sites in the rat GRIN2B promoter region were screened. The associations of these SNPs with microwave-induced rat brain dysfunction and with rat pheochromocytoma-12 (PC12) cell function were investigated. Wistar rats (n = 160) were exposed to microwave radiation (30 mW/cm(2) for 5 min/day, 5 days/week, over a period of 2 months). Screening of the GRIN2B promoter region revealed a stable C-to-T variant at nucleotide position -217 that was not induced by microwave exposure. The learning and memory ability, amino acid contents in the hippocampus and cerebrospinal fluid, and NR2B expression were then investigated in the different genotypes. Following microwave exposure, NR2B protein expression decreased, while the Glu contents in the hippocampus and CSF increased, and memory impairment was observed in the TT genotype but not the CC and CT genotypes. In PC12 cells, the effects of the T allele were more pronounced than those of the C allele on transcription factor binding ability, transcriptional activity, NR2B mRNA, and protein expression. These effects may be related to the detrimental role of the T allele and the protective role of the C allele in rat brain function and PC12 cells exposed to microwave radiation.

  4. Evaluation of NR2B peptide as subunit vaccines against experimental neuropathic pain

    Institute of Scientific and Technical Information of China (English)

    WANG Gong-ming; TIAN Yu-ke; CHEN Jian-ping; TIAN Xu-bi; GAO Feng; YANG Hui; AN Ke; MA Guo-ping

    2007-01-01

    Background NR2B containing N-methyl-D-aspartate (NMDA) receptor plays an important role in the facilitation and maintenance of neuropathic pain. The discrete distribution of NR2B subunit in the central nervous system (CNS) may support reduced side effects of agents that act selectively at this site. Therefore, we investigated the hypothesis that a humoral autoimmune response targeting the NR2B subunit of NMDA receptor relieves pain like behaviours produced by peripheral injury.Methods Rats were immunized subcutaneously with NR2B-Keyhole Limpet Hemocyanin (NR2B-KLH) three times at two-week intervals. NR2B specific IgG titres in sera and cerebrospinal fluid were determined by indirect ELISA. Seven days after the third immunization, 2 of the 3 terminal branches of the sciatic nerve (tibial and common peroneal nerves) were tightly ligated. Behavioural testing was carried out on every other day after surgery, until 7 days after surgery. The lumbar spinal cord (L4-6) was removed on day 7 after ligation. The expression of NR2B protein in the lumbar spinal cord was determined using Western blotting.Results After the second vaccination, NR2B specific IgG in sera was detected to be >0.5 μg/ml in six of nine rats. After the third vaccination, all the immunized rats had >2.2 μg/ml. Titres of NR2B specific IgG in sera peaked 42 days post initial immunization and persisted for over 70 days. No NR2B specific IgG was detected in sera from PBS or KLH group.The behavioural thresholds in NR2B group were significantly higher than those in PBS and KLH groups on day 7 after ligation. NR2B specific IgG in CSF in NR2B group could not be detected on day 1 before spinal dissection; but could be detected on day 7 after surgery. The expression of NR2B protein in group NR2B was significantly lower than in PBS and KLH groups on day 7 after surgery.Conclusion The NR2B peptide could be used as a vaccine against neuropathic pain, which could be associated with reduction of NR2B protein in

  5. genetic overexpression of NR2B subunit enhances social recognition memory for different strains and species.

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    Stephanie A Jacobs

    Full Text Available The ability to learn and remember conspecifics is essential for the establishment and maintenance of social groups. Many animals, including humans, primates and rodents, depend on stable social relationships for survival. Social learning and social recognition have become emerging areas of interest for neuroscientists but are still not well understood. It has been established that several hormones play a role in the modulation of social recognition including estrogen, oxytocin and arginine vasopression. Relatively few studies have investigated how social recognition might be improved or enhanced. In this study, we investigate the role of the NMDA receptor in social recognition memory, specifically the consequences of altering the ratio of the NR2B:NR2A subunits in the forebrain regions in social behavior. We produced transgenic mice in which the NR2B subunit of the NMDA receptor was overexpressed postnatally in the excitatory neurons of the forebrain areas including the cortex, amygdala and hippocampus. We investigated the ability of both our transgenic animals and their wild-type littermate to learn and remember juvenile conspecifics using both 1-hr and 24-hr memory tests. Our experiments show that the wild-type animals and NR2B transgenic mice preformed similarly in the 1-hr test. However, transgenic mice showed better performances in 24-hr tests of recognizing animals of a different strain or animals of a different species. We conclude that NR2B overexpression in the forebrain enhances social recognition memory for different strains and animal species.

  6. Association Study of N-Methyl-D-Aspartate Receptor Subunit 2B (GRIN2B Polymorphisms and Schizophrenia Symptoms in the Han Chinese Population.

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    Yongfeng Yang

    Full Text Available Schizophrenia (SZ is a common and complex psychiatric disorder that has a significant genetic component. The glutamatergic system is the major excitatory neurotransmitter system in the central nervous system, and is mediated by N-methyl-D-aspartate (NMDA receptors. Disturbances in this system have been hypothesized to play a major role in SZ pathogenesis. Several studies have revealed that the NMDA receptor subunit 2B (GRIN2B potentially associates with SZ and its psychiatric symptoms. In this study, we performed a case-control study to identify polymorphisms of the GRIN2B gene that may confer susceptibility to SZ in the Han Chinese population. Thirty-four single nucleotide polymorphisms (SNPs were genotyped in 528 paranoid SZ patients and 528 control subjects. A significant association was observed in allele and genotype between SZ and controls at rs2098469 (χ2 = 8.425 and 4.994; p = 0.025 and 0.014, respectively. Significant associations were found in the allele at rs12319804 (χ2 = 4.436; p = 0.035, as well as in the genotype at rs12820037 and rs7298664 between SZ and controls (χ2 = 11.162 and 38.204; p = 0.003 and 4.27×10(-8, respectively. After applying the Bonferroni correction, rs7298664 still had significant genotype associations with SZ (p = 1.71×10(-7. In addition, rs2098469 genotype and allele frequencies, and 12820037 allele frequencies were nominally associated with SZ. Three haplotypes, CGA (rs10845849-rs12319804-rs10845851, CC (rs12582848-rs7952915, and AAGAC (rs2041986-rs11055665-rs7314376-rs7297101-rs2098469, had significant differences between SZ and controls (χ2 = 4.324, 4.582, and 4.492; p = 0.037, 0.032, and 0.034, respectively. In addition, three SNPs, rs2098469, rs12820037, and rs7298664, were significantly associated with cognition factors PANSS subscores in SZ (F = 16.799, 7.112, and 13.357; p = 0.000, 0.017, and 0.000, respectively. In conclusion, our study provides novel evidence for an association between GRIN2B

  7. Association Study of N-Methyl-D-Aspartate Receptor Subunit 2B (GRIN2B) Polymorphisms and Schizophrenia Symptoms in the Han Chinese Population

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    Zhang, Hongxing; Yang, Ge; Wang, Xiujuan; Ding, Minli; Jiang, Tianzi; Lv, Luxian

    2015-01-01

    Schizophrenia (SZ) is a common and complex psychiatric disorder that has a significant genetic component. The glutamatergic system is the major excitatory neurotransmitter system in the central nervous system, and is mediated by N-methyl-D-aspartate (NMDA) receptors. Disturbances in this system have been hypothesized to play a major role in SZ pathogenesis. Several studies have revealed that the NMDA receptor subunit 2B (GRIN2B) potentially associates with SZ and its psychiatric symptoms. In this study, we performed a case–control study to identify polymorphisms of the GRIN2B gene that may confer susceptibility to SZ in the Han Chinese population. Thirty-four single nucleotide polymorphisms (SNPs) were genotyped in 528 paranoid SZ patients and 528 control subjects. A significant association was observed in allele and genotype between SZ and controls at rs2098469 (χ2 = 8.425 and 4.994; p = 0.025 and 0.014, respectively). Significant associations were found in the allele at rs12319804 (χ2 = 4.436; p = 0.035), as well as in the genotype at rs12820037 and rs7298664 between SZ and controls (χ2 = 11.162 and 38.204; p = 0.003 and 4.27×10-8, respectively). After applying the Bonferroni correction, rs7298664 still had significant genotype associations with SZ (p = 1.71×10-7). In addition, rs2098469 genotype and allele frequencies, and 12820037 allele frequencies were nominally associated with SZ. Three haplotypes, CGA (rs10845849—rs12319804—rs10845851), CC (rs12582848—rs7952915), and AAGAC (rs2041986—rs11055665—rs7314376—rs7297101—rs2098469), had significant differences between SZ and controls (χ2 = 4.324, 4.582, and 4.492; p = 0.037, 0.032, and 0.034, respectively). In addition, three SNPs, rs2098469, rs12820037, and rs7298664, were significantly associated with cognition factors PANSS subscores in SZ (F = 16.799, 7.112, and 13.357; p = 0.000, 0.017, and 0.000, respectively). In conclusion, our study provides novel evidence for an association between

  8. NMDA receptor NR2B subunits contribute to PTZ-kindling-induced hippocampal astrocytosis and oxidative stress.

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    Zhu, Xinjian; Dong, Jingde; Shen, Kai; Bai, Ying; Zhang, Yuan; Lv, Xuan; Chao, Jie; Yao, Honghong

    2015-05-01

    The N-methyl-d-aspartate (NMDA) receptor plays an important role in the pathophysiology of several neurological diseases, including epilepsy. The present study investigated the effect of NMDA receptor NR2B subunits on pentylenetetrazole (PTZ)-kindling-induced pathological and biochemical events in mice. Our results showed that PTZ-kindling up-regulates the expression of NMDA receptor NR2B subunits in the hippocampus and that kindled mice were characterized by significant astrocytosis and neuron loss in the hippocampus. Oxidative stress, including excessive malondialdehyde (MDA) production and decreased enzymatic activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-PX), were detected in the hippocampus after the mice were fully kindled. Additionally, expression of brain-derived neurotrophic factor (BDNF) in the hippocampus was found to be up-regulated in PTZ-kindled mice. However, selectively blocking NMDA receptor NR2B subunits by ifenprodil significantly suppressed PTZ-kindling-induced hippocampal astrocytosis, oxidative stress and neuron loss. Furthermore, blocking NMDA receptor NR2B subunits also abolished PTZ-kindling-induced BDNF expression. These results indicate that NMDA receptor NR2B subunits contribute to epilepsy-associated pathological and biochemical events, including hippocampal astrocytosis, oxidative stress and neuron loss, and these events might be correlated with up-regulation of BDNF expression.

  9. Co-activation of NR2A and NR2B subunits induces resistance to fear extinction.

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    Leaderbrand, Katherine; Corcoran, Kevin A; Radulovic, Jelena

    2014-09-01

    Unpredictable stress is known to profoundly enhance susceptibility to fear and anxiety while reducing the ability to extinguish fear when threat is no longer present. Accordingly, partial aversive reinforcement, via random exposure to footshocks, induces fear that is resistant to extinction. Here we sought to determine the hippocampal mechanisms underlying susceptibility versus resistance to context fear extinction as a result of continuous (CR) and partial (PR) reinforcement, respectively. We focused on N-methyl-D-aspartate receptor (NMDAR) subunits 2A and B (NR2A and NR2B) as well as their downstream signaling effector, extracellular signal-regulated kinase (ERK), based on their critical role in the acquisition and extinction of fear. Pharmacological inactivation of NR2A, but not NR2B, blocked extinction after CR, whereas inactivation of NR2A, NR2B, or both subunits facilitated extinction after PR. The latter finding suggests that co-activation of NR2A and NR2B contributes to persistent fear following PR. In contrast to CR, PR increased membrane levels of ERK and NR2 subunits after the conditioning and extinction sessions, respectively. In parallel, nuclear activation of ERK was significantly reduced after the extinction session. Thus, co-activation and increased surface expression of NR2A and NR2B, possibly mediated by ERK, may cause persistent fear. These findings suggest that patients with post-traumatic stress disorder (PTSD) may benefit from antagonism of specific NR2 subunits.

  10. Perinatal exposure to PTU delays switching from NR2B to NR2A subunits of the NMDA receptor in the rat cerebellum.

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    Kobayashi, Kumiko; Tsuji, Ryozo; Yoshioka, Takafumi; Mino, Terumasa; Seki, Takaki

    2006-03-01

    Certain kinds of developmental neurotoxicants are considered to act by affecting the levels of thyroid hormones, which are essential for the brain development of both humans and experimental animals. Hypothyroidism experimentally induced in rats with propylthiouracil (PTU) offers a useful animal model for developmental neurotoxicity. The purpose of the present study was to clarify developmental alterations in gene expression caused by PTU in this model, with the focus on eight genes implicated in neural network formation or synaptic functions, such as the brain-derived neurotrophic factor (BDNF) and NMDA receptors 2A/2B. First, we measured the developmental profile of gene expression in vehicle-dosed rat cerebellum by quantitative RT-PCR and then examined the effects of PTU on mRNA levels on postnatal day (PND) 22, when most of the cerebellar structures in mature animals are already formed. PTU induced up-regulation of NR2B mRNA and down-regulation of NR2A and BDNF mRNAs in the cerebellum on PND 22, but there were no changes in the other genes (growth associated protein-43, L1, neuronal cell adhesion molecule, synaptophysin, post synaptic density-95). Examination of the effects of PTU on maturation of NMDAR subunits (NR2A/NR2B) demonstrated changes in relative expression on PND 14, but not on PND 4, with recovery after maturation. The profile of NMDAR subunits in vehicle-dosed rats showed a shift from NR2B to NR2A during development. These results suggest PTU can delay this switching from NR2B to NR2A subunits in the maturation of NMDA receptors.

  11. Crystal structure of catalytic domain of the initiation factor 2B epsilon subunit

    DEFF Research Database (Denmark)

    Boesen, Thomas; Mohammad, Sarah S.; Pavitt, Graham D.;

    this motif is involved in binding to the N-terminal part of the eIF2β subunit The aliphatic residues in the AA box motifs are involved in specific contacts in the hydrophobic core of the C-terminal helices important for maintaining the overall structure, whereas acidic residues in the motifs form a clustered......-terminal two helices contain the catalytic part of the domain, whereas the C-terminal six helices harbor the two Aromatic Acidic (AA) box motifs. This motif is also found in initiation factor 5, the GTPase activator protein of eIF2, and furthermore in mammalian initiation factor 4G. In eIF2B and eIF5......, surface exposed acidic patch which might interact with the lysine boxes of eIF2β. Interestingly, Tryptophan 699 was found to be solvent exposed and involved in crystal packing. This residue could possibly be important for the specific interaction with eIF2β. Furthermore, the structure shows the location...

  12. NR2B subunit in the prefrontal cortex: A double-edged sword for working memory function and psychiatric disorders

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    Monaco, Sarah A.; Gulchina, Yelena; Gao, Wen-Jun

    2015-01-01

    The prefrontal cortex (PFC) is a brain region featured with working memory function. The exact mechanism of how working memory operates within the PFC circuitry is unknown, but persistent neuronal firing recorded from prefrontal neurons during a working memory task is proposed to be the neural correlate of this mnemonic encoding. The PFC appears to be specialized for sustaining persistent firing, with N-methyl-D-aspartate (NMDA) receptors, especially slow-decay NR2B subunits, playing an essential role in the maintenance of sustained activity and normal working memory function. However, the NR2B subunit serves as a double-edged sword for PFC function. Because of its slow kinetics, NR2B endows the PFC with not only “neural psychic” properties, but also susceptibilities for neuroexcitotoxicity and psychiatric disorders. This review aims to clarify the interplay among working memory, the PFC, and NMDA receptors; demonstrate the importance of the NR2B subunit in the maintenance of persistent activity; understand the risks and vulnerabilities of how NR2B is related to the development of neuropsychiatric disorders; identify gaps that currently exist in our understanding of these processes; and provide insights regarding future directions that may clarify these issues. We conclude that the PFC is a specialized brain region with distinct delayed maturation, unique neuronal circuitry, and characteristic NMDA receptor function. The unique properties and development of NMDA receptors, especially enrichment of NR2B subunits, endows the PFC with not only the capability to generate sustained activity for working memory, but also serves as a major vulnerability to environmental insults and risk factors for psychiatric disorders. PMID:26143512

  13. Kalirin Binds the NR2B Subunit of the NMDA Receptor, Altering Its Synaptic Localization and Function

    KAUST Repository

    Kiraly, D. D.

    2011-08-31

    The ability of dendritic spines to change size and shape rapidly is critical in modulating synaptic strength; these morphological changes are dependent upon rearrangements of the actin cytoskeleton. Kalirin-7 (Kal7), a Rho guanine nucleotide exchange factor localized to the postsynaptic density (PSD), modulates dendritic spine morphology in vitro and in vivo. Kal7 activates Rac and interacts with several PSD proteins, including PSD-95, DISC-1, AF-6, and Arf6. Mice genetically lacking Kal7 (Kal7KO) exhibit deficient hippocampal long-term potentiation (LTP) as well as behavioral abnormalities in models of addiction and learning. Purified PSDs from Kal7KO mice contain diminished levels of NR2B, an NMDA receptor subunit that plays a critical role in LTP induction. Here we demonstrate that Kal7KO animals have decreased levels of NR2B-dependent NMDA receptor currents in cortical pyramidal neurons as well as a specific deficit in cell surface expression of NR2B. Additionally, we demonstrate that the genotypic differences in conditioned place preference and passive avoidance learning seen in Kal7KO mice are abrogated when animals are treated with an NR2B-specific antagonist during conditioning. Finally, we identify a stable interaction between the pleckstrin homology domain of Kal7 and the juxtamembrane region of NR2B preceding its cytosolic C-terminal domain. Binding of NR2B to a protein that modulates the actin cytoskeleton is important, as NMDA receptors require actin integrity for synaptic localization and function. These studies demonstrate a novel and functionally important interaction between the NR2B subunit of the NMDA receptor and Kalirin, proteins known to be essential for normal synaptic plasticity.

  14. Structure of the gene encoding the murine protein kinase CK2 beta subunit

    DEFF Research Database (Denmark)

    Boldyreff, B; Issinger, O G

    1995-01-01

    The mouse protein kinase CK2 beta subunit gene (Csnk2b) is composed of seven exons contained within 7874 bp. The exon and intron lengths extend from 76 to 321 and 111 to 1272 bp, respectively. The lengths of the murine coding exons correspond exactly to the lengths of the exons in the human CK2...

  15. Copy Number Variation of UGT 2B Genes in Indian Families Using Whole Genome Scans

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    Avinash M. Veerappa

    2016-01-01

    Full Text Available Background and Objectives. Uridine diphospho-glucuronosyltransferase 2B (UGT2B is a family of genes involved in metabolizing steroid hormones and several other xenobiotics. These UGT2B genes are highly polymorphic in nature and have distinct polymorphisms associated with specific regions around the globe. Copy number variations (CNVs status of UGT2B17 in Indian population is not known and their disease associations have been inconclusive. It was therefore of interest to investigate the CNV profile of UGT2B genes. Methods. We investigated the presence of CNVs in UGT2B genes in 31 members from eight Indian families using Affymetrix Genome-Wide Human SNP Array 6.0 chip. Results. Our data revealed >50% of the study members carried CNVs in UGT2B genes, of which 76% showed deletion polymorphism. CNVs were observed more in UGT2B17 (76.4% than in UGT2B15 (17.6%. Molecular network and pathway analysis found enrichment related to steroid metabolic process, carboxylesterase activity, and sequence specific DNA binding. Interpretation and Conclusion. We report the presence of UGT2B gene deletion and duplication polymorphisms in Indian families. Network analysis indicates the substitutive role of other possible genes in the UGT activity. The CNVs of UGT2B genes are very common in individuals indicating that the effect is neutral in causing any suspected diseases.

  16. N-methyl-D-aspartate receptor NR2B subunit involved in depression-like behaviours in lithium chloride-pilocarpine chronic rat epilepsy model.

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    Peng, Wei-Feng; Ding, Jing; Li, Xin; Fan, Fan; Zhang, Qian-Qian; Wang, Xin

    2016-01-01

    Depression is a common comorbidity in patients with epilepsy with unclear mechanisms. This study is to explore the role of glutamate N-methyl-D-aspartate (NMDA) receptor NR1, NR2A and NR2B subunits in epilepsy-associated depression. Lithium chloride (Licl)-pilocarpine chronic rat epilepsy model was established and rats were divided into epilepsy with depression (EWD) and epilepsy without depression (EWND) subgroups based on forced swim test. Expression of NMDA receptor NR1, NR2A and NR2B subunits was measured by western blot and immunofluorescence methods. The immobility time (IMT) was significantly greater in Licl-pilocarpine model group than in Control group, which was also greater in EWD group than in EWND group. No differences of spontaneous recurrent seizure (SRS) counts over two weeks and latency were found between EWD and EWND groups. The number of NeuN positive cells was significantly less in Licl-pilocarpine model group than in Control group, but had no difference between EWD and EWND groups. The ratios of phosphorylated NR1 (p-NR1)/NR1 and p-NR2B/NR2B were significantly greater in the hippocampus in EWD group than in EWND group. Moreover, the expression of p-NR1 and p-NR2B in the CA1 subfield of hippocampus were both greater in Licl-pilocarpine model group than Control group. Selective blockage of NR2B subunit with ifenprodil could alleviate depression-like behaviours of Licl-pilocarpine rat epilepsy model. In conclusion, glutamate NMDA receptor NR2B subunit was involved in promoting depression-like behaviours in the Licl-pilocarpine chronic rat epilepsy model and might be a target for treating epilepsy-associated depression.

  17. The selectivity of conantokin-G for ion channel inhibition of NR2B subunit-containing NMDA receptors is regulated by amino acid residues in the S2 region of NR2B.

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    Sheng, Zhenyu; Liang, Zhong; Geiger, James H; Prorok, Mary; Castellino, Francis J

    2009-08-01

    The conantokins are short, naturally occurring peptides that inhibit ion flow through N-methyl-d-aspartate receptor (NMDAR) channels. One member of this peptide family, conantokin-G (con-G), shows high selectivity for antagonism of NR2B-containing NMDAR channels, whereas other known conantokins are less selective inhibitors with regard to the nature of the NR2 subunit of the NMDAR complex. In order to define the molecular determinants of NR2B that govern con-G selectivity, we evaluated the ability of con-G to inhibit NMDAR ion channels expressed in human embryonic kidney (HEK)293 cells transfected with NR1, in combination with various NR2A/2B chimeras and point mutants, by electrophysiology using cells voltage-clamped in the whole-cell configuration. We found that a variant of the con-G-insensitive subunit, NR2A, in which the 158 residues comprising the S2 peptide segment (E(657)-I(814)) were replaced by the corresponding S2 region of NR2B (E(658)-I(815)), results in receptors that are highly sensitive to inhibition by con-G. Of the 22 amino acids that are different between the NR2A-S2 and the NR2B-S2 regions, exchange of one of these, M(739) of NR2B for the equivalent K(738) of NR2A, was sufficient to completely import the inhibitory activity of con-G into NR1b/NR2A-containing NMDARs. Some reinforcement of this effect was found by substitution of a second amino acid, K(755) of NR2B for Y(754) of NR2A. The discovery of the molecular determinants of NR2B selectivity with con-G has implications for the design of subunit-selective neurobiological probes and drug therapies, in addition to advancing our understanding of NR2B- versus NR2A-mediated neurological processes.

  18. Leptin reverses corticosterone-induced inhibition of neural stem cell proliferation through activating the NR2B subunits of NMDA receptors

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    Shi, Wen-Zhu [Anesthesia and Operation Center, Hainan Branch of Chinese PLA General Hospital, Hainan 572013 (China); Anesthesia and Operation Center, Chinese PLA General Hospital, Beijing 100853 (China); Miao, Yu-Liang [Department of Anesthesiology, PLA No. 306 Hospital, Beijing 100101 (China); Guo, Wen-Zhi [Department of Anesthesiology, Beijing Military General Hospital of Chinese People’s Liberation Army, Beijing 100700 (China); Wu, Wei, E-mail: wwzwgk@163.com [Department of Head and Neck Surgery of Otolaryngology, PLA No. 306 Hospital, Beijing 100101 (China); Li, Bao-Wei [Department of Head and Neck Surgery of Otolaryngology, PLA No. 306 Hospital, Beijing 100101 (China); An, Li-Na [Department of Anesthesiology, Armed Police General Hospital, Beijing 100039 (China); Fang, Wei-Wu [Department of Anesthesiology, PLA No. 306 Hospital, Beijing 100101 (China); Mi, Wei-Dong, E-mail: elite2005gg@163.com [Anesthesia and Operation Center, Chinese PLA General Hospital, Beijing 100853 (China)

    2014-04-25

    Highlights: • Leptin promotes the proliferation of neural stem cells isolated from embryonic mouse hippocampus. • Leptin reverses corticosterone-induced inhibition of neural stem cell proliferation. • The effects of leptin are partially mediated by upregulating NR2B subunits. - Abstract: Corticosterone inhibits the proliferation of hippocampal neural stem cells (NSCs). The removal of corticosterone-induced inhibition of NSCs proliferation has been reported to contribute to neural regeneration. Leptin has been shown to regulate brain development, improve angiogenesis, and promote neural regeneration; however, its effects on corticosterone-induced inhibition of NSCs proliferation remain unclear. Here we reported that leptin significantly promoted the proliferation of hippocampal NSCs in a concentration-dependent pattern. Also, leptin efficiently reversed the inhibition of NSCs proliferation induced by corticosterone. Interestingly, pre-treatment with non-specific NMDA antagonist MK-801, specific NR2B antagonist Ro 25-6981, or small interfering RNA (siRNA) targeting NR2B, significantly blocked the effect of leptin on corticosterone-induced inhibition of NSCs proliferation. Furthermore, corticosterone significantly reduced the protein expression of NR2B, whereas pre-treatment with leptin greatly reversed the attenuation of NR2B expression caused by corticosterone in cultured hippocampal NSCs. Our findings demonstrate that leptin reverses the corticosterone-induced inhibition of NSCs proliferation. This process is, at least partially mediated by increased expression of NR2B subunits of NMDA receptors.

  19. Gene targeting of CK2 catalytic subunits

    Science.gov (United States)

    Lou, David Y.; Toselli, Paul; Landesman-Bollag, Esther; Dominguez, Isabel

    2013-01-01

    Protein kinase CK2 is a highly conserved and ubiquitous serine–threonine kinase. It is a tetrameric enzyme that is made up of two regulatory CK2β subunits and two catalytic subunits, either CK2α/CK2α, CK2α/ CK2α′, or CK2α′/CK2α′. Although the two catalytic subunits diverge in their C termini, their enzymatic activities are similar. To identify the specific function of the two catalytic subunits in development, we have deleted them individually from the mouse genome by homologous recombination. We have previously reported that CK2α′is essential for male germ cell development, and we now demonstrate that CK2α has an essential role in embryogenesis, as mice lacking CK2α die in mid-embryogenesis, with cardiac and neural tube defects. PMID:18594950

  20. Gene targeting of CK2 catalytic subunits.

    Science.gov (United States)

    Seldin, David C; Lou, David Y; Toselli, Paul; Landesman-Bollag, Esther; Dominguez, Isabel

    2008-09-01

    Protein kinase CK2 is a highly conserved and ubiquitous serine-threonine kinase. It is a tetrameric enzyme that is made up of two regulatory CK2beta subunits and two catalytic subunits, either CK2alpha/CK2alpha, CK2alpha/CK2alpha', or CK2alpha'/CK2alpha'. Although the two catalytic subunits diverge in their C termini, their enzymatic activities are similar. To identify the specific function of the two catalytic subunits in development, we have deleted them individually from the mouse genome by homologous recombination. We have previously reported that CK2alpha' is essential for male germ cell development, and we now demonstrate that CK2alpha has an essential role in embryogenesis, as mice lacking CK2alpha die in mid-embryogenesis, with cardiac and neural tube defects.

  1. Propofol effectively inhibits lithium-pilocarpine-induced status epilepticus in rats via downregulation of N-methyl-D-aspartate receptor 2B subunit expression

    Institute of Scientific and Technical Information of China (English)

    Henglin Wang; Zhuoqiang Wang; Weidong Mi; Cong Zhao; Yanqin Liu; Yongan Wang; Haipeng Sun

    2012-01-01

    Status epilepticus was induced via intraperitoneal injection of lithium-pilocarpine. The inhibitory ef-fects of propofol on status epilepticus in rats were judged based on observation of behavior, elec-troencephalography and 24-hour survival rate. Propofol (12.5-100 mg/kg) improved status epilep-ticus in a dose-dependent manner, and significantly reduced the number of deaths within 24 hours of lithium-pilocarpine injection. Western blot results showed that, 24 hours after induction of status epilepticus, the levels of N-methyl-D-aspartate receptor 2A and 2B subunits were significantly in-creased in rat cerebral cortex and hippocampus. Propofol at 50 mg/kg significantly suppressed the increase in N-methyl-D-aspartate receptor 2B subunit levels, but not the increase in N-methyl-D-aspartate receptor 2A subunit levels. The results suggest that propofol can effectively inhibit status epilepticus induced by lithium-pilocarpine. This effect may be associated with down-regulation of N-methyl-D-aspartate receptor 2B subunit expression after seizures.

  2. Competitive binding of postsynaptic density 95 and Ca2+-calmodulin dependent protein kinase Ⅱ to N-methyl-D-aspartate receptor subunit 2B in rat brain

    Institute of Scientific and Technical Information of China (English)

    Fan-jie MENG; Jun GUO; Bo SONG; Xue-bo YAN; Guang-yi ZHANG

    2004-01-01

    AIM: To investigate the interactions among postsynaptic density 95 (PSD-95), Ca2+-calmodulin dependent protein kinase Ⅱα (CaMKⅡα), and N-methyl-D-aspartate receptor subunit 2B (NR2B) during ischemia and reperfusion in hippocampus of rats. METHODS: Brain ischemia was induced by four-vessel occlusion procedure in rats. Immunoprecipitation and immunoblotting were performed to study the interactions and phosphorylation of proteins. The association-dissociation of PSD-95 and CaMKⅡα to and from N-methyl-D-aspartate (NMDA) receptor induced by ischemia and reperfusion and the effects of 1-[N,O-bis-(5-isoquinolinesulfonyl)-N-methyl-L-tyrosyl]-4-phenyl-piperazine (KN-62, a selective inhibitor of CaMKⅡ) on these protein interactions were investigated. Coimmunoprecipitation and immunoblotting were performed for the studies of interactions among proteins. RESULTS: The alternations of the binding level of PSD-95 and CaMKⅡα to NR2B during ischemia and reperfusion demonstrated the negative correlation to each other. Pre-administration of KN62 through both cerebral ventricles inhibited the 10 min ischemia-induced increase of the binding of PSD-95 to NR2B and, on the contrary, promoted the binding of CaMKⅡα to NR2B. CONCLUSION: PSD-95 competes with CaMKⅡ to bind to NR2B during ischemia and reperfusion in rat hippocampus.

  3. Gene silencing of NR2B-containing NMDA receptor by intrathecal injection of short hairpin RNA reduces formalin-induced nociception in C57BL/6 mouse.

    Science.gov (United States)

    Zhang, Rao-Xiang; Yan, Xue-Bin; Gu, Yong-Hong; Huang, Dong; Gan, Li; Han, Rui; Huang, Li-Hua

    2013-09-01

    Spinal NR2B-containing N-methyl-D-aspartate receptors (NR2B) play a critical role in the formation of central sensitization and persistent pain. Previous studies show that gene silencing of the spinal NR2B subunit by small interfering RNA (siRNA) could alleviate nociception in animals. The siRNA is a 19- to 23-nt RNA duplex, which can be synthesized in vitro or derived from short hairpin RNAs (shRNAs). In the present study, we investigated whether intrathecal injection of shRNAs targeting NR2B (GRIN2B shRNA) could affect nociception on formalin-induced pain in mice. Our results showed that intrathecal injection of GRIN2B shRNA could decrease NR2B mRNA and protein expression levels and hence effectively relieve formalin-induced nociception in mice, suggesting that intrathecal delivery of GRIN2B shRNA can be an efficient way to silence the target gene and provide new insights into the treatment of chronic pain.

  4. Genes for resistance to stripe rust on chromosome 2B and their application in wheat breeding

    Institute of Scientific and Technical Information of China (English)

    Peigao Luo; Xueyun Hu; Huaiyu Zhang; Zhenglong Ren

    2009-01-01

    Stripe rust,caused by Puccinia striiformis f.sp.tritici,is one of the most damaging diseases of wheat worldwide.Growing resistant cultivars is the most economic and environmental friendly way to control the disease.There are many resistance genes to stripe rust located on wheat chromosome 2B.Here,we propose a strategy to construct the recombinant wheat chromosome 2B with multiple resistances to stripe rust by making crosses between wheat lines or cultivars carrying Yr genes and using marker-assisted selection,based on the reported information about resistance spectrum,chromosomal location,and linked markers of the genes.Pyramiding the resistance genes on 2B would afford a valuable strategy to control the disease by cultivating varieties with durable resistance.The possibility,efficiency,and prospect of the suggested strategy are reviewed in the paper.

  5. Tyrosine phosphorylation of the N-Methyl-D-Aspartate receptor 2B subunit in spinal cord contributes to remifentanil-induced postoperative hyperalgesia: the preventive effect of ketamine

    Directory of Open Access Journals (Sweden)

    Cui Songqin

    2009-12-01

    Full Text Available Abstract Background Experimental and clinical studies showed that intraoperative infusionof remifentanil has been associated with postoperative hyperalgesia. Previous reports suggested that spinal N-methyl-D-aspartate (NMDA receptors may contribute to the development and maintenance of opioid-induced hyperalgesia. In the present study, we used a rat model of postoperative pain to investigate the role of tyrosine phosphorylation of NMDA receptor 2B (NR2B subunit in spinal cord in the postoperative hyperalgesia induced by remifentanil and the intervention of pretreatment with ketamine. Results Intraoperative infusion of remifentanil (0.04 mg/kg, subcutaneous significantly enhanced mechanical allodynia and thermal hyperalgesia induced by the plantar incision during the postoperative period (each lasting between 2 h and 48 h, which was attenuated by pretreatment with ketamine (10 mg/kg, subcutaneous. Correlated with the pain behavior changes, immunocytochemical and western blotting experiments in our study revealed that there was a marked increase in NR2B phosphorylation at Tyr1472 in the superficial dorsal horn after intraoperative infusion of remifentanil, which was attenuated by pretreatment with ketamine. Conclusions This study provides direct evidence that tyrosine phosphorylation of the NR2B at Tyr1472 in spinal dosal horn contributes to postoperative hyperalgesia induced by remifentanil and supports the potential therapeutic value of ketamine for improving postoperative hyperalgesia induced by remifentanil.

  6. Crystal structure of the catalytic domain of the initiation factor 2B epsilon subunit from saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Boesen, Thomas; Pavitt, Graham D.; Andersen, Gregers Rom

    -terminal domain harbors the two aa-box motifs involved in binding to the N-terminal part of the initiation factor 2 β subunit. Aliphatic residues in the aa-box motifs are involved in specific contacts in the hydrophobic core of the C-terminal bundle important for maintaining the overall structure, whereas, acidic...... residues in the motifs form a surface exposed acidic patch which might interact with the lysine boxes of initiation factor 2 β. Interestingly, tryptophan 699 was found to be solvent exposed and involved in crystal packing. This residue could possibly be important for the specific interaction...

  7. Mapping of the Mouse Actin Capping Protein Beta Subunit Gene

    Directory of Open Access Journals (Sweden)

    Cooper John A

    2000-07-01

    Full Text Available Abstract Background Capping protein (CP, a heterodimer of α and β subunits, is found in all eukaryotes. CP binds to the barbed ends of actin filaments in vitro and controls actin assembly and cell motility in vivo. Vertebrates have three isoforms of CPβ produced by alternatively splicing from one gene; lower organisms have one gene and one isoform. Results We isolated genomic clones corresponding to the β subunit of mouse CP and identified its chromosomal location by interspecies backcross mapping. Conclusions The CPβ gene (Cappb1 mapped to Chromosome 4 between Cdc42 and D4Mit312. Three mouse mutations, snubnose, curly tail, and cribriform degeneration, map in the vicinity of the β gene.

  8. No evidence for a role of Ile587Val polymorphism of EIF2B5 gene in multiple sclerosis in Kashmir Valley of India.

    Science.gov (United States)

    Zahoor, Insha; Asimi, Ravouf; Haq, Ehtishamul

    2015-12-15

    Multiple sclerosis (MS) is an inflammatory neurodegenerative disease of the nervous system with a profound genetic element. It is already known that alterations in Eukaryotic Translation Initiation Factor 2B (EIF2B) gene encoding the five subunits of eIF2B complex cause Vanishing White Matter (VWM) disease of the brain and emerging evidences have advocated certain resemblances between MS and VWM in terms of clinical and epidemiological characteristics, thus validating the association study between EIF2B and MS. Moreover, a recent study has implicated EIF2B5 Ile587Val (rs843358) polymorphism as a susceptibility factor for MS. In order to investigate the association of EIF2B5 Ile587Val polymorphism with MS susceptibility in Kashmir region in India, we screened EIF2B5 Exon 13 in 30 MS patients and 65 controls (a total of 95 participants). During the present course of study, we could not find statistically significant difference in the frequency of Ile587Val between MS patients and controls, thus indicating that such alteration does not appear to influence MS development in Kashmiri population. Our results provide evidence against a major role for Ile587Val polymorphism in MS susceptibility.

  9. MOLECULAR CLONING AND HETEROLOGOUS EXPRESSION OF HUMAN INTERFERON ALPHA2b GENE

    Directory of Open Access Journals (Sweden)

    I. Made Artika

    2013-01-01

    Full Text Available Human alpha Interferons (hIFNα have been shown to have antiviral, antiproliferative and immunomodulatory activities. The human interferon alpha2b (hIFNα2b, is one of the human interferon alpha2 sub variants, naturally synthesized as a polypeptide of 188 amino acid residues, the first 23 residues of which represents a signal peptide. In the present study, the hIFNα2b gene was expressed after being fused with Glutathione S-Transferase (GST gene. The hIFNα2b gene was amplified from human genomic DNA by using a pair of specific primers, cloned into an Escherichia coli expression vector and expressed in E. coli cells under the direction of the tac promoter. The expressed protein was purified using a one-step affinity chromatography column containing immobilized gluthatione-bound resin. The purified protein was shown to react specifically with anti-human-interferon-alpha antibody, confirming that the protein was the human interferon alpha molecule. This strategy has the potential to be used as an alternative mean for production of pure human interferon α proteins for therapeutic purposes and for further studies on their molecular characterization and mechanism of action.

  10. Closely linked H2B genes in the marine copepod, Tigriopus californicus indicate a recent gene duplication or gene conversion event.

    Science.gov (United States)

    Brown, D; Cook, A; Wagner, M; Wells, D

    1992-01-01

    Two nonallelic histone gene clusters were characterized in the marine copepod, Tigriopus californicus. The DNA sequence of one of the clusters reveals six genes in the contiguous arrangement of H2B, H1, H3, H4, H2B and H2A. The order of genes within the second cluster is H3, H4, H2B and H2A. There is no evidence for the presence of an H1 gene in this cluster. Comparison of the three copepod H2B genes reveals a high degree of similarity between the 5' upstream regions and between the amino terminal halves of the two H2B genes found within the same cluster. From these data we infer that gene duplication and/or gene conversion events occurred within this cluster in the recent past.

  11. Characterization of the gene for the a subunit of human factor XIII (plasma transglutaminase), a blood coagulation factor

    Energy Technology Data Exchange (ETDEWEB)

    Ichinose, A.; Davie, E.W. (Univ. of Washington, Seattle (USA))

    1988-08-01

    Factor XIII (plasma transglutaminase, fibrin stabilizing factor) is a glycoprotein that circulates in blood as a tetramer (a{sub 2}b{sub 2}) consisting of two a and two b subunits. The primary structures of the a and b subunits of human factor XIII have been reported by a combination of cDNA cloning and amino acid sequence analysis. To establish the gene structure of the a subunit for factor XIII, several human genomic libraries were screened by using the cDNA encoding the a subunit as a probe. Among {approx}5 {times} 10{sup 7} recombinant phage, 121 have been shown to contain an insert encoding a portion of the a subunit. Twenty-five unique clones were than characterized by restriction mapping, Southern blotting, and DNA sequencing. Overlapping clones encoding the a subunit of factor XIII span >160 kilobases. DNA sequence analysis revealed that the activation peptide released by thrombin, the active site cysteine region, the two putative calcium-binding regions, and the thrombin cleavage site leading to inactivation are encoded by separate exons. This suggest that the introns may separate the a subunit into functional and structural domains. A comparison of the amino acid sequence deduced from the genomic DNA sequence with those deduced from cDNA or determined by amino acid sequence analysis of the plasma and placental proteins revealed apparent amino acid polymorphisms in six positions of the polypeptide chain of the a subunit.

  12. Immune Response in Calves Vaccinated with Type Three Secretion System Antigens and Shiga Toxin 2B Subunit of Escherichia coli O157:H7

    Science.gov (United States)

    Martorelli, Luisina; Garbaccio, Sergio; Vilte, Daniel A.; Albanese, Adriana A.; Mejías, María P.; Palermo, Marina S.; Mercado, Elsa C.; Ibarra, Cristina E.; Cataldi, Angel A.

    2017-01-01

    Ruminants are the primary reservoir of Shiga-toxin producing Escherichia coli (STEC) O157:H7 and the main source of infection for humans. The aim of this study was to assess the immunogenic properties of a candidate vaccine consisting on the recombinant proteins of E. coli O157:H7 IntiminC280, the carboxy-terminal fraction of Intimin γ, EspB and the fusion protein between the B subunit of Stx2 and Brucella Lumazine Synthase (BLS)(BLS-Stx2B), in Holstein Fresian calves.To accomplish this goal we vaccinated calves with two doses of different vaccine formulations: 2 antigens (IntiminC280, EspB), 3 antigens (IntiminC280, EspB, BLS-Stx2B), BLS-Stx2B alone and a control non-vaccinated group. All antigens were expressed as recombinant proteins in E. coli. Specific IgG titres increased in vaccinated calves and the inclusion of BLS-Stx2B in the formulation seems to have a stimulatory effect on the humoral response to IntiminC280 and EspB after the booster. The neutralizing activity of antibodies against these two antigens was assessed in Red Blood Cell lysis assays and adherence to Hep-2 cells as a correlate of T3SS activity. Both sera from animals vaccinated with 2 or 3 antigens inhibited both virulence properties. Serological response to Stx2 was observed in animals vaccinated only with BLS-Stx2B and with 3 antigens and neutralization of Stx2 cytotoxicity was also observed in both groups. In conclusion, immunization of calves with BLS-Stx2B, IntiminC280 and EspB elicited a potent humoral response able to neutralize Shiga toxin 2 cytotoxity and the T3SS virulence properties in vitro. These results suggest that this formulation is a good candidate vaccine to reduce STEC shedding in cattle and needs to be further assessed in vivo. PMID:28046078

  13. Inhibition of human UGT2B7 gene expression in transgenic mice by the constitutive androstane receptor.

    Science.gov (United States)

    Yueh, M F; Mellon, P L; Tukey, R H

    2011-06-01

    The xenobiotic receptors, constitutive androstane receptor (CAR), and pregnane X receptor (PXR) regulate and alter the metabolism of xenobiotic substrates. Among the 19 functional UDP-glucuronosyltransferases (UGTs) in humans, UGT2B7 is involved in the metabolism of many structurally diverse xenobiotics and plays an important role in the clearance and detoxification of many therapeutic drugs. To examine whether this gene is regulated by CAR and PXR in vivo, transgenic mice expressing the entire UGT2B7 gene (TgUGT2B7) were created. Gene expression profiles revealed that UGT2B7 is differentially expressed in liver, kidney, adipocytes, brain, and estrogen-sensitive tissues, such as ovary and uterus. Liver UGT2B7 expression levels were decreased when TgUGT2B7 mice were treated with the CAR ligand 1,4-b-s-[2-(3,5,-dichloropyridyloxy)] (TCPOBOP) but not the PXR ligand pregnenolone 16α-carbonitrile. Although TCPOBOP decreased the levels of UGT2B7 mRNA in TgUGT2B7 mice, it had no affect on Tg(UGT2B7)Car(-/-) mice, adding support for a CAR-dependent mechanism contributing toward UGT2B7 gene suppression. Expression of promoter constructs in HepG2 cells showed the CAR-dependent inhibition was linked to hepatocyte nuclear factor-4α (HNF4α)-mediated transactivation of the UGT2B7 promoter. The inhibitory effect of CAR on UGT2B7 gene expression was validated in chromatin immunoprecipitation assays in which TCPOBOP treatment blocked HNF4α binding to the UGT2B7 promoter. These results suggest that HNF4α plays an important role in the constitutive expression of hepatic UGT2B7, and CAR acts as a negative regulator by interfering with HNF4α binding activity.

  14. CONSTRUCTION AND STUDY OF Althaea officinalis TRANSGENIC ROOTS CULTURE WITH HUMAN INTERFERON α2B GENE

    Directory of Open Access Journals (Sweden)

    N. A. Matvieieva

    2013-04-01

    Full Text Available The aim of our work was to obtain Althaea officinalis L. «hairy» root culture with human interferon α2b gene (ifn-α2b, to measure fructans content and antiviral activity of extracts from the transgenic roots. Transformation of leaf and root explants was carried out by means of Agrobacterium rhizogenes-mediated transformation. Antiviral activity was measured by the reduction in cytopathic effect of vesicular stomatitis virus (Indiana strain in bovine kidney cells line MDBK. Transformation frequency was 100% for leaf and root explants. RT-PCR confirmed ifn- α2b gene transcription. The clones of transgenic roots differed in mass increasing from 0, 036 ± 0,008 up to 0,371 ± 0,019 g in 30 days cultivation and in fructan synthesis from 67,2± 4,47 up to 154,6 ± 6,62 mg/g roots dry weight. Extracts from «hairy»roots culture were characterized by high antiviral activity against vesicular stomatitis virus — up to 26 000 IU/ g of roots fresh weight. In some cases the genetic transformation shown to lead increasing the growth rate and increasing the level of fructan synthesis in transgenic A. officinalis roots. Extracts from cultivated in vitro marshmallow transgenic roots were characterized by high level of antiviral activity against vesicular stomatitis virus. Thus, there were obtained transgenic A. officinalis roots, characterized by high growth rate, significant accumulation of fructans and high antiviral activity.

  15. Differential expression of G protein alpha and ß subunit genes during development of Phytophthora infestans

    NARCIS (Netherlands)

    Laxalt, A.M.; Latijnhouwers, M.; Hulten, van M.; Govers, F.

    2002-01-01

    A G protein subunit gene (pigpa1) and a G protein subunit gene (pigpb1) were isolated from the oomycete Phytophthora infestans, the causal agent of potato late blight. Heterotrimeric G proteins are evolutionary conserved GTP-binding proteins that are composed of ,, and subunits and participate in di

  16. Fyn Kinase regulates GluN2B subunit-dominant NMDA receptors in human induced pluripotent stem cell-derived neurons.

    Science.gov (United States)

    Zhang, Wen-Bo; Ross, P Joel; Tu, YuShan; Wang, Yongqian; Beggs, Simon; Sengar, Ameet S; Ellis, James; Salter, Michael W

    2016-04-04

    NMDA receptor (NMDAR)-mediated fast excitatory neurotransmission is implicated in a broad range of physiological and pathological processes in the mammalian central nervous system. The function and regulation of NMDARs have been extensively studied in neurons from rodents and other non-human species, and in recombinant expression systems. Here, we investigated human NMDARs in situ by using neurons produced by directed differentiation of human induced pluripotent stem cells (iPSCs). The resultant cells showed electrophysiological characteristics demonstrating that they are bona fide neurons. In particular, human iPSC-derived neurons expressed functional ligand-gated ion channels, including NMDARs, AMPA receptors, GABAA receptors, as well as glycine receptors. Pharmacological and electrophysiological properties of NMDAR-mediated currents indicated that these were dominated by receptors containing GluN2B subunits. The NMDAR currents were suppressed by genistein, a broad-spectrum tyrosine kinase inhibitor. The NMDAR currents were also inhibited by a Fyn-interfering peptide, Fyn(39-57), but not a Src-interfering peptide, Src(40-58). Together, these findings are the first evidence that tyrosine phosphorylation regulates the function of NMDARs in human iPSC-derived neurons. Our findings provide a basis for utilizing human iPSC-derived neurons in screening for drugs targeting NMDARs in neurological disorders.

  17. Expansion of transducin subunit gene families in early vertebrate tetraploidizations.

    Science.gov (United States)

    Lagman, David; Sundström, Görel; Ocampo Daza, Daniel; Abalo, Xesús M; Larhammar, Dan

    2012-10-01

    Hundreds of gene families expanded in the early vertebrate tetraploidizations including many gene families in the phototransduction cascade. We have investigated the evolution of the heterotrimeric G-proteins of photoreceptors, the transducins, in relation to these events using both phylogenetic analyses and synteny comparisons. Three alpha subunit genes were identified in amniotes and the coelacanth, GNAT1-3; two of these were identified in amphibians and teleost fish, GNAT1 and GNAT2. Most tetrapods have four beta genes, GNB1-4, and teleosts have additional duplicates. Finally, three gamma genes were identified in mammals, GNGT1, GNG11 and GNGT2. Of these, GNGT1 and GNGT2 were found in the other vertebrates. In frog and zebrafish additional duplicates of GNGT2 were identified. Our analyses show all three transducin families expanded during the early vertebrate tetraploidizations and the beta and gamma families gained additional copies in the teleost-specific genome duplication. This suggests that the tetraploidizations contributed to visual specialisations.

  18. Functional characterization of cadmium-responsive garlic gene AsMT2b: A new member of metallothionein family

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    A new gene of metallothionein (MT) family was cloned from garlic (Allium sativum) seedlings using RACE method and designated AsMT2b. The full length of AsMT2b cDNA was 520 bp encoding 80 amino acids. The deduced amino acids of AsMT2b showed that AsMT2b contained the characteristic structure of type 2 MT proteins, but the number and arrangement of the cysteine residues in the N- and C-terminal domains was different from other type 2 MT proteins. Semi-quantitative reverse transcriptase-PCR showed that transcript levels of AsMT2b were enhanced only in response to higher concentrations or longer incubation time of Cd. Such an expression pattern of AsMT2b greatly differed from that of other type 2 MT genes. Yeast cells transformed with this gene had improved resistance to Cd. AsMT2b overexpressing Arabidopsis showed stronger Cd tolerance and higher Cd accumulation compared with wild-type plants. These results suggest that AsMT2b should be useful in phytoremediation of Cd-polluted soil in the future.

  19. Dopamine receptor D5 deficiency results in a selective reduction of hippocampal NMDA receptor subunit NR2B expression and impaired memory.

    Science.gov (United States)

    Moraga-Amaro, Rodrigo; González, Hugo; Ugalde, Valentina; Donoso-Ramos, Juan Pablo; Quintana-Donoso, Daisy; Lara, Marcelo; Morales, Bernardo; Rojas, Patricio; Pacheco, Rodrigo; Stehberg, Jimmy

    2016-04-01

    Pharmacological evidence associates type I dopamine receptors, including subtypes D1 and D5, with learning and memory. Analyses using genetic approaches have determined the relative contribution of dopamine receptor D1 (D1R) in cognitive tasks. However, the lack of drugs that can discriminate between D1R and D5R has made the pharmacological distinction between the two receptors difficult. Here, we aimed to determine the role of D5R in learning and memory. In this study we tested D5R knockout mice and wild-type littermates in a battery of behavioral tests, including memory, attention, locomotion, anxiety and motivational evaluations. Our results show that genetic deficiency of D5R significantly impairs performance in the Morris water maze paradigm, object location and object recognition memory, indicating a relevant role for D5R in spatial memory and recognition memory. Moreover, the lack of D5R resulted in decreased exploration and locomotion. In contrast, D5R deficiency had no impact on working memory, anxiety and depressive-like behavior, measured using the spontaneous alternation, open-field, tail suspension test, and forced swimming test. Electrophysiological analyses performed on hippocampal slices showed impairment in long-term-potentiation in mice lacking D5R. Further analyses at the molecular level showed that genetic deficiency of D5R results in a strong and selective reduction in the expression of the NMDA receptor subunit NR2B in the hippocampus. These findings demonstrate the relevant contribution of D5R in memory and suggest a functional interaction of D5R with hippocampal glutamatergic pathways.

  20. Relationship between human evolution and neurally mediated syncope disclosed by the polymorphic sites of the adrenergic receptor gene α2B-AR.

    Directory of Open Access Journals (Sweden)

    Tomoyoshi Komiyama

    Full Text Available The objective of this study was to clarify the effects of disease on neurally mediated syncope (NMS during an acute stress reaction. We analyzed the mechanism of the molecular interaction and the polymorphisms of the alpha-2 adrenoreceptor (α2B-AR gene as the potential psychiatric cause of incentive stress. We focused on the following three genotypes of the repeat polymorphism site at Glu 301-303 in the α2B-AR gene: Glu12/12, Glu12/9, and Glu9/9. On the basis of our clinical research, NMS is likely to occur in people with the Glu12/9 heterotype. To verify this, we assessed this relationship with the interaction of Gi protein and adenylate cyclase by in silico analysis of the Glu12/9 heterotype. By measuring the difference in the dissociation time of the Gi-α subunit twice, we found that the Glu12/9 heterotype suppressed the action of adenylate cyclase longer than the Glu homotypes. As this difference in the Glu repeat number effect is thought to be one of the causes of NMS, we investigated the evolutionary significance of the Glu repeat number. Glu8 was originally repeated in simians, while the Glu12 repeats occurred over time during the evolution of bipedalism in humans. Taken with the Glu12 numbers, NMS would likely become a defensive measure to prevent significant blood flow to the human brain.

  1. Screening for coding variants in FTO and SH2B1 genes in Chinese patients with obesity.

    Directory of Open Access Journals (Sweden)

    Zhaojing Zheng

    Full Text Available OBJECTIVE: To investigate potential functional variants in FTO and SH2B1 genes among Chinese children with obesity. METHODS: Sanger sequencing of PCR products of all FTO and SH2B1 exons and their flanking regions were performed in 338 Chinese Han children with obesity and 221 age- and sex-matched lean controls. RESULTS: A total of seven and five rare non-synonymous variants were identified in FTO and SH2B1, respectively. The overall frequencies of FTO and SH2B1 rare non-synonymous variants were similar in obese and lean children (2.37% and 0.90% vs. 1.81% and 1.36%, P>0.05. However, four out of the seven variants in FTO were novel and all were unique to obese children (p>0.05. None of the novel variants was consistently being predicted to be deleterious. Four out of five variants in SH2B1 were novel and one was unique to obese children (p>0.05. One variant (L293R that was consistently being predicted as deleterious in SH2B1 gene was unique to lean control. While rare missense mutations were more frequently detected in girls from obesity as well as lean control than boys, the difference was not statistically significant. In addition, it's shown that the prevalence of rare missense mutations of FTO as well as SH2B1 was similar across different ethnic groups. CONCLUSION: The rare missense mutations of FTO and SH2B1 did not confer risks of obesity in Chinese Han children in our cohort.

  2. Screening for Coding Variants in FTO and SH2B1 Genes in Chinese Patients with Obesity

    Science.gov (United States)

    Huang, Xiaodong; Yang, Peirong; Li, Juan; Ding, Yu; Yao, Ru-en; Geng, Juan; Shen, Yongnian; Shen, Yiping; Fu, Qihua; Yu, Yongguo

    2013-01-01

    Objective To investigate potential functional variants in FTO and SH2B1 genes among Chinese children with obesity. Methods Sanger sequencing of PCR products of all FTO and SH2B1 exons and their flanking regions were performed in 338 Chinese Han children with obesity and 221 age- and sex-matched lean controls. Results A total of seven and five rare non-synonymous variants were identified in FTO and SH2B1, respectively. The overall frequencies of FTO and SH2B1 rare non-synonymous variants were similar in obese and lean children (2.37% and 0.90% vs. 1.81% and 1.36%, P>0.05). However, four out of the seven variants in FTO were novel and all were unique to obese children (p>0.05). None of the novel variants was consistently being predicted to be deleterious. Four out of five variants in SH2B1 were novel and one was unique to obese children (p>0.05). One variant (L293R) that was consistently being predicted as deleterious in SH2B1 gene was unique to lean control. While rare missense mutations were more frequently detected in girls from obesity as well as lean control than boys, the difference was not statistically significant. In addition, it's shown that the prevalence of rare missense mutations of FTO as well as SH2B1 was similar across different ethnic groups. Conclusion The rare missense mutations of FTO and SH2B1 did not confer risks of obesity in Chinese Han children in our cohort. PMID:23825611

  3. A Special Extract of Bacopa monnieri (CDRI-08 Restores Learning and Memory by Upregulating Expression of the NMDA Receptor Subunit GluN2B in the Brain of Scopolamine-Induced Amnesic Mice

    Directory of Open Access Journals (Sweden)

    Rakesh Rai

    2015-01-01

    Full Text Available In the present communication, we have investigated effects of the CDRI-08, a well characterized extract of Bacopa monnieri, on expression of the GluN2B subunit of NMDAR in various brain regions of the scopolamine-induced amnesic mice. Our behavioral data reveal that scopolamine-treated amnesic mice exhibit significant decline in the spatial memory compared to the normal control mice. Our RT-PCR and immunoblotting data revealed that the scopolamine treatment resulted in a significant downregulation of the NMDAR GluN2B subunit expression in prefrontal cortex and hippocampus. Our enzyme assay data revealed that scopolamine caused a significant increase in the acetylcholinesterase activity in both the brain regions. Further, oral administration of the CDRI-08 to scopolamine-treated amnesic mice restored the spatial memory which was found to be associated with significant upregulation of the GluN2B subunit expression and decline in the acetylcholinesterase activity in prefrontal cortex as well as hippocampus towards their levels in the normal control mice. Our study provides the evidence for the mechanism underlying role of the Bacopa monnieri extract (CDRI-08 in restoring spatial memory in amnesic mice, which may have therapeutic implications.

  4. Identification of PPAP2B as a novel recurrent translocation partner gene of HMGA2 in lipomas.

    Science.gov (United States)

    Bianchini, Laurence; Birtwisle, Loïc; Saâda, Esma; Bazin, Audrey; Long, Elodie; Roussel, Jean-François; Michiels, Jean-François; Forest, Fabien; Dani, Christian; Myklebost, Ola; Birtwisle-Peyrottes, Isabelle; Pedeutour, Florence

    2013-06-01

    Most lipomas are characterized by translocations involving the HMGA2 gene in 12q14.3. These rearrangements lead to the fusion of HMGA2 with an ectopic sequence from the translocation chromosome partner. Only five fusion partners of HMGA2 have been identified in lipomas so far. The identification of novel fusion partners of HMGA2 is important not only for diagnosis in soft tissue tumors but also because these genes might have an oncogenic role in other tumors. We observed that t(1;12)(p32;q14) was the second most frequent translocation in our series of lipomas after t(3;12)(q28;q14.3). We detected overexpression of HMGA2 mRNA and protein in all t(1;12)(p32;q14) lipomas. We used a fluorescence in situ hybridization-based positional cloning strategy to characterize the 1p32 breakpoint. In 11 cases, we identified PPAP2B, a member of the lipid phosphate phosphatases family as the 1p32 target gene. Reverse transcription-polymerase chain reaction analysis followed by nucleotide sequencing of the fusion transcript indicated that HMGA2 3' untranslated region (3'UTR) fused with exon 6 of PPAP2B in one case. In other t(1;12) cases, the breakpoint was extragenic, located in the 3'region flanking PPAP2B 3'UTR. Moreover, in one case showing a t(1;6)(p32;p21) we observed a rearrangement of PPAP2B and HMGA1, which suggests that HMGA1 might also be a fusion partner for PPAP2B. Our results also revealed that adipocytic differentiation of human mesenchymal stem cells derived from adipose tissue was associated with a significant decrease in PPAP2B mRNA expression suggesting that PPAP2B might play a role in adipogenesis.

  5. Promoter of soybean early nodulin gene enod2B is induced by rhizobial Nod factors in transgenic rice

    Institute of Scientific and Technical Information of China (English)

    WANG Yanzhang; YU Guanqiao; SHEN Shanjiong(San Chiun Shen); ZHU Jiabi

    2004-01-01

    Nod factors, which are signaling molecules produced by Rhizobia, are the principal determinants of host specificity in Rhizobium-legume symbiosis. Nod factors can elicit a number of characteristic developmental responses in the roots of legumes, such as depolarization of the membrane potential in epidermal cells, specific expression of early nodulin genes and changes in the flux of calcium in root hairs, deformation of root hairs, cell division in the root cortex and formation of the nodule primordium. Whether the rice plant can respond to signaling molecules (I.e. Nod factors) is an important question, as it could establish the potential for symbiotic nitrogen fixation in rice. The promoter of the soybean (Glycine max) early nodulin gene Gmenod2B fused to the β-glucuronidase (GUS) reporter gene was used as a molecular marker to explore whether Nod factors can be recognized by rice cells as signaling molecules. Transgenic rice plants harboring the chimeric gene Gmenod2BP-GUS were obtained via an Agrobacterium tumefaciens-mediated system. NodNGR factors produced by a broad-host-range Rhizobium strain NGR234(pA28) were used as probes to investigate the activity of the Gmenod2B promoter in rice. Our results showed that the early nodulin gene Gmenod2B promoter was induced by NodNGR factors in transgenic rice, and that it was specifically expressed in rice plant roots. Moreover, GUS gene expression driven by the Gmenod2B promoter in transgenic rice was regulated by nitrogen status. These findings indicated that rice possessed the ability to respond to Nod factor signals, and that this signal transduction system resulted in activation of the Gmenod2B promoter. Thus, we predict that the Nod-factor inducible nodulin expression system, which is similar to Rhizobium-legume symbiosis, may also exist in rice.

  6. Involvement of the GluN2A and GluN2B subunits in synaptic and extrasynaptic N-methyl-D-aspartate receptor function and neuronal excitotoxicity.

    Science.gov (United States)

    Zhou, Xianju; Ding, Qi; Chen, Zhuoyou; Yun, Huifang; Wang, Hongbing

    2013-08-16

    GluN2A and GluN2B are the major subunits of functional NMDA receptors (NMDAR). Previous studies have suggested that GluN2A and GluN2B may differentially mediate NMDAR function at synaptic and extrasynaptic locations and play opposing roles in excitotoxicity, such as neurodegeneration triggered by ischemic stroke and brain injury. By using pharmacological and molecular approaches to suppress or enhance the function of GluN2A and GluN2B in cultured cortical neurons, we examined NMDAR-mediated, bidirectional regulation of prosurvival signaling (i.e. the cAMP response element-binding protein (CREB)-Bdnf cascade) and cell death. Inhibition of GluN2A or GluN2B attenuated the up-regulation of prosurvival signaling triggered by the activation of either synaptic or extrasynaptic NMDAR. Inhibition of GluN2A or GluN2B also attenuated the down-regulation of prosurvival signaling triggered by the coactivation of synaptic and extrasynaptic receptors. The effects of GluN2B on CREB-Bdnf signaling were larger than those of GluN2A. Consistently, compared with suppression of GluN2A, suppression of GluN2B resulted in more reduction of NMDA- and oxygen glucose deprivation-induced excitotoxicity as well as NMDAR-mediated elevation of intracellular calcium. Moreover, excitotoxicity and down-regulation of CREB were exaggerated in neurons overexpressing GluN2A or GluN2B. Together, we found that GluN2A and GluN2B are involved in the function of both synaptic and extrasynaptic NMDAR, demonstrating that they play similar rather than opposing roles in NMDAR-mediated bidirectional regulation of prosurvival signaling and neuronal death.

  7. Cholinergic cells in the nucleus basalis of mice express the N-methyl-D-aspartate-receptor subunit NR2C and its replacement by the NR2B subunit enhances frontal and amygdaloid acetylcholine levels

    NARCIS (Netherlands)

    De Souza Silva, M. A.; Dolga, Amalia; Pieri, I.; Marchetti, L.; Eisel, U. L. M.; Huston, J. P.; Dere, E.

    2006-01-01

    It is known that glutamatergic and cholinergic systems interact functionally at the level of the cholinergic basal forebrain. The N-methyl-D-aspartate receptor (NMDA-R) is a multiprotein complex composed of NR1, NR2 and/or NR3 subunits. The subunit composition of NMDA-R of cholinergic cells in the n

  8. Significant prognostic values of nuclear genes encoding mitochondrial complex I subunits in tumor patients.

    Science.gov (United States)

    Li, L D; Sun, H F; Bai, Y; Gao, S P; Jiang, H L; Jin, W

    2016-01-01

    In cancer biology, it remains still open question concerning the oncogenic versus oncosuppressor behavior of metabolic genes, which includes those encoding mitochondrial complex I (CI) subunits. The prognostic value of nuclear genome mRNAs expression of CI subunits is to be evaluated in the tumor patients. We used the Kaplan Meier plotter database, the cBio Cancer Genomics Portal, and the Oncomine in which gene expression data and survival information were from thousands of tumor patients to assess the relevance of nuclear genome mRNAs level of CI subunits to patients' survival, as well as their alterations in gene and expression level in tumors. We presented that the relative expression level of overwhelming majority of the nuclear genes of CI subunits with survival significance (overall survival, relapse free survival, progression free survival, distant metastasis free survival, post progression survival, and first progression), had consistent effects for patients in each type of four tumors separately, including breast cancer, ovarian cancer, lung cancer, and gastric cancer. However, in gene level, frequent cumulative or individual alteration of these genes could not significantly affect patients' survival and the overexpression of the individual gene was not ubiquitous in tumors versus normal tissues. Given that reprogrammed energy metabolism was viewed as an emerging hallmark of tumor, thus tumor patients' survival might potentially to be evaluated by certain threshold for overall expression of CI subunits. Comprehensive understanding of the nuclear genome encoded CI subunits may have guiding significance for the diagnosis and prognosis in tumor patients.

  9. Mapping of a liver phosphorylase kinase [alpha]-subunit gene on the mouse x chromosome

    Energy Technology Data Exchange (ETDEWEB)

    Geng, Yan; Derry, J.M.J.; Barnard, P.J. (MRC Molecular Neurobiology Unit, Cambridge (United Kingdom)); Hendrickx, J.; Coucke, P.; Willems, P.R. (Univ. of Antwerp (Belgium))

    1993-01-01

    Phosphorylase kinase (PHK) is a regulatory enzyme of the glycogenolytic pathway composed of a complex of four subunits. We recently mapped the muscle [alpha]-subunit gene (Phka) to the mouse X chromosome in a region syntenic with the proximal long arm of the human X chromosome and containing the human homologue of this gene, PHKA. We now report the mapping of the liver [alpha]-subunit gene to the telomeric end of the mouse X chromosome. This mapping position would suggest a location for the human liver [alpha]-subunit gene on the proximal short arm of the X chromosome, a region recently implicated in X-linked liver glycogenosis (XLG). 20 refs., 2 figs.

  10. Genes involved in protein metabolism of the probiotic lactic acid bacterium Lactobacillus delbrueckii UFV H2b20.

    Science.gov (United States)

    Do Carmo, A P; da Silva, D F; De Oliveira, M N V; Borges, A C; De Carvalho, A F; De Moraes, C A

    2011-09-01

    A basic requirement for the prediction of the potential use of lactic acid bacteria (LAB) in the dairy industry is the identification of specific genes involved in flavour-forming pathways. The probiotic Lactobacillus delbrueckii UFV H2b20 was submitted to a genetic characterisation and phylogenetic analysis of genes involved in protein catabolism. Eight genes belonging to this system were identified, which possess a closely phylogenetic relationship to NCFM strains representative, as it was demonstrated for oppC and oppBII, encoding oligopeptide transport system components. PepC, PepN, and PepX might be essential for growth of LAB, probiotic or not, since the correspondent genes are always present, including in L. delbrueckii UFV H2b20 genome. For pepX gene, a probable link between carbohydrate catabolism and PepX expression may exists, where it is regulated by PepR1/CcpA-like, a common feature between Lactobacillus strains and also in L. delbrueckii UFV H2b20. The well conserved evolutionary history of the ilvE gene is evidence that the pathways leading to branched-chain amino acid degradation, such as isoleucine and valine, are similar among L. delbrueckii subsp. bulgaricus strains and L. delbrueckii UFV H2b20. Thus, the involvement of succinate in flavour formation can be attributed to IlvE activity. The presence of aminopeptidase G in L. delbrueckii UFV H2b20 genome, which is absent in several strains, might improve the proteolytic activity and effectiveness. The nucleotide sequence encoding PepG revealed that it is a cysteine endopeptidase, belonging to Peptidase C1 superfamily; sequence analysis showed 99% identity with L. delbrueckii subsp. bulgaricus ATCC 11842 pepG, whereas protein sequence analysis revealed 100% similarity with PepG from the same organism. The present study proposes a schematic model to explain how the proteolytic system of the probiotic L. delbrueckii UFV H2b20 works, based on the components identified so far.

  11. Role of the NR2A/2B subunits of the N-methyl-D-aspartate receptor in glutamate-induced glutamic acid decarboxylase alteration in cortical GABAergic neurons in vitro.

    Science.gov (United States)

    Monnerie, H; Hsu, F-C; Coulter, D A; Le Roux, P D

    2010-12-29

    The vulnerability of brain neuronal cell subpopulations to neurologic insults varies greatly. Among cells that survive a pathological insult, for example ischemia or brain trauma, some may undergo morphological and/or biochemical changes that may compromise brain function. The present study is a follow-up of our previous studies that investigated the effect of glutamate-induced excitotoxicity on the GABA synthesizing enzyme glutamic acid decarboxylase (GAD65/67)'s expression in surviving DIV 11 cortical GABAergic neurons in vitro [Monnerie and Le Roux, (2007) Exp Neurol 205:367-382, (2008) Exp Neurol 213:145-153]. An N-methyl-D-aspartate receptor (NMDAR)-mediated decrease in GAD expression was found following glutamate exposure. Here we examined which NMDAR subtype(s) mediated the glutamate-induced change in GAD protein levels. Western blotting techniques on cortical neuron cultures showed that glutamate's effect on GAD proteins was not altered by NR2B-containing diheteromeric (NR1/NR2B) receptor blockade. By contrast, blockade of triheteromeric (NR1/NR2A/NR2B) receptors fully protected against a decrease in GAD protein levels following glutamate exposure. When receptor location on the postsynaptic membrane was examined, extrasynaptic NMDAR stimulation was observed to be sufficient to decrease GAD protein levels similar to that observed after glutamate bath application. Blocking diheteromeric receptors prevented glutamate's effect on GAD proteins after extrasynaptic NMDAR stimulation. Finally, NR2B subunit examination with site-specific antibodies demonstrated a glutamate-induced, calpain-mediated alteration in NR2B expression. These results suggest that glutamate-induced excitotoxic NMDAR stimulation in cultured GABAergic cortical neurons depends upon subunit composition and receptor location (synaptic vs. extrasynaptic) on the neuronal membrane. Biochemical alterations in surviving cortical GABAergic neurons in various disease states may contribute to the altered

  12. Sequence and expression of the CAPA/CAP2b gene in the tobacco hawkmoth, Manduca sexta.

    Science.gov (United States)

    Loi, Poh Kheng; Tublitz, Nathan J

    2004-10-01

    The gene coding for cardioacceleratory peptide 2b (CAP2b; pELYAFPRV) has been isolated and sequenced from the moth Manduca sexta (GenBank accession #AY649544). Because of its significant homology to the CAPA gene in Drosophila melanogaster, this gene is called the Manduca CAPA gene. The Manduca CAPA gene is 958 nucleotides long with 29 untranslated nucleotides from the beginning of the sequence to the putative start initiation site. The CAPA gene has a single open reading frame, 441 nucleotides long, that codes for a predicted precursor protein of 147 amino acids. The predicted prepropeptide encodes a single copy of each of three deduced propeptides, a CAP2b propeptide, with a Q substituted for an E at the N-terminus (QLYAFPRVa), and two novel CAP2b-related propeptides (DGVLNLYPFPRVa and TEGPGMWFGPRLa). To reduce confusion and to adopt a more standardized nomenclature, we rename pELYAFPRVa as Mas-CAPA-1 and assign the names of Mas-CAPA-2 to DGVLNLYPFPRVa and Mas-PK-1 (Pyrokinin-1) to TEGPGMWFGPRLa. The spatial and temporal expression pattern of the CAPA gene in the Manduca central nervous system (CNS) was determined in all major post-embryonic stages using in situ hybridization techniques. The CAPA gene is expressed in a total of 27 pairs of neurons in the post-embryonic Manduca CNS. A total of 16 pairs of cells is observed in the brain, two pairs in the sub-esophageal ganglion (SEG), one pair in the third thoracic ganglion (T3), one pair in each unfused abdominal ganglion (A1-A6) and two pairs in the fused terminal ganglion. The mRNA from the CAPA gene is present in nearly every ganglion in each post-embryonic stage. The number of cells expressing the CAPA gene varies during post-embryonic life, starting at 54 cells in first-instar larvae and declining to a minimum of 14 cells midway through adult development.

  13. Activation of glycine site and GluN2B subunit of NMDA receptors is necessary for ERK/CREB signaling cascade in rostral anterior cingulate cortex in rats: Implications for affective pain

    Institute of Scientific and Technical Information of China (English)

    Hong Cao; Wen-Hua Ren; Mu-Ye Zhu; Zhi-Qi Zhao; Yu-Qiu Zhang

    2012-01-01

    Objective The rostral anterior cingulate cortex (rACC) is implicated in processing the emotional component of pain.N-methyl-D-aspartate receptors (NMDARs) are highly expressed in the rACC and mediate painrelated affect by activating a signaling pathway that involves cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA) and/or extracellular regulated kinase (ERK)/cAMP-response element-binding protein (CREB).The present study investigated the contributions of the NMDAR glycine site and GluN2B subunit to the activation of ERK and CREB both in vitro and in vivo in rat rACC.Methods Immunohistochemistry and Western blot analysis were used to separately assess the expression of phospho-ERK (pERK) and phospho-CREB (pCREB) in vitro and in vivo.Double immunostaining was also used to determine the colocalization of pERK and pCREB.Results Both bath application of NMDA in brain slices in vitro and intraplantar injection of formalin into the rat hindpaw in vivo induced significant up-regulation of pERK and pCREB in the rACC,which was inhibited by the NMDAR antagonist DL-2-amino-5-phospho-novaleric acid.Selective blockade of the NMDAR GluN2B subunit and the glycinebinding site,or degradation of endogenous D-serine,a co-agonist for the glycine site,significantly decreased the upregulation of pERK and pCREB expression in the rACC.Further,the activated ERK predominantly colocalized with CREB.Conclusion Either the glycine site or the GluN2B subunit of NMDARs participates in the phosphorylation of ERK and CREB induced by bath application of NMDA in brain slices or hindpaw injection of 5% formalin in rats,and these might be fundamental molecular mechanisms underlying pain affect.

  14. The SHORT-ROOT-like gene PtSHR2B is involved in Populus phellogen activity.

    Science.gov (United States)

    Miguel, Andreia; Milhinhos, Ana; Novák, Ondřej; Jones, Brian; Miguel, Célia M

    2016-03-01

    SHORT-ROOT (SHR) is a GRAS transcription factor first characterized for its role in the specification of the stem cell niche and radial patterning in Arabidopsis thaliana (At) roots. Three SHR-like genes have been identified in Populus trichocarpa (Pt). PtSHR1 shares high similarity with AtSHR over the entire length of the coding sequence. The two other Populus SHR-like genes, PtSHR2A and PtSHR2B, are shorter in their 5' ends when compared with AtSHR. Unlike PtSHR1, that is expressed throughout the cambial zone of greenhouse-grown Populus trees, PtSHR2Bprom:uidA expression was detected in the phellogen. Additionally, PtSHR1 and PtSHR2B expression patterns markedly differ in the shoot apex and roots of in vitro plants. Transgenic hybrid aspen expressing PtSHR2B under the 35S constitutive promoter showed overall reduced tree growth while the proportion of bark increased relative to the wood. Reverse transcription-quantitative PCR (RT-qPCR) revealed increased transcript levels of cytokinin metabolism and response-related genes in the transgenic plants consistent with an increase of total cytokinin levels. This was confirmed by cytokinin quantification by LC-MS/MS. Our results indicate that PtSHR2B appears to function in the phellogen and therefore in the regulation of phellem and periderm formation, possibly acting through modulation of cytokinin homeostasis. Furthermore, this work points to a functional diversification of SHR after the divergence of the Populus and Arabidopsis lineages. This finding may contribute to selection and breeding strategies of cork oak in which, unlike Populus, the phellogen is active throughout the entire tree lifespan, being at the basis of a highly profitable cork industry.

  15. Nuclear respiratory factor 2 regulates the expression of the same NMDA receptor subunit genes as NRF-1: both factors act by a concurrent and parallel mechanism to couple energy metabolism and synaptic transmission.

    Science.gov (United States)

    Priya, Anusha; Johar, Kaid; Wong-Riley, Margaret T T

    2013-01-01

    Neuronal activity and energy metabolism are tightly coupled processes. Previously, we found that nuclear respiratory factor 1 (NRF-1) transcriptionally co-regulates energy metabolism and neuronal activity by regulating all 13 subunits of the critical energy generating enzyme, cytochrome c oxidase (COX), as well as N-methyl-d-aspartate (NMDA) receptor subunits 1 and 2B, GluN1 (Grin1) and GluN2B (Grin2b). We also found that another transcription factor, nuclear respiratory factor 2 (NRF-2 or GA-binding protein) regulates all subunits of COX as well. The goal of the present study was to test our hypothesis that NRF-2 also regulates specific subunits of NMDA receptors, and that it functions with NRF-1 via one of three mechanisms: complementary, concurrent and parallel, or a combination of complementary and concurrent/parallel. By means of multiple approaches, including in silico analysis, electrophoretic mobility shift and supershift assays, in vivo chromatin immunoprecipitation of mouse neuroblastoma cells and rat visual cortical tissue, promoter mutations, real-time quantitative PCR, and western blot analysis, NRF-2 was found to functionally regulate Grin1 and Grin2b genes, but not any other NMDA subunit genes. Grin1 and Grin2b transcripts were up-regulated by depolarizing KCl, but silencing of NRF-2 prevented this up-regulation. On the other hand, over-expression of NRF-2 rescued the down-regulation of these subunits by the impulse blocker TTX. NRF-2 binding sites on Grin1 and Grin2b are conserved among species. Our data indicate that NRF-2 and NRF-1 operate in a concurrent and parallel manner in mediating the tight coupling between energy metabolism and neuronal activity at the molecular level.

  16. Biodistribution and elimination kinetics of systemic Stx2 by the Stx2A and Stx2B subunit-specific human monoclonal antibodies in mice

    Directory of Open Access Journals (Sweden)

    Sheoran Abhineet

    2012-06-01

    Full Text Available Abstract Background Hemolytic uremic syndrome (HUS leading to acute kidney failure, is a condition linked to the production of primarily Shiga toxin 2 (Stx2 by some E. coli serotypes. We have previously shown that Stx2 A subunit-specific human monoclonal antibody (HuMAb 5C12, and B subunit-specific HuMAb 5H8 inhibit cultured cell death, and protect mice and piglets from fatal Stx2-intoxication. We have also shown that 5H8 blocks binding of Stx2 to its cell-surface receptor globotriaosyl ceramide (Gb3, whereas Stx2 when complexed with 5C12 binds Gb3 with higher affinity than Stx2. The mechanism by which 5C12 neutralizes Stx2 in vitro involves trapping of Stx2 in the recycling endosomes and releasing it into the extracellular environment. Because of the clinical implications associated with the formation of Stx2/antibody complexes and the potential for trapping and clearance through a severely damaged kidney associated with HUS, we investigated the likely site(s of Stx2/antibody localization and clearance in intoxicated mice treated with antibody or placebo. Results Mice were injected with radiolabeled Stx2 (125I-Stx2 4 hours after administration of 5C12, 5H8, or phosphate buffered saline (PBS and the sites of localization of labeled Stx2, were investigated 3, 24 and 48 hours later. The liver recorded statistically much higher concentrations of labeled Stx2 for groups receiving 5C12 and 5H8 antibodies after 3, 24 and 48 hours, as compared with the PBS group. In contrast, highest levels of labeled Stx2 were detected in the kidneys of the PBS group at all 3 sampling times. Mice receiving either of the two HuMAbs were fully protected against the lethal effect of Stx2, as compared with the fatal outcome of the control group. Conclusions The results suggest that HuMAbs 5C12 and 5H8 promoted hepatic accumulation and presumably clearance of toxin/antibody complexes, significantly diverting Stx2 localization in the kidneys, the target of Stx2 and the

  17. Piroxicam inhibits NMDA receptor-mediated excitotoxicity through allosteric inhibition of the GluN2B subunit: an in silico study elucidating a novel mechanism of action of the drug.

    Science.gov (United States)

    Mazumder, Muhammed Khairujjaman; Borah, Anupom

    2014-12-01

    Hyperactivation of GluN2B subunit containing N-methyl-d-aspartate receptors (NMDARs) significantly contributes to the development of several neurodegenerative diseases through a process called excitotoxicity. NMDARs are voltage-gated Ca2+ channels which when activated lead to excessive influx of Ca2+ into neurons thereby exacerbating several calcium-dependent pathways that cause oxidative stress and apoptosis. Several drugs are presently in use to counter the NMDAR-mediated excitotoxic events among which Ifenprodil and its derivatives are GluN2B selective allosteric antagonists. Certain non-steroidal anti-inflammatory drugs (NSAIDs) have also been reported to inhibit NMDARs and the resultant pathologies. Meanwhile, Piroxicam, which is a NSAID, has been reported to be protective in cerebral ischemia-induced neurodegeneration through various pathways. Since Piroxicam has more number of interacting groups as compared to other NSAIDs and also has structural similarities with Ifenprodil, we thought it prudent that Piroxicam may inhibit NMDARs similar to Ifenprodil. By using molecular docking as a tool, we validated the hypothesis and hereby report for the first time that Piroxicam can inhibit GluN2B containing NMDARs through allosteric mode similar to the well known selective antagonist--Ifenprodil; and thus can be a therapeutic drug for the prevention of excitotoxic neurodegeneration.

  18. Mutations in the histone methyltransferase gene KMT2B cause complex early-onset dystonia

    NARCIS (Netherlands)

    Meyer, Esther; Carss, Keren J.; Rankin, Julia; Nichols, John M. E.; Grozeva, Detelina; Joseph, Agnel P.; Mencacci, Niccolo E.; Papandreou, Apostolos; Ng, Joanne; Barra, Serena; Ngoh, Adeline; Ben-Pazi, Hilla; Willemsen, Michel A.; Arkadir, David; Barnicoat, Angela; Bergman, Hagai; Bhate, Sanjay; Boys, Amber; Darin, Niklas; Foulds, Nicola; Gutowski, Nicholas; Hills, Alison; Houlden, Henry; Hurst, Jane A.; Israe, Zvi; Kaminska, Margaret; Limousin, Patricia; Lumsden, Daniel; Mckee, Shane; Misra, Shibalik; Mohammed, Shekeeb S.; Nakou, Vasiliki; Nicolai, Joost; Nilsson, Magnus; Pall, Hardev; Peall, Kathryn J.; Peters, Gregory B.; Prabhakar, Prab; Reuter, Miriam S.; Rump, Patrick; Sege, Reeval; Sinnema, Margje; Smith, Martin; Turnpenny, Peter; White, Susan M.; Wieczorek, Dagmar; Wiethoff, Sarah; Wilson, Brian T.; Winter, Gidon; Wragg, Christopher; Pope, Simon; Heales, Simon J. H.; Morrogh, Deborah; Pittman, Alan; Carr, Lucinda J.; Perez-Duenas, Belen; Lin, Jean-Pierre; Reis, Andre; Gahl, William A.; Toro, Camilo; Bhatia, Kailash P.; Wood, Nicholas W.; Kamsteeg, Erik-Jan; Chong, Wui K.; Gissen, Paul; Topf, Maya; Dale, Russell C.; Chubby, Jonathan R.; Raymond, F. Lucy; Kurian, Manju A.

    2017-01-01

    Histone lysine methylation, mediated by mixed-lineage leukemia (MLL) proteins, is now known to be critical in the regulation of gene expression, genomic stability, cell cycle and nuclear architecture. Despite MLL proteins being postulated as essential for normal development, little is known about th

  19. ATXN2 and its neighbouring gene SH2B3 are associated with increased ALS risk in the Turkish population.

    Directory of Open Access Journals (Sweden)

    Suna Lahut

    Full Text Available Expansions of the polyglutamine (polyQ domain (≥ 34 in Ataxin-2 (ATXN2 are the primary cause of spinocerebellar ataxia type 2 (SCA2. Recent studies reported that intermediate-length (27-33 expansions increase the risk of Amyotrophic Lateral Sclerosis (ALS in 1-4% of cases in diverse populations. This study investigates the Turkish population with respect to ALS risk, genotyping 158 sporadic, 78 familial patients and 420 neurologically healthy controls. We re-assessed the effect of ATXN2 expansions and extended the analysis for the first time to cover the ATXN2 locus with 18 Single Nucleotide Polymorphisms (SNPs and their haplotypes. In accordance with other studies, our results confirmed that 31-32 polyQ repeats in the ATXN2 gene are associated with risk of developing ALS in 1.7% of the Turkish ALS cohort (p=0.0172. Additionally, a significant association of a 136 kb haplotype block across the ATXN2 and SH2B3 genes was found in 19.4% of a subset of our ALS cohort and in 10.1% of the controls (p=0.0057, OR: 2.23. ATXN2 and SH2B3 encode proteins that both interact with growth receptor tyrosine kinases. Our novel observations suggest that genotyping of SNPs at this locus may be useful for the study of ALS risk in a high percentage of individuals and that ATXN2 and SH2B3 variants may interact in modulating the disease pathway.

  20. Early transcription factor subunits are encoded by vaccinia virus late genes.

    Science.gov (United States)

    Gershon, P D; Moss, B

    1990-06-01

    The vaccinia virus early transcription factor (VETF) was shown to be a virus-encoded heterodimer. The gene for the 82-kDa subunit was identified as open reading frame (ORF) A8L, based on the N-terminal sequence of factor purified by using DNA-affinity magnetic beads. The 70-kDa subunit of VETF was refractory to N-terminal analysis, and so N-terminal sequences were obtained for three internal tryptic peptides. All three peptides matched sequences within ORF D6R. ORFs A8L and D6R are located within the central region of the vaccinia virus genome and are separated by about 13,600 base pairs. Proteins corresponding to the 3' ends of ORFs A8L and D6R were overexpressed in Escherichia coli and used to prepare antisera that bound to the larger and smaller subunits, respectively, of affinity-purified VETF. Immunoblot analysis of proteins from infected cells indicated that both subunits are expressed exclusively in the late phase of infection, just prior to their packaging in virus particles. The two subunits of VETF have no significant local or overall amino acid sequence homology to one another, to other entries in biological sequence data bases including bacterial sigma factors, or to recently determined sequences of some eukaryotic transcription factors. The 70-kDa subunit, however, has motifs in common with a super-family of established and putative DNA and RNA helicases.

  1. Mutations in genes encoding subunits of RNA polymerases I and III cause Treacher Collins syndrome.

    NARCIS (Netherlands)

    Dauwerse, J.G.; Dixon, J.; Seland, S.; Ruivenkamp, C.A.; Haeringen, A. van; Hoefsloot, L.H.; Peters, D.J.; Boers, A.C.; Daumer-Haas, C.; Maiwald, R.; Zweier, C.; Kerr, B.; Cobo, A.M.; Toral, J.F.; Hoogeboom, A.J.M.; Lohmann, D.R.; Hehr, U.; Dixon, M.J.; Breuning, M.H.; Wieczorek, D.

    2011-01-01

    We identified a deletion of a gene encoding a subunit of RNA polymerases I and III, POLR1D, in an individual with Treacher Collins syndrome (TCS). Subsequently, we detected 20 additional heterozygous mutations of POLR1D in 252 individuals with TCS. Furthermore, we discovered mutations in both allele

  2. Short hairpin RNA targeting 2B gene of coxsackievirus B3 exhibits potential antiviral effects both in vitro and in vivo

    Directory of Open Access Journals (Sweden)

    Yao Hailan

    2012-08-01

    Full Text Available Abstract Background Coxsackievirus B3 is an important infectious agent of viral myocarditis, pancreatitis and aseptic meningitis, but there are no specific antiviral therapeutic reagents in clinical use. RNA interference-based technology has been developed to prevent the viral infection. Methods To evaluate the impact of RNA interference on viral replication, cytopathogenicity and animal survival, short hairpin RNAs targeting the viral 2B region (shRNA-2B expressed by a recombinant vector (pGCL-2B or a recombinant lentivirus (Lenti-2B were tansfected in HeLa cells or transduced in mice infected with CVB3. Results ShRNA-2B exhibited a significant effect on inhibition of viral production in HeLa cells. Furthermore, shRNA-2B improved mouse survival rate, reduced the viral tissues titers and attenuated tissue damage compared with those of the shRNA-NC treated control group. Lenti-2B displayed more effective role in inhibition of viral replication than pGCL-2B in vivo. Conclusions Coxsackievirus B3 2B is an effective target of gene silencing against coxsackievirus B3 infection, suggesting that shRNA-2B is a potential agent for further development into a treatment for enterviral diseases.

  3. A common deletion in the uridine diphosphate glucuronyltransferase (UGT) 2B17 gene is a strong determinant of androgen excretion in healthy pubertal boys

    DEFF Research Database (Denmark)

    Juul, A; Sørensen, K; Aksglaede, L;

    2008-01-01

    BACKGROUND: Testosterone (T) is excreted in urine as water-soluble glucuronidated and sulfated conjugates. The ability to glucuronidate T and other steroids depends on a number of different glucuronidases (UGT) of which UGT2B17 is essential. The aim of the study was to evaluate the influence of UGT......2B17 genotypes on urinary excretion of androgen metabolites in pubertal boys. STUDY DESIGN: A clinical study of 116 healthy boys aged 8-19 yr. UGT2B17 genotyping was performed using quantitative PCR. Serum FSH, LH, T, estradiol (E2), and SHBG were analyzed by immunoassays, and urinary levels...... of androgen metabolites were quantitated by gas chromatography/mass spectrometry in all subjects. RESULTS: Ten of 116 subjects (9%) presented with a homozygote deletion of the UGT2B17 gene (del/del), whereas 52 and 54 boys were hetero- and homozygous carriers of the UGT2B17 gene (del/ins and ins...

  4. [Phenotypic variations in Aicardi-Goutieres syndrome caused by RNASEH2B gene mutations: report of two new cases].

    Science.gov (United States)

    Ortiz-Madinaveitia, Saturnino; Conejo-Moreno, David; López-Pisón, Javier; Peña-Segura, José Luis; Serrano-Madrid, M Luisa; Durán-Palacios, Ingrid C; Peláez-Cabo, Pilar

    2016-02-16

    Introduccion. El sindrome de Aicardi-Goutieres es un trastorno inmunitario raro debido a mutaciones en siete genes que codifican proteinas llamadas TREX1, el complejo ribonucleasa H2, SAMHD1, ADAR e IFIH1 (MAD5), las cuales estan implicadas en el metabolismo de los acidos nucleicos. A continuacion se presentan dos nuevos casos por mutacion en el gen RNASEH2B, uno de los cuales presenta una mutacion no descrita hasta la fecha. Casos clinicos. Caso 1: varon que consulto porque desde los 5 meses, coincidiendo con cuadros febriles de repeticion, presentaba perdida de los items madurativos adquiridos hasta la fecha. Caso 2: niño de 4 meses que desde los 2 meses mostraba gran irritabilidad con dificultades en la alimentacion, asociado a un grave retraso psicomotor. En ambos casos se constato un aumento de las pterinas en el liquido cefalorraquideo, principalmente de la neopterina, con calcificaciones en los ganglios basales. El diagnostico se confirmo mediante secuenciacion del gen RNASEH2B; el caso 2 presentaba una mutacion no descrita en la literatura medica. Conclusiones. Los casos corresponden a la descripcion clasica realizada por Aicardi-Goutieres. Debe tenerse en cuenta este sindrome ante un paciente con un cuadro de encefalopatia subaguda de comienzo en el primer año de vida, distonia/espasticidad en grado variable e importante afectacion/regresion del desarrollo psicomotor, especialmente si asocia aumento de las pterinas (neopterina) en el liquido cefalorraquideo y calcificaciones en los ganglios basales.

  5. The molecular prevalence and MSA-2b gene-based genetic diversity of Babesia bovis in dairy cattle in Thailand.

    Science.gov (United States)

    Simking, Pacharathon; Saengow, Sinsamuth; Bangphoomi, Kunan; Sarataphan, Nachai; Wongnarkpet, Sirichai; Inpankaew, Tawin; Jittapalapong, Sathaporn; Munkhjargal, Tserendorj; Sivakumar, Thillaiampalam; Yokoyama, Naoaki; Igarashi, Ikuo

    2013-11-08

    Bovine babesiosis is an economically significant disease that affects dairy farming operations in Thailand. In the present study, 1824 blood-DNA samples prepared from cattle bred in 4 different regions of the country (North, Northeast, Central, and South) were screened using a nested PCR for the specific detection of Babesia bovis. While the overall prevalence of B. bovis was 8.8%, the Central region of Thailand was found to be a high-risk area of the country, as the prevalence of the parasite was 15.0%. The positive rate was relatively higher among the animals of 1-5 years of age. The genetic diversity among the B. bovis parasites was also studied based on their MSA-2b gene, and the findings showed that the Thai sequences were dispersed across 8 of 13 total clades observed in the phylogram. Three of these clades were formed only of Thai sequences. Similarity among the deduced MSA-2b amino acid sequences determined in the present study was 68.3-100%. In conclusion, the present study found that all the locations surveyed were infected with B. bovis and that the parasite populations in Thailand were genetically diverse. Our findings highlight the need for further studies in Thailand to generate more information before a sound control strategy could be implemented against B. bovis.

  6. A heart-hand syndrome gene: Tfap2b plays a critical role in the development and remodeling of mouse ductus arteriosus and limb patterning.

    Directory of Open Access Journals (Sweden)

    Feng Zhao

    Full Text Available BACKGROUND: Patent ductus arteriosus (PDA is one of the most common forms of congenital heart disease. Mutations in transcription factor TFAP2B cause Char syndrome, a human disorder characterized by PDA, facial dysmorphysm and hand anomalies. Animal research data are needed to understand the mechanisms. The aim of our study was to elucidate the pathogenesis of Char syndrome at the molecular level. METHODOLOGY/PRINCIPAL FINDINGS: Gene expression of Tfap2b during mouse development was studied, and newborns of Tfap2b-deficient mice were examined to identify phenotypes. Gel shift assays had been carried out to search for Tfap2 downstream genes. Promoters of candidate genes were cloned into a reporter construct and used to demonstrate their regulation by Tfap2b in cell transfection. In situ hybridizations showed that the murine transcription factor Tfap2b was expressed during the entire development of mouse ductus arteriosus. Histological examination of ductus arteriosus from Tfap2b knockout mice 6 hours after birth revealed that they were not closed. Consequently, the lungs of Tfap2b(-/- mice demonstrated progressive congestion of the pulmonary capillaries, which was postulated to result secondarily from PDA. In addition, Tfap2b was expressed in the limb buds, particularly in the posterior limb field during development. Lack of Tfap2b resulted in bilateral postaxial accessory digits. Further study indicated that expressions of bone morphogenetic protein (Bmp genes, which are reported to be involved in the limb patterning and ductal development, were altered in limb buds of Tfap2b-deficient embryos, due to direct control of Bmp2 and Bmp4 promoter activity by Tfap2b. CONCLUSIONS/SIGNIFICANCE: Tfap2b plays important roles in the development of mouse ductus arteriosus and limb patterning. Loss of Tfap2b results in altered Bmp expression that may cause the heart-limb defects observed in Tfap2b mouse mutants and Char syndrome patients. The Tfap2b knockout

  7. STEADY-STATE TRANSCRIPT LEVELS OF CYTOCHROME-C-OXIDASE GENES DURING HUMAN MYOGENESIS INDICATE SUBUNIT SWITCHING OF SUBUNIT VIA AND COEXPRESSION OF SUBUNIT VIIA ISOFORMS

    NARCIS (Netherlands)

    TAANMAN, JW; HERZBERG, NH; DEVRIES, H; BOLHUIS, PA; VANDENBOGERT, C

    1992-01-01

    Steady-state levels of the mitochondrial rRNAs, of mRNAs for mitochondrially and nuclear-encoded subunits of cytochrome c oxidase and for the beta-subunit of ATP synthase were assessed by Northern blot hybridizations during the in vitro differentiation of human myoblasts. Transcript levels of the so

  8. Genetic variations of glycinin subunit genes among cultivated and wild type soybean species

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Glycinin is a predominant storage protein in most soybean accessions. It is a hexamer constituted by five major subunits, which can be classified into two groups. Group Ⅰ contains Gl, G2 and G3, and Group Ⅱ contains G4 and G5. The genes encoding these subunits have been designated from Gyl to Gy5, respectively. In the present study, Gyl genomic fragments were cloned from wild accessions of subgenera Glycine glycine, Glycine soja and a cultivar of Glycine max. Their sequences and the deduced amino acid sequences were compared. The residues critical for assembling of G1 subunits from the wild perennial accession were conservative. The Gy4 fragments were cloned from two wild perennial accessions and compared with that from subgenus Soja. The intron 3 of Gy4 had abundant variations between the subgenera G. Soja and G. Glycine as well as within the subgenus G. Glycine. Abundant variations existed in the disordered regions 3 and 4 of G4 subunits from two wild perennial accessions. The genomic organization of glycinin genes was analyzed in 19 accessions from subgenera Soja and Glycine. The hybridization patterns were identical among the accessions of subgenus Soja. On the contrary, abundant polymorphisms existed between the accessions from subgenus Glycine. These results indicated that glycinin genes have high degree of conservation within subgenus Soja but more variations within subgenus Glycine.

  9. Glutathione S-transferase Ya subunit gene: identification of regulatory elements required for basal level and inducible expression.

    OpenAIRE

    Telakowski-Hopkins, C A; King, R. G.; Pickett, C B

    1988-01-01

    The function of the 5'-flanking region of a rat glutathione S-transferase Ya subunit structural gene has been examined in homologous and heterologous cells. By using the 5'-flanking region of the Ya subunit gene fused to the structural gene encoding chloramphenicol acetyltransferase, we have identified two cis-acting regulatory elements in the upstream region of the Ya gene. One element is required for maximum basal level expression in homologous cells, whereas maximum basal level expression ...

  10. A remarkably stable TipE gene cluster: evolution of insect Para sodium channel auxiliary subunits

    Directory of Open Access Journals (Sweden)

    Li Jia

    2011-11-01

    Full Text Available Abstract Background First identified in fruit flies with temperature-sensitive paralysis phenotypes, the Drosophila melanogaster TipE locus encodes four voltage-gated sodium (NaV channel auxiliary subunits. This cluster of TipE-like genes on chromosome 3L, and a fifth family member on chromosome 3R, are important for the optional expression and functionality of the Para NaV channel but appear quite distinct from auxiliary subunits in vertebrates. Here, we exploited available arthropod genomic resources to trace the origin of TipE-like genes by mapping their evolutionary histories and examining their genomic architectures. Results We identified a remarkably conserved synteny block of TipE-like orthologues with well-maintained local gene arrangements from 21 insect species. Homologues in the water flea, Daphnia pulex, suggest an ancestral pancrustacean repertoire of four TipE-like genes; a subsequent gene duplication may have generated functional redundancy allowing gene losses in the silk moth and mosquitoes. Intronic nesting of the insect TipE gene cluster probably occurred following the divergence from crustaceans, but in the flour beetle and silk moth genomes the clusters apparently escaped from nesting. Across Pancrustacea, TipE gene family members have experienced intronic nesting, escape from nesting, retrotransposition, translocation, and gene loss events while generally maintaining their local gene neighbourhoods. D. melanogaster TipE-like genes exhibit coordinated spatial and temporal regulation of expression distinct from their host gene but well-correlated with their regulatory target, the Para NaV channel, suggesting that functional constraints may preserve the TipE gene cluster. We identified homology between TipE-like NaV channel regulators and vertebrate Slo-beta auxiliary subunits of big-conductance calcium-activated potassium (BKCa channels, which suggests that ion channel regulatory partners have evolved distinct lineage

  11. Stress-induced co-expression of two alternative oxidase (VuAox1 and 2b) genes in Vigna unguiculata.

    Science.gov (United States)

    Costa, José Hélio; Mota, Erika Freitas; Cambursano, Mariana Virginia; Lauxmann, Martin Alexander; de Oliveira, Luciana Maia Nogueira; Silva Lima, Maria da Guia; Orellano, Elena Graciela; Fernandes de Melo, Dirce

    2010-05-01

    Cowpea (Vigna unguiculata) alternative oxidase is encoded by a small multigene family (Aox1, 2a and 2b) that is orthologous to the soybean Aox family. Like most of the identified Aox genes in plants, VuAox1 and VuAox2 consist of 4 exons interrupted by 3 introns. Alignment of the orthologous Aox genes revealed high identity of exons and intron variability, which is more prevalent in Aox1. In order to determine Aox gene expression in V. unguiculata, a steady-state analysis of transcripts involved in seed development (flowers, pods and dry seeds) and germination (soaked seeds) was performed and systemic co-expression of VuAox1 and VuAox2b was observed during germination. The analysis of Aox transcripts in leaves from seedlings under different stress conditions (cold, PEG, salicylate and H2O2 revealed stress-induced co-expression of both VuAox genes. Transcripts of VuAox2a and 2b were detected in all control seedlings, which was not the case for VuAox1 mRNA. Estimation of the primary transcript lengths of V. unguiculata and soybean Aox genes showed an intron length reduction for VuAox1 and 2b, suggesting that the two genes have converged in transcribed sequence length. Indeed, a bioinformatics analysis of VuAox1 and 2b promoters revealed a conserved region related to a cis-element that is responsive to oxidative stress. Taken together, the data provide evidence for co-expression of Aox1 and Aox2b in response to stress and also during the early phase of seed germination. The dual nature of VuAox2b expression (constitutive and induced) suggests that the constitutive Aox2b gene of V. unguiculata has acquired inducible regulatory elements.

  12. Identification of compound heterozygous mutations in the ITGA2B gene in a Chinese patient with Glanzmann thrombasthenia

    Institute of Scientific and Technical Information of China (English)

    ZHENG Jia-yong; JIN Yan-hui; ZHU Yong-lin; JIN Pei-pei; ZHANG De-ting; JIN Zi-bing

    2010-01-01

    Background Glanzmann thrombasthenia (GT) is an autosomat recessive bleeding disorder characterized by the tendency to hemorrhage and the inability of platelets to aggregate in response to agonists. GT is caused by a defect of the platelet glycoprotein IIb/IIIa complex. The objective of this study was to describe the clinical features and the genetic cause of GT in a 6-year-old girl from south China.Methods A three-generation family was studied. The proband patient aged 6 years and her parents undertook examinations of platelet counts, blood film, bleeding time, platelet aggregation, and flow cytometry. All coding exons of the ITGA2B and ITGB3 genes were amplified by polymerase chain reaction (PCR), and direct sequencing was performed for mutational screening on the patient and normal controls consisted of 52 healthy blood donors. Reverse transcription PCR was conducted to test for exon skipping.Results The proposita patient showed dispersing platelets, prolonged bleeding time, and severely reduced platelet aggregation in response to the physiological agonists adenosine diphosphate (ADP), epinephrine, collagen, and ristocetin. Flow cytometric measurements showed that the contents of allb and β3 were significantly decreased. Sequencing results demonstrated two different types of heterozygous mutations existed in the allb gene (c.2930delG and IVS15-1delG). The compound mutations were also confirmed in the patient's mother and father separately.Conclusions The allbp3 deficiency of the proband was caused by two compound ITGA2B mutations, which were first reported in Chinese GT patients. The IVS15-1delG was first confirmed to cause an exon skipping.

  13. Gyrase activity and number of copies of the gyrase B subunit gene in Haemophilus influenzae

    Energy Technology Data Exchange (ETDEWEB)

    Cabrera-Juarez, E.; Setlow, J.K.

    1985-11-01

    Gyrase activities in extracts of various strains of Haemophilus influenzae can differ by more than an order of magnitude. Measurements of in vitro activity and copy number indicated that most of these differences arose from variations in the number of copies of the gene for the gyrase B subunit, with some strains containing multicopy plasmids coding for that subunit. The quantitative relationship between gyrase and copy number depended on the mutations in the plasmids and in the host. The possibility that the in vivo gyrase activity did not reflect the in vitro data was explored by measurement of alkaline phosphatase and ATPase activity in the extracts. Alkaline phosphatase activity increased with increasing gyrase activity measured in vitro, but ATPase activity did not. The authors conclude that extra supercoiling enhanced transcription of the alkaline phosphatase gene but not the ATPase gene and that it is unlikely that there is much discrepancy between gyrase activity assayed in vitro and the activity in the cell.

  14. Conservation of the Nrf2-Mediated Gene Regulation of Proteasome Subunits and Glucose Metabolism in Zebrafish

    Directory of Open Access Journals (Sweden)

    Vu Thanh Nguyen

    2016-01-01

    Full Text Available The Keap1-Nrf2 system is an evolutionarily conserved defense mechanism against oxidative and xenobiotic stress. Besides the exogenous stress response, Nrf2 has been found to regulate numerous cellular functions, including protein turnover and glucose metabolism; however, the evolutionary origins of these functions remain unknown. In the present study, we searched for novel target genes associated with the zebrafish Nrf2 to answer this question. A microarray analysis of zebrafish embryos that overexpressed Nrf2 revealed that 115 candidate genes were targets of Nrf2, including genes encoding proteasome subunits and enzymes involved in glucose metabolism. A real-time quantitative PCR suggested that the expression of 3 proteasome subunits (psma3, psma5, and psmb7 and 2 enzymes involved in glucose metabolism (pgd and fbp1a were regulated by zebrafish Nrf2. We thus next examined the upregulation of these genes by an Nrf2 activator, diethyl maleate, using Nrf2 mutant zebrafish larvae. The results of real-time quantitative PCR and whole-mount in situ hybridization showed that all of these 5 genes were upregulated by diethyl maleate treatment in an Nrf2-dependent manner, especially in the liver. These findings implied that the Nrf2-mediated regulation of the proteasome subunits and glucose metabolism is evolutionarily conserved among vertebrates.

  15. GABAA receptor subunit gene expression in human prefrontal cortex: comparison of schizophrenics and controls

    Science.gov (United States)

    Akbarian, S.; Huntsman, M. M.; Kim, J. J.; Tafazzoli, A.; Potkin, S. G.; Bunney, W. E. Jr; Jones, E. G.; Bloom, F. E. (Principal Investigator)

    1995-01-01

    The prefrontal cortex of schizophrenics is hypoactive and displays changes related to inhibitory, GABAergic neurons, and GABAergic synapses. These changes include decreased levels of glutamic acid decarboxylase (GAD), the enzyme for GABA synthesis, upregulation of muscimol binding, and downregulation of benzodiazepine binding to GABAA receptors. Studies in the visual cortex of nonhuman primates have demonstrated that gene expression for GAD and for several GABAA receptor subunit polypeptides is under control of neuronal activity, raising the possibility that similar mechanisms in the hypoactive prefrontal cortex of schizophrenics may explain the abnormalities in GAD and in GABAA receptor regulation. In the present study, which is the first of its type on human cerebral cortex, levels of mRNAs for six GABAA receptor subunits (alpha 1, alpha 2, alpha 5, beta 1, beta 2, gamma 2) and their laminar expression patterns were analyzed in the prefrontal cortex of schizophrenics and matched controls, using in situ hybridization histochemistry and densitometry. Three types of laminar expression pattern were observed: mRNAs for the alpha 1, beta 2, and gamma 2 subunits, which are the predominant receptor subunits expressed in the mature cortex, were expressed at comparatively high levels by cells of all six cortical layers, but most intensely by cells in lower layer III and layer IV. mRNAs for the alpha 2, alpha 5, and beta 1 subunits were expressed at lower levels; alpha 2 and beta 1 were expressed predominantly by cells in layers II, III, and IV; alpha 5 was expressed predominantly in layers IV, V, and VI. There were no significant changes in overall mRNA levels for any of the receptor subunits in the prefrontal cortex of schizophrenics, and the laminar expression pattern of all six receptor subunit mRNAs did not differ between schizophrenics and controls. Because gene expression for GABAA receptor subunits is not consistently altered in the prefrontal cortex of

  16. GRIN2B Gene and Associated Brain Cortical White Matter Changes in Bipolar Disorder: A Preliminary Combined Platform Investigation

    Directory of Open Access Journals (Sweden)

    Carissa Nadia Kuswanto

    2013-01-01

    Full Text Available Abnormalities in glutamate signaling and glutamate toxicity are thought to be important in the pathophysiology of bipolar disorder (BD. Whilst previous studies have found brain white matter changes in BD, there is paucity of data about how glutamatergic genes affect brain white matter integrity in BD. Based on extant neuroimaging data, we hypothesized that GRIN2B risk allele is associated with reductions of brain white matter integrity in the frontal, parietal, temporal, and occipital regions and cingulate gyrus in BD. Fourteen patients with BD and 22 healthy controls matched in terms of age, gender and handedness were genotyped using blood samples and underwent diffusion tensor imaging. Compared to G allele, brain FA values were significantly lower in BD patients with risk T allele in left frontal region (P=0.001, right frontal region (P=0.002, left parietal region (P=0.001, left occipital region (P=0.001, right occipital region (P<0.001, and left cingulate gyrus (P=0.001. Further elucidation of the interactions between different glutamate genes and their relationships with such structural, functional brain substrates will enhance our understanding of the link between dysregulated glutamatergic neurotransmission and neuroimaging endophenotypes in BD.

  17. Global view of transcriptome in the brains of aged NR2B transgenic mice*****

    Institute of Scientific and Technical Information of China (English)

    Chunxia Li; Men Su; Huimin Wang; Yinghe Hu

    2013-01-01

    NR2B subunits are involved in regulating aging, in particular, age-related learning and memory deficits. We examined 19-month-old NR2B transgenic mice and their littermate controls. First, we detected expression of the NR2B subunit gene, Grin2b, in the neocortex of transgenic mice using real-time PCR. Next, we used microarrays to examine differences in neocortical gene expression. Pathway and signal-net analyses identified multiple pathways altered in the transgenic mice, in-cluding the P53, Jak-STAT, Wnt, and Notch pathways, as wel as regulation of the actin cytoskeleton and neuroactive ligand-receptor interactions. Further signal-net analysis highlighted the P53 and insulin-like growth factor pathways as key regulatory pathways. Our results provide new insight into understanding the molecular mechanisms of NR2B regulated age-related memory storage, normal organismal aging and age-related disease.

  18. Potentially harmful advantage to athletes: a putative connection between UGT2B17 gene deletion polymorphism and renal disorders with prolonged use of anabolic androgenic steroids

    Directory of Open Access Journals (Sweden)

    Barker James

    2010-04-01

    Full Text Available Abstract Background and objective With prolonged use of anabolic androgenic steroids (AAS, occasional incidents of renal disorders have been observed. Independently, it has also been established that there are considerable inter-individual and inter-ethnic differences, in particular with reference to the uridine diphosphate-glucuronosyltransferase 2B17 (UGT2B17 gene, in metabolising these compounds. This report postulates the association of deletion polymorphism in the UGT2B17 gene with the occurrence of renal disorders on chronic exposure to AAS. Presentation of the hypothesis The major deactivation and elimination pathway of AASs is through glucuronide conjugation, chiefly catalyzed by the UGT2B17 enzyme, followed by excretion in urine. Excretion of steroids is affected in individuals with a deletion mutation in the UGT2B17 gene. We hypothesize that UGT2B17 deficient individuals are more vulnerable to developing renal disorders with prolonged use of AAS owing to increases in body mass index and possible direct toxic effects of steroids on the kidneys. Elevated serum levels of biologically active steroids due to inadequate elimination can lead to prolonged muscle build up. An increase in body mass index may cause renal injuries due to sustained elevated glomerular pressure and flow rate. Testing the hypothesis In the absence of controlled clinical trials in humans, observational studies can be carried out. Real time PCR with allelic discrimination should be employed to examine the prevalence of different UGT2B17 genotypes in patients with impaired renal function and AAS abuse. In individuals with the UGT2B17 deletion polymorphism, blood tests, biofluid analyses, urinalysis, and hair analyses following the administration of an anabolic steroid can be used to determine the fate of the substance once in the body. Implications of the hypothesis If the hypothesis is upheld, anabolic steroid users with a deletion mutation in the UGT2B17 gene may be

  19. Bone cancer pain induce anxiety-like behavior and high expression of NR2B subunit in anterior cingulate cortex of rats%骨癌痛诱发大鼠焦虑样行为和前扣带回脑区NR2B 的上调表达

    Institute of Scientific and Technical Information of China (English)

    赵宇; 刘瑾瑜

    2016-01-01

    Objective To investigate the effect of bone cancer pain on emotion and NMDA re-ceptor NR2B subunit expression level in anterior cingulate cortex (ACC)in rats.Methods One hun-dred and fifty healthy male Wistar rats weighing 200-250 g aged 3 months old were randomly divided into 3 groups (n = 50 in each group):sham operation group (group S),bone cancer pain group (group BCP),RO25-6981 group (group RO).The BCP model was established by inoculating Walker 256 breast cancer cells into right intra-tibial.Rats in group S were given the same dose of d-hanks. Group RO was injected intraperitoneally with RO25-6981 (5 mg/kg/d)on the day of inoculation, while rats in the group S and group BCP were given the same dose of normal saline.Mechanical with-drawal threshold (MWT)and thermal withdrawal latency (TWL)of right hind legs were measured on day 7,10,14 after inoculation respectively.Elevated plus-maze test was carried out to investigate the effect of bone cancer pain on emotion in rats after pain threshold detection,then the percentage of the times entering the open arms (OE)and the percentage of the time staying in the open arms (OT) duration the total period were evaluated.Then the anterior cingulate cortex tissue was removed to e-valuate the NR2B protein and mRNA expression levels by RT-PCR,Western blot and immunofluo-rescence methods on day 14 after elevated plus-maze test.Results All the parameters did not differ with significant difference between group S and group RO.MWT decreased and TWL shortened on day 7,10,14 after inoculation in group BCP compared with those before inoculation and those of group S and group RO.OE and OT in group BCP reduced remarkably than those before inoculation and those of group S and group RO.Relative absorbance of NR2B mRNA,the expression of NR2B pro-tein,average NR2B relative fluorescence intensity value is obviously higher than that of group S and group RO (P <0.05).Conclusion Bone cancer pain can induce pain-related aversion and anxiety

  20. The human thyrotropin beta-subunit gene differs in 5' structure from murine TSH-beta genes.

    Science.gov (United States)

    Guidon, P T; Whitfield, G K; Porti, D; Kourides, I A

    1988-12-01

    The gene encoding the beta-subunit of human thyrotropin (hTSH-beta) was isolated, and its nucleotide sequence was determined. The gene is 4.3 kb in length, consists of three exons and two introns, and is present as a single copy as determined by Southern blot analysis of total genomic DNA. The protein coding portion of the gene, which includes exons 2 and 3, was isolated from a human genomic phage library, while exon 1, which encodes only 5' untranslated mRNA sequence, was isolated from a plasmid library of size-selected genomic DNA fragments. Here we describe the isolation of the 5' untranslated exon of the hTSH-beta subunit and 5'-flanking region. The structure of the hTSH-beta gene is very similar to the previously characterized TSH-beta genes from mouse and rat. The genes from all three species have two distinct promoter regions, but while both promoters are utilized by the murine TSH-beta genes, the human TSH-beta gene apparently utilizes only the proximal promoter for transcription initiation. A striking difference in hTSH-beta gene structure compared to the murine genes is that exon 1 of the human gene is 36 nucleotides. An analysis of the mouse, rat, and human exon 1 and 5'-flanking region shows a high percentage of sequence homology, with the exception of a 9-nucleotide insertion 13 bases 3' from the proximal TATA box found in the human gene but not found in the other two species. We propose that this insertion results in the additional length of human exon 1 compared to the mouse and rat genes. By isolating the promoter region of the hTSH-beta gene, we can begin to identify specific sequences involved in the regulation of hTSH gene expression.

  1. Expression of five acetylcholine receptor subunit genes in Brugia malayi adult worms.

    Science.gov (United States)

    Li, Ben-Wen; Rush, Amy C; Weil, Gary J

    2015-12-01

    Acetylcholine receptors (AChRs) are required for body movement in parasitic nematodes and are targets of "classical" anthelmintic drugs such as levamisole and pyrantel and of newer drugs such as tribendimidine and derquantel. While neurotransmission explains the effects of these drugs on nematode movement, their effects on parasite reproduction are unexplained. The levamisole AChR type (L-AChRs) in Caenorhabditis elegans is comprised of five subunits: Cel-UNC-29, Cel-UNC-38, Cel-UNC-63, Cel-LEV-1 and Cel-LEV-8. The genome of the filarial parasite Brugia malayi contains nine AChRs subunits including orthologues of Cel-unc-29, Cel-unc-38, and Cel-unc-63. We performed in situ hybridization with RNA probes to localize the expression of five AChR genes (Bm1_35890-Bma-unc-29, Bm1_20330-Bma-unc-38, Bm1_38195-Bma-unc-63, Bm1_48815-Bma-acr-26 and Bm1_40515-Bma-acr-12) in B. malayi adult worms. Four of these genes had similar expression patterns with signals in body muscle, developing embryos, spermatogonia, uterine wall adjacent to stretched microfilariae, wall of V as deferens, and lateral cord. Three L-AChR subunit genes (Bma-unc-29, Bma-unc-38 and Bma-unc-63) were expressed in body muscle, which is a known target of levamisole. Bma-acr-12 was co-expressed with these levamisole subunit genes in muscle, and this suggests that its protein product may form receptors with other alpha subunits. Bma-acr-26 was expressed in male muscle but not in female muscle. Strong expression signals of these genes in early embryos and gametes in uterus and testis suggest that AChRs may have a role in nervous system development of embryogenesis and spermatogenesis. This would be consistent with embryotoxic effects of drugs that target these receptors in filarial worms. Our data show that the expression of these receptor genes is tightly regulated with regard to localization in adult worms and developmental stage in embryos and gametes. These results may help to explain the broad effects of

  2. Gene trap mutagenesis of hnRNP A2/B1: a cryptic 3' splice site in the neomycin resistance gene allows continued expression of the disrupted cellular gene

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    DeGregori James V

    2003-01-01

    Full Text Available Abstract Background Tagged sequence mutagenesis is a process for constructing libraries of sequenced insertion mutations in embryonic stem cells that can be transmitted into the mouse germline. To better predict the functional consequences of gene entrapment on cellular gene expression, the present study characterized the effects of a U3Neo gene trap retrovirus inserted into an intron of the hnRNP A2/B1 gene. The mutation was selected for analysis because it occurred in a highly expressed gene and yet did not produce obvious phenotypes following germline transmission. Results Sequences flanking the integrated gene trap vector in 1B4 cells were used to isolate a full-length cDNA whose predicted amino acid sequence is identical to the human A2 protein at all but one of 341 amino acid residues. hnRNP A2/B1 transcripts extending into the provirus utilize a cryptic 3' splice site located 28 nucleotides downstream of the neomycin phosphotransferase start codon. The inserted Neo sequence and proviral poly(A site function as an 3' terminal exon that is utilized to produce hnRNP A2/B1-Neo fusion transcripts, or skipped to produce wild-type hnRNP A2/B1 transcripts. This results in only a modest disruption of hnRNPA2/B1 gene expression. Conclusions Expression of the occupied hnRNP A2/B1 gene and utilization of the viral poly(A site are consistent with an exon definition model of pre-mRNA splicing. These results reveal a mechanism by which U3 gene trap vectors can be expressed without disrupting cellular gene expression, thus suggesting ways to improve these vectors for gene trap mutagenesis.

  3. Lack of evidence for mutations or deletions in the CDKN2A/p16 and CDKN2B/p15 genes of Brazilian neuroblastoma patients

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    Bassi C.L.

    2004-01-01

    Full Text Available Neuroblastoma, the most common extracranial tumor in childhood, has a wide spectrum of clinical and biological features. The loss of heterozygosity within the 9p21 region has been reported as a prognostic factor. Two tumor suppressor genes located in this region, the CDKN2B/p15 and CDKN2A/p16 (cyclin-dependent kinase inhibitors 2B and 2A, respectively genes, play a critical role in cell cycle progression and are considered to be targets for tumor inactivation. We analyzed CDKN2B/p15 and CDKN2A/p16 gene alterations in 11 patients, who ranged in age from 4 months to 13 years (male/female ratio was 1.2:1. The most frequent stage of the tumor was stage IV (50%, followed by stages II and III (20% and stage I (10%. The samples were submitted to the multiplex PCR technique for homozygous deletion analysis and to single-strand conformation polymorphism and nucleotide sequencing for mutation analysis. All exons of both genes were analyzed, but no deletion was detected. One sample exhibited shift mobility specific for exon 2 in the CDKN2B/p15 gene, not confirmed by DNA sequencing. Homozygous deletions and mutations are not involved in the inactivation mechanism of the CDKN2B/p15 and CDKN2A/p16 genes in neuroblastoma; however, these two abnormalities do not exclude other inactivation pathways. Recent evidence has shown that the expression of these genes is altered in this disease. Therefore, other mechanisms of inactivation, such as methylation of promoter region and unproperly function of proteins, may be considered in order to estimate the real contribution of these genes to neuroblastoma genesis or disease progression.

  4. Genetic differentiation of the mitochondrial cytochrome oxidase C subunit I gene in genus Paramecium (Protista, Ciliophora.

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    Yan Zhao

    Full Text Available BACKGROUND: The mitochondrial cytochrome c oxidase subunit I (COI gene is being used increasingly for evaluating inter- and intra-specific genetic diversity of ciliated protists. However, very few studies focus on assessing genetic divergence of the COI gene within individuals and how its presence might affect species identification and population structure analyses. METHODOLOGY/PRINCIPAL FINDINGS: We evaluated the genetic variation of the COI gene in five Paramecium species for a total of 147 clones derived from 21 individuals and 7 populations. We identified a total of 90 haplotypes with several individuals carrying more than one haplotype. Parsimony network and phylogenetic tree analyses revealed that intra-individual diversity had no effect in species identification and only a minor effect on population structure. CONCLUSIONS: Our results suggest that the COI gene is a suitable marker for resolving inter- and intra-specific relationships of Paramecium spp.

  5. Induction of UDP-glucuronosyltransferase 2B15 gene expression by the major active metabolites of tamoxifen, 4-hydroxytamoxifen and endoxifen, in breast cancer cells.

    Science.gov (United States)

    Chanawong, Apichaya; Hu, Dong Gui; Meech, Robyn; Mackenzie, Peter I; McKinnon, Ross A

    2015-06-01

    We previously reported upregulation of UGT2B15 by 17β-estradiol in breast cancer MCF7 cells via binding of the estrogen receptor α (ERα) to an estrogen response unit (ERU) in the proximal UGT2B15 promoter. In the present study, we show that this ERα-mediated upregulation was significantly reduced by two ER antagonists (fulvestrant and raloxifene) but was not affected by a third ER antagonist, 4-hydroxytamoxifen (4-OHTAM), a major active tamoxifen (TAM) metabolite. Furthermore, we found that, similar to 17β-estradiol, 4-OHTAM and endoxifen (another major active TAM metabolite) elevated UGT2B15 mRNA levels, and that this stimulation was significantly abrogated by fulvestrant. Further experiments using 4-OHTAM revealed a critical role for ERα in this regulation. Specifically; knockdown of ERα expression by anti-ERα small interfering RNA reduced the 4-OHTAM-mediated induction of UGT2B15 expression; 4-OHTAM activated the wild-type but not the ERU-mutated UGT2B15 promoter; and chromatin immunoprecipitation assays showed increased ERα occupancy at the UGT2B15 ERU in MCF7 cells upon exposure to 4-OHTAM. Together, these data indicate that both 17β-estradiol and the antiestrogen 4-OHTAM upregulate UGT2B15 in MCF7 cells via the same ERα-signaling pathway. This is consistent with previous observations that both 17β-estradiol and TAM upregulate a common set of genes in MCF7 cells via the ER-signaling pathway. As 4-OHTAM is a UGT2B15 substrate, the upregulation of UGT2B15 by 4-OHTAM in target breast cancer cells is likely to enhance local metabolism and inactivation of 4-OHTAM within the tumor. This represents a potential mechanism that may reduce TAM therapeutic efficacy or even contribute to the development of acquired TAM resistance.

  6. Multiple thyrotropin β-subunit and thyrotropin receptor-related genes arose during vertebrate evolution.

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    Gersende Maugars

    Full Text Available Thyroid-stimulating hormone (TSH is composed of a specific β subunit and an α subunit that is shared with the two pituitary gonadotropins. The three β subunits derive from a common ancestral gene through two genome duplications (1R and 2R that took place before the radiation of vertebrates. Analysis of genomic data from phylogenetically relevant species allowed us to identify an additional Tshβ subunit-related gene that was generated through 2R. This gene, named Tshβ2, present in cartilaginous fish, little skate and elephant shark, and in early lobe-finned fish, coelacanth and lungfish, was lost in ray-finned fish and tetrapods. The absence of a second type of TSH receptor (Tshr gene in these species suggests that both TSHs act through the same receptor. A novel Tshβ sister gene, named Tshβ3, was generated through the third genomic duplication (3R that occurred early in the teleost lineage. Tshβ3 is present in most teleost groups but was lostin tedraodontiforms. The 3R also generated a second Tshr, named Tshrb. Interestingly, the new Tshrb was translocated from its original chromosomic position after the emergence of eels and was then maintained in its new position. Tshrb was lost in tetraodontiforms and in ostariophysians including zebrafish although the latter species have two TSHs, suggesting that TSHRb may be dispensable. The tissue distribution of duplicated Tshβs and Tshrs was studied in the European eel. The endocrine thyrotropic function in the eel would be essentially mediated by the classical Tshβ and Tshra, which are mainly expressed in the pituitary and thyroid, respectively. Tshβ3 and Tshrb showed a similar distribution pattern in the brain, pituitary, ovary and adipose tissue, suggesting a possible paracrine/autocrine mode of action in these non-thyroidal tissues. Further studies will be needed to determine the binding specificity of the two receptors and how these two TSH systems are interrelated.

  7. Fragmentation of the large subunit ribosomal RNA gene in oyster mitochondrial genomes

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    Milbury Coren A

    2010-09-01

    Full Text Available Abstract Background Discontinuous genes have been observed in bacteria, archaea, and eukaryotic nuclei, mitochondria and chloroplasts. Gene discontinuity occurs in multiple forms: the two most frequent forms result from introns that are spliced out of the RNA and the resulting exons are spliced together to form a single transcript, and fragmented gene transcripts that are not covalently attached post-transcriptionally. Within the past few years, fragmented ribosomal RNA (rRNA genes have been discovered in bilateral metazoan mitochondria, all within a group of related oysters. Results In this study, we have characterized this fragmentation with comparative analysis and experimentation. We present secondary structures, modeled using comparative sequence analysis of the discontinuous mitochondrial large subunit rRNA genes of the cupped oysters C. virginica, C. gigas, and C. hongkongensis. Comparative structure models for the large subunit rRNA in each of the three oyster species are generally similar to those for other bilateral metazoans. We also used RT-PCR and analyzed ESTs to determine if the two fragmented LSU rRNAs are spliced together. The two segments are transcribed separately, and not spliced together although they still form functional rRNAs and ribosomes. Conclusions Although many examples of discontinuous ribosomal genes have been documented in bacteria and archaea, as well as the nuclei, chloroplasts, and mitochondria of eukaryotes, oysters are some of the first characterized examples of fragmented bilateral animal mitochondrial rRNA genes. The secondary structures of the oyster LSU rRNA fragments have been predicted on the basis of previous comparative metazoan mitochondrial LSU rRNA structure models.

  8. An AS-PCR assay for accurate genotyping of FAD2A/FAD2B genes in peanuts (Arachis hypogaea L.

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    Yu, H. T.

    2013-09-01

    Full Text Available FAD2A and FAD2B are homoeologous genes from A and B genomes in cultivated peanuts (Arachis hypogaea L. encoding fatty acid desaturates which convert oleate to linoleate. To study the genetics of oleate and breed high oleate peanut cultivars, a simple allele specific-PCR (AS-PCR protocol for the accurate genotyping of FAD2A/FAD2B was developed to discriminate the wild and mutant allele of both genes (FAD2A 448 G > A and FAD2B 441_442insA. The results may serve to develop a feasible procedure for producing highly desired high oleate peanut cultivars through hybridization.FAD2A y FAD2B son genes homólogos de genomas A y B en cacahuete cultivado (Arachis hypogaea L. que codifican las desaturasas de ácidos grasos que convierten oleato a linoleato. Para estudiar la genética de las variedades mejoradas de cacahuete alto oleato, fue desarrollado un protocolo sencillo del alelo específico PCR-(AS-PCR para la determinación precisa del genotipo de FAD2A/FAD2B y discriminar el alelo silvestre y el mutante de ambos genes (FAD2A 448 G > A y FAD2B 441_442insA. Los resultados pueden servir para desarrollar un procedimiento viable para la producción de cultivares deseados de cacahuetes alto oleato a través de hibridación.

  9. Synonymous Codon Usage Bias and Overexpression of a Synthetic Gene Encoding Interferon α2b in Yeast

    Institute of Scientific and Technical Information of China (English)

    Bin FANG; Bu-feng LIANG; Guang-yuan HE

    2007-01-01

    To achieve higher level expression of Interferon α2b (IFN-α2b) in methylotrophic yeast (Pichia pastoris), a cDNA fragment coding for the mature IFN-α2b was designed and synthesized based on the synonymous codon bias of P. pastoris and optimized G+C content. The synthetic IFN-α2b was inserted into the secreted expression vector pPICZαA, and then integrated into P. pastoris GS115 genome by electroporation. Multi-copy integrants in the Mut+ recombinant P. pastoris strain were screened by high concentrations of Zeocin. 120 hours culturing allowed expression of the IFN-α2b transformant up to 810 mg/L as detected by SDS-PAGE and quantitative methods. In addition, Western blot analysis showed that the recombinant proteins had immunogenicity. The significant antiviral activity of the recombinant IFN-α2b protein was verified by WISH/ VSV system, which was 3.3×105 IU/mL.

  10. Apoptosis Gene Hunting Using Retroviral Expression Cloning: Identification of Vacuolar ATPase Subunit E

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    Claire L. Anderson

    2003-01-01

    Full Text Available Over the past 10-15 years there has been an explosion of interest in apoptosis. The delayed realisation that cell death is an essential part of life for any multicellular organism has meant that, despite the recent and rapid developments of the last decade, the precise biochemical pathways involved in apoptosis remain incomplete and potentially novel genes may, as yet, remain undiscovered. The hunt is therefore on to bridge the remaining gaps in our knowledge. Our contribution to this research effort utilises a functional cloning approach to isolate important regulatory genes involved in apoptosis. This mini-review focuses on the use and advantages of a retroviral expression cloning strategy and describes the isolation and identification of one such potential apoptosis regulatory gene, namely that encoding vacuolar ATPase subunit E.

  11. NR2B GENE EXPRESSION CHANGE IN WISTAR RAT PRACTICING AEROBIC EXERCISE COMPARING TO SOYBEAN (GLYCINE MAX OR PHYLLANTHUS NIRURI INTAKES AND SOYBEAN AND PHYLLANTHUS NIRURI COMPOSITION INTAKE

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    Vita Murniati Tarawan

    2015-09-01

    Full Text Available Objective: To examine the relationship between nutrition and brain memory. Methods: This study was an experimental laboratory study conducted during the period of June 2011 to July 2012 at the Biomedical and Biochemistry laboratory, Faculty of Medicine, Universitas Padjadjaran. The subjects were 56 8-week-old male Wistar rats weighing approximately 200–250 grams which were divided into 8 groups with different treatments. The treatment groups received no exercise or exercise and soybean (Glycine max, Phyllanthus niruri, or combination of both. Results: NR2B gene expression changes found is described as follows: (1 without practicing exercise (3.8 and after exercise (4.6; (2 Glycine max minus exercise (2.86 and Glycine max and exercise (3.17; (3 Phyllanthus niruri minus exercises (4.7 and Phyllanthus niruri and exercise (4.9; and (4 Glycine max and Phyllanthus niruri combination minus exercise (3.14 and Glycine max and Phyllanthus niruri combination and exercise (4.83. Conclusions: This study determines that exercises and Phyllanthus niruri intake enhance NR2B gene expressions. Glycine max inhibits the NR2B gene expressions. Glycine max and Phyllanthus niruri combination, both with and without practicing exercises, enhance NR2B gene expressions. Therefore, practicing exercise and Phyllanthus niruri intake might cause brain cell apoptosis while Glycine max intake inhibits brain cell apoptosis.

  12. Traxoprodil, a selective antagonist of the NR2B subunit of the NMDA receptor, potentiates the antidepressant-like effects of certain antidepressant drugs in the forced swim test in mice.

    Science.gov (United States)

    Poleszak, Ewa; Stasiuk, Weronika; Szopa, Aleksandra; Wyska, Elżbieta; Serefko, Anna; Oniszczuk, Anna; Wośko, Sylwia; Świąder, Katarzyna; Wlaź, Piotr

    2016-08-01

    One of the newest substances, whose antidepressant activity was shown is traxoprodil, which is a selective antagonist of the NR2B subunit of the NMDA receptor. The main goal of the present study was to evaluate the effect of traxoprodil on animals' behavior using the forced swim test (FST), as well as the effect of traxoprodil (10 mg/kg) on the activity of antidepressants, such as imipramine (15 mg/kg), fluoxetine (5 mg/kg), escitalopram (2 mg/kg) and reboxetine (2.5 mg/kg). Serotonergic lesion and experiment using the selective agonists of serotonin receptors 5-HT1A and 5-HT2 was conducted to evaluate the role of the serotonergic system in the antidepressant action of traxoprodil. Brain concentrations of tested agents were determined using HPLC. The results showed that traxoprodil at a dose of 20 and 40 mg/kg exhibited antidepressant activity in the FST and it was not related to changes in animals' locomotor activity. Co-administration of traxoprodil with imipramine, fluoxetine or escitalopram, each in subtherapeutic doses, significantly affected the animals' behavior in the FST and, what is important, these changes were not due to the severity of locomotor activity. The observed effect of traxoprodil is only partially associated with serotonergic system and is independent of the effect on the 5-HT1A and 5-HT2 serotonin receptors. The results of an attempt to assess the nature of the interaction between traxoprodil and the tested drugs show that in the case of joint administration of traxoprodil and fluoxetine, imipramine or escitalopram, there were interactions in the pharmacokinetic phase.

  13. [Association polymorphic variants of GRIN2B gene with paranoid schizophrenia and response to common neuroleptics in Russians and Tatars from Bashkortostan Republic].

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    Gareeva, A E; Zakirov, D F; Khusnutdinova, E K

    2013-09-01

    An analysis of the association of paranoid schizophrenia seeking with polymorphic variants of GRIN2B gene was performed in order to identify genetic risk factors of disease development and genetic markers of the response to therapy by neuroleptics in Russian and Tatar patients from Bashkortostan Republic (BB). In the course of the analysis, we revealed the following: 1) genetic markers of increased risk of developing paranoid schizophrenia in various ethnic groups, including, in Tatars, the GRIN2B* T/*Tgenotype (p = 0.003; OR = 2.33) and GRIN2B*T allele (p = 0.001; OR = 2.36), rs1805247; in Russians, the GRIN2B*T/*T genotype (p = 0.038; OR = 2.12) and GRIN2B* T allele (p = 0.028; OR = 2.03), rs1805247, genotype GRIN2B*A/*A (p = 0.042; OR = 2.12), rs1805476; 2) genetic markers of the reduced risk of developing paranoid schizophrenia; 3) genetic markers of therapy response and the risk of side effects development during neuroleptics (haloperidol) treatment in Bashkortostan. The significant interethnic diversity of genetic factors related to the risk of this disease development was noted.

  14. Construction of Recombinant Plasmid Containing S. Mutans F-ATPase β Subunit Gene

    Institute of Scientific and Technical Information of China (English)

    YU Dan-ni; JIANG Li

    2005-01-01

    objective: construct a homologous recombinant plasmid which was expected to be transformed into S. mutans Methods: a region at the 5' terminus of the S. mutans F-ATPase β subunit gene was amplified by PCR, the PCR product was inserted into vector pVA891, yielding recombinant plasmid. Results: the DNA sequence of the recombinant plasmid was identified correct in whole by restriction endonuclease and DNA sequence techniques. Conclusion: the recombinant plasmid of S. mutans DNA was cloned in effect ,it may assist in construction of homologues recombinant mutant.

  15. High basal expression of interferon-stimulated genes in human bronchial epithelial (BEAS-2B cells contributes to influenza A virus resistance.

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    Lai-Giea Seng

    Full Text Available Respiratory epithelial cells play a key role in influenza A virus (IAV pathogenesis and host innate response. Transformed human respiratory cell lines are widely used in the study of IAV-host interactions due to their relative convenience, and inherent difficulties in working with primary cells. Transformed cells, however, may have altered susceptibility to virus infection. Proper characterization of different respiratory cell types in their responses to IAV infection is therefore needed to ensure that the cell line chosen will provide results that are of relevance in vivo. We compared replication kinetics of human H1N1 (A/USSR/77 IAVs in normal primary human bronchial epithelial (NHBE and two commonly used respiratory epithelial cell lines namely BEAS-2B and A549 cells. We found that IAV replication was distinctly poor in BEAS-2B cells in comparison with NHBE, A549 and Madin-Darby canine kidney (MDCK cells. IAV resistance in BEAS-2B cells was accompanied by an activated antiviral state with high basal expression of interferon (IFN regulatory factor-7 (IRF-7, stimulator of IFN genes (STING and IFN stimulated genes (ISGs. Treatment of BEAS-2B cells with a pan-Janus-activated-kinase (JAK inhibitor decreased IRF-7 and ISG expression and resulted in increased IAV replication. Therefore, the use of highly resistant BEAS-2B cells in IAV infection may not reflect the cytopathogenicity of IAV in human epithelial cells in vivo.

  16. Sequence analysis of dolphin ferritin H and L subunits and possible iron-dependent translational control of dolphin ferritin gene

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    Sasaki Yukako

    2008-10-01

    Full Text Available Abstract Background Iron-storage protein, ferritin plays a central role in iron metabolism. Ferritin has dual function to store iron and segregate iron for protection of iron-catalyzed reactive oxygen species. Tissue ferritin is composed of two kinds of subunits (H: heavy chain or heart-type subunit; L: light chain or liver-type subunit. Ferritin gene expression is controlled at translational level in iron-dependent manner or at transcriptional level in iron-independent manner. However, sequencing analysis of marine mammalian ferritin subunits has not yet been performed fully. The purpose of this study is to reveal cDNA-derived amino acid sequences of cetacean ferritin H and L subunits, and demonstrate the possibility of expression of these subunits, especially H subunit, by iron. Methods Sequence analyses of cetacean ferritin H and L subunits were performed by direct sequencing of polymerase chain reaction (PCR fragments from cDNAs generated via reverse transcription-PCR of leukocyte total RNA prepared from blood samples of six different dolphin species (Pseudorca crassidens, Lagenorhynchus obliquidens, Grampus griseus, Globicephala macrorhynchus, Tursiops truncatus, and Delphinapterus leucas. The putative iron-responsive element sequence in the 5'-untranslated region of the six different dolphin species was revealed by direct sequencing of PCR fragments obtained using leukocyte genomic DNA. Results Dolphin H and L subunits consist of 182 and 174 amino acids, respectively, and amino acid sequence identities of ferritin subunits among these dolphins are highly conserved (H: 99–100%, (99→98 ; L: 98–100%. The conserved 28 bp IRE sequence was located -144 bp upstream from the initiation codon in the six different dolphin species. Conclusion These results indicate that six different dolphin species have conserved ferritin sequences, and suggest that these genes are iron-dependently expressed.

  17. The dnaN gene codes for the beta subunit of DNA polymerase III holoenzyme of escherichia coli.

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    Burgers, P M; Kornberg, A; Sakakibara, Y

    1981-09-01

    An Escherichia coli mutant, dnaN59, stops DNA synthesis promptly upon a shift to a high temperature; the wild-type dnaN gene carried in a transducing phage encodes a polypeptide of about 41,000 daltons [Sakakibara, Y. & Mizukami, T. (1980) Mol. Gen. Genet. 178, 541-553; Yuasa, S. & Sakakibara, Y. (1980) Mol. Gen. Genet. 180, 267-273]. We now find that the product of dnaN gene is the beta subunit of DNA polymerase III holoenzyme, the principal DNA synthetic multipolypeptide complex in E. coli. The conclusion is based on the following observations: (i) Extracts from dnaN59 cells were defective in phage phi X174 and G4 DNA synthesis after the mutant cells had been exposed to the increased temperature. (ii) The enzymatic defect was overcome by addition of purified beta subunit but not by other subunits of DNA polymerase III holoenzyme or by other replication proteins required for phi X174 DNA synthesis. (iii) Partially purified beta subunit from the dnaN mutant, unlike that from the wild type, was inactive in reconstituting the holoenzyme when mixed with the other purified subunits. (iv) Increased dosage of the dnaN gene provided by a plasmid carrying the gene raised cellular levels of the beta subunit 5- to 6-fold.

  18. Gene regulation, alternative splicing, and posttranslational modification of troponin subunits in cardiac development and adaptation: a focused review.

    Science.gov (United States)

    Sheng, Juan-Juan; Jin, Jian-Ping

    2014-01-01

    Troponin plays a central role in regulating the contraction and relaxation of vertebrate striated muscles. This review focuses on the isoform gene regulation, alternative RNA splicing, and posttranslational modifications of troponin subunits in cardiac development and adaptation. Transcriptional and posttranscriptional regulations such as phosphorylation and proteolysis modifications, and structure-function relationships of troponin subunit proteins are summarized. The physiological and pathophysiological significances are discussed for impacts on cardiac muscle contractility, heart function, and adaptations in health and diseases.

  19. Agrobacterium-mediated transformation of the β-subunit gene in 7S globulin protein in soybean using RNAi technology.

    Science.gov (United States)

    Qu, J; Liu, S Y; Wang, P W; Guan, S Y; Fan, Y G; Yao, D; Zhang, L; Dai, J L

    2016-04-26

    The objective of this study was to use RNA interference (RNAi) to improve protein quality and decrease anti-nutritional effects in soybean. Agrobacterium tumefaciens-mediated transformation was conducted using RNAi and an expression vector containing the 7S globulin β-subunit gene. The BAR gene was used as the selective marker and cotyledonary nodes of soybean genotype Jinong 27 were chosen as explant material. Regenerated plants were detected by molecular biology techniques. Transformation of the β-subunit gene in the 7S protein was detected by PCR, Southern blot, and q-PCR. Positive plants (10 T0, and 6 T1, and 13 T2) were tested by PCR. Hybridization bands were detected by Southern blot analysis in two of the T1 transgenic plants. RNAi expression vectors containing the soybean 7S protein β-subunit gene were successfully integrated into the genome of transgenic plants. qRT-PCR analysis in soybean seeds showed a clear decrease in expression of the soybean β-subunit gene. The level of 7S protein β-subunit expression in transgenic plants decreased by 77.5% as compared to that of the wild-type plants. This study has established a basis for the application of RNAi to improve the anti-nutritional effects of soybean.

  20. The toxic effects of Tris-(2,3-dibromopropyl)isocyanurate(TBC) on genes expression of bmp2b and bmp4 of zebrafish embryos

    Institute of Scientific and Technical Information of China (English)

    JIA Wan-jun

    2016-01-01

    We exposed zebrafish embryos to Tris-(2,3-dibromopropyl)isocyanurate(TBC)at the concentration of 20ppb, 100ppb, 400ppb, 1000ppb for 120h and 0.1%DMSO was set as the control group. Bmp2b and bmp4 were chosen perform RT-PCR to determine their genes expression level. The results showed that, TBC influenced their genes expression level in some extent and it significantly raised the genes expression level at the concentration of 20ppb.

  1. Molecular cloning and analysis of zebrafish voltage-gated sodium channel beta subunit genes: implications for the evolution of electrical signaling in vertebrates

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    Zhong Tao P

    2007-07-01

    Full Text Available Abstract Background Action potential generation in excitable cells such as myocytes and neurons critically depends on voltage-gated sodium channels. In mammals, sodium channels exist as macromolecular complexes that include a pore-forming alpha subunit and 1 or more modulatory beta subunits. Although alpha subunit genes have been cloned from diverse metazoans including flies, jellyfish, and humans, beta subunits have not previously been identified in any non-mammalian species. To gain further insight into the evolution of electrical signaling in vertebrates, we investigated beta subunit genes in the teleost Danio rerio (zebrafish. Results We identified and cloned single zebrafish gene homologs for beta1-beta3 (zbeta1-zbeta3 and duplicate genes for beta4 (zbeta4.1, zbeta4.2. Sodium channel beta subunit loci are similarly organized in fish and mammalian genomes. Unlike their mammalian counterparts, zbeta1 and zbeta2 subunit genes display extensive alternative splicing. Zebrafish beta subunit genes and their splice variants are differentially-expressed in excitable tissues, indicating tissue-specific regulation of zbeta1-4 expression and splicing. Co-expression of the genes encoding zbeta1 and the zebrafish sodium channel alpha subunit Nav1.5 in Chinese Hamster Ovary cells increased sodium current and altered channel gating, demonstrating functional interactions between zebrafish alpha and beta subunits. Analysis of the synteny and phylogeny of mammalian, teleost, amphibian, and avian beta subunit and related genes indicated that all extant vertebrate beta subunits are orthologous, that beta2/beta4 and beta1/beta3 share common ancestry, and that beta subunits are closely related to other proteins sharing the V-type immunoglobulin domain structure. Vertebrate sodium channel beta subunit genes were not identified in the genomes of invertebrate chordates and are unrelated to known subunits of the para sodium channel in Drosophila. Conclusion The

  2. Mutations in the Gene Encoding the Ancillary Pilin Subunit of the Streptococcus suis srtF Cluster Result in Pili Formed by the Major Subunit Only

    Science.gov (United States)

    Fittipaldi, Nahuel; Takamatsu, Daisuke; la Cruz Domínguez-Punaro, María de; Lecours, Marie-Pier; Montpetit, Diane; Osaki, Makoto; Sekizaki, Tsutomu; Gottschalk, Marcelo

    2010-01-01

    Pili have been shown to contribute to the virulence of different Gram-positive pathogenic species. Among other critical steps of bacterial pathogenesis, these structures participate in adherence to host cells, colonization and systemic virulence. Recently, the presence of at least four discrete gene clusters encoding putative pili has been revealed in the major swine pathogen and emerging zoonotic agent Streptococcus suis. However, pili production by this species has not yet been demonstrated. In this study, we investigated the functionality of one of these pili clusters, known as the srtF pilus cluster, by the construction of mutant strains for each of the four genes of the cluster as well as by the generation of antibodies against the putative pilin subunits. Results revealed that the S. suis serotype 2 strain P1/7, as well as several other highly virulent invasive S. suis serotype 2 isolates express pili from this cluster. However, in most cases tested, and as a result of nonsense mutations at the 5′ end of the gene encoding the minor pilin subunit (a putative adhesin), pili were formed by the major pilin subunit only. We then evaluated the role these pili play in S. suis virulence. Abolishment of the expression of srtF cluster-encoded pili did not result in impaired interactions of S. suis with porcine brain microvascular endothelial cells. Furthermore, non-piliated mutants were as virulent as the wild type strain when evaluated in a murine model of S. suis sepsis. Our results show that srtF cluster-encoded, S. suis pili are atypical compared to other Gram-positive pili. In addition, since the highly virulent strains under investigation are unlikely to produce other pili, our results suggest that pili might be dispensable for critical steps of the S. suis pathogenesis of infection. PMID:20052283

  3. NUCLEOTIDE-SEQUENCE OF THE LAST EXON OF THE GENE FOR HUMAN CYTOCHROME-C-OXIDASE SUBUNIT-VIB AND ITS FLANKING REGIONS

    NARCIS (Netherlands)

    TAANMAN, JW; SCHRAGE, C; BOKMA, E; REUVEKAMP, P; AGSTERIBBE, E; DEVRIES, H

    1991-01-01

    A human genomic clone encompassing the last exon of the gene for cytochrome c oxidase subunit VIb and a human genomic clone containing the most distal end of this gene were characterized. The last exon of the gene codes for the 17 C-terminal amino acid residues of the subunit and the 3' noncoding re

  4. DC-SCRIPT is a novel regulator of the tumor suppressor gene CDKN2B and induces cell cycle arrest in ERα-positive breast cancer cells.

    Science.gov (United States)

    Ansems, Marleen; Søndergaard, Jonas Nørskov; Sieuwerts, Anieta M; Looman, Maaike W G; Smid, Marcel; de Graaf, Annemarie M A; de Weerd, Vanja; Zuidscherwoude, Malou; Foekens, John A; Martens, John W M; Adema, Gosse J

    2015-02-01

    Breast cancer is one of the most common causes of cancer-related deaths in women. The estrogen receptor (ERα) is well known for having growth promoting effects in breast cancer. Recently, we have identified DC-SCRIPT (ZNF366) as a co-suppressor of ERα and as a strong and independent prognostic marker in ESR1 (ERα gene)-positive breast cancer patients. In this study, we further investigated the molecular mechanism on how DC-SCRIPT inhibits breast cancer cell growth. DC-SCRIPT mRNA levels from 190 primary ESR1-positive breast tumors were related to global gene expression, followed by gene ontology and pathway analysis. The effect of DC-SCRIPT on breast cancer cell growth and cell cycle arrest was investigated using novel DC-SCRIPT-inducible MCF7 breast cancer cell lines. Genome-wide expression profiling of DC-SCRIPT-expressing MCF7 cells was performed to investigate the effect of DC-SCRIPT on cell cycle-related gene expression. Findings were validated by real-time PCR in a cohort of 1,132 ESR1-positive breast cancer patients. In the primary ESR1-positive breast tumors, DC-SCRIPT expression negatively correlated with several cell cycle gene ontologies and pathways. DC-SCRIPT expression strongly reduced breast cancer cell growth in vitro, breast tumor growth in vivo, and induced cell cycle arrest. In addition, in the presence of DC-SCRIPT, multiple cell cycles related genes were differentially expressed including the tumor suppressor gene CDKN2B. Moreover, in 1,132 primary ESR1-positive breast tumors, DC-SCRIPT expression also correlated with CDKN2B expression. Collectively, these data show that DC-SCRIPT acts as a novel regulator of CDKN2B and induces cell cycle arrest in ESR1-positive breast cancer cells.

  5. Analysis of the cytochrome c oxidase subunit II (COX2) gene in giant panda, Ailuropoda melanoleuca.

    Science.gov (United States)

    Ling, S S; Zhu, Y; Lan, D; Li, D S; Pang, H Z; Wang, Y; Li, D Y; Wei, R P; Zhang, H M; Wang, C D; Hu, Y D

    2017-01-23

    The giant panda, Ailuropoda melanoleuca (Ursidae), has a unique bamboo-based diet; however, this low-energy intake has been sufficient to maintain the metabolic processes of this species since the fourth ice age. As mitochondria are the main sites for energy metabolism in animals, the protein-coding genes involved in mitochondrial respiratory chains, particularly cytochrome c oxidase subunit II (COX2), which is the rate-limiting enzyme in electron transfer, could play an important role in giant panda metabolism. Therefore, the present study aimed to isolate, sequence, and analyze the COX2 DNA from individuals kept at the Giant Panda Protection and Research Center, China, and compare these sequences with those of the other Ursidae family members. Multiple sequence alignment showed that the COX2 gene had three point mutations that defined three haplotypes, with 60% of the sequences corresponding to haplotype I. The neutrality tests revealed that the COX2 gene was conserved throughout evolution, and the maximum likelihood phylogenetic analysis, using homologous sequences from other Ursidae species, showed clustering of the COX2 sequences of giant pandas, suggesting that this gene evolved differently in them.

  6. The β-conglycinin deficiency in wild soybean is associated with the tail-to-tail inverted repeat of the α-subunit genes.

    Science.gov (United States)

    Tsubokura, Yasutaka; Hajika, Makita; Kanamori, Hiroyuki; Xia, Zhengjun; Watanabe, Satoshi; Kaga, Akito; Katayose, Yuichi; Ishimoto, Masao; Harada, Kyuya

    2012-02-01

    β-conglycinin, a major seed protein in soybean, is composed of α, α', and β subunits sharing a high homology among them. Despite its many health benefits, β-conglycinin has a lower amino acid score and lower functional gelling properties compared to glycinin, another major soybean seed protein. In addition, the α, α', and β subunits also contain major allergens. A wild soybean (Glycine soja Sieb et Zucc.) line, 'QT2', lacks all of the β-conglycinin subunits, and the deficiency is controlled by a single dominant gene, Scg-1 (Suppressor of β-conglycinin). This gene was characterized using a soybean cultivar 'Fukuyutaka', 'QY7-25', (its near-isogenic line carrying the Scg-1 gene), and the F₂ population derived from them. The physical map of the Scg-1 region covered by lambda phage genomic clones revealed that the two α-subunit genes, a β-subunit gene, and a pseudo α-subunit gene were closely organized. The two α-subunit genes were arranged in a tail-to-tail orientation, and the genes were separated by 197 bp in Scg-1 compared to 3.3 kb in the normal allele (scg-1). In addition, small RNA was detected in immature seeds of the mutants by northern blot analysis using an RNA probe of the α subunit. These results strongly suggest that β-conglycinin deficiency in QT2 is controlled by post-transcriptional gene silencing through the inverted repeat of the α subunits.

  7. A novel mutation in the sodium channel α1 subunit gene in a child with Dravet syndrome in Turkey

    Institute of Scientific and Technical Information of China (English)

    Mutluay Arslan; Ulu(c) Yi(s); Hande (C)a(g)layan; R1dvan Akin

    2013-01-01

    Dravet syndrome is a rare epileptic encephalopathy characterized by frequent seizures beginning in the first year of life and behavioral disorders. Mutations in the sodium channel α1 subunit gene are the main cause of this disease. We report two patients with refractory seizures and psychomotor retardation in whom the final diagnosis was Dravet syndrome with confirmed mutations in the sodium channel α1 subunit gene. The mutation identified in the second patient was a novel frame shift mutation, which resulted from the deletion of five nucleotides in exon 24.

  8. Genetic diversity of the VP1/VP2 gene of canine parvovirus type 2b amplified from clinical specimens in Brazil Diversidade genética no gene VP1/VP2 do parvovirus canino tipo 2b amplificado de material clínico no Brasil

    OpenAIRE

    Cesar A. D. Pereira; Edison Luiz Durigon

    2000-01-01

    We evaluated the genetic diversity in the VP1/VP2 gene of CPV type 2b isolates from symptomatic dogs in Brazil. A total of 21 isolates collected from 1990 through 1995 previously typed as CPV2b by PCR assay were studied. Overall we found a high degree of similarity among sequences from different CPV clinical isolates collected. Genetic analysis of this selected region gave no indication of a specific Brazilian parvovirus lineage.Neste estudo foi avaliada a diversidade genética no gene VP1/VP2...

  9. Dynamic expression of genes encoding subunits of inward rectifier potassium (Kir) channels in the yellow fever mosquito Aedes aegypti.

    Science.gov (United States)

    Yang, Zhongxia; Statler, Bethanie-Michelle; Calkins, Travis L; Alfaro, Edna; Esquivel, Carlos J; Rouhier, Matthew F; Denton, Jerod S; Piermarini, Peter M

    2017-02-01

    Inward rectifier potassium (Kir) channels play fundamental roles in neuromuscular, epithelial, and endocrine function in mammals. Recent research in insects suggests that Kir channels play critical roles in the development, immune function, and excretory physiology of fruit flies and/or mosquitoes. Moreover, our group has demonstrated that mosquito Kir channels may serve as valuable targets for the development of novel insecticides. Here we characterize the molecular expression of 5 mRNAs encoding Kir channel subunits in the yellow fever mosquito, Aedes aegypti: Kir1, Kir2A-c, Kir2B, Kir2B', and Kir3. We demonstrate that 1) Kir mRNA expression is dynamic in whole mosquitoes, Malpighian tubules, and the midgut during development from 4th instar larvae to adult females, 2) Kir2B and Kir3 mRNA levels are reduced in 4th instar larvae when reared in water containing an elevated concentration (50mM) of KCl, but not NaCl, and 3) Kir mRNAs are differentially expressed in the Malpighian tubules, midgut, and ovaries within 24h after blood feeding. Furthermore, we provide the first characterization of Kir mRNA expression in the anal papillae of 4th instar larval mosquitoes, which indicates that Kir2A-c is the most abundant. Altogether, the data provide the first comprehensive characterization of Kir mRNA expression in Ae. aegypti and offer insights into the putative physiological roles of Kir subunits in this important disease vector.

  10. [Polymorphisms of 2B-adrenergic receptor and endothelial NO-Synthase genes in genesis of the hereditary sick sinus node syndrome].

    Science.gov (United States)

    Chernova, A A; Nikulina, S Iu; Shul'man, V A; Kukushkina, T S; Voevoda, M I; Maksimov, V N

    2011-01-01

    In this work we have demonstrated for the first time on the clinico-genetic material association between hereditary sick sinus node syndrome (SSNS) ADRA2B and eNOS genes polymorphisms. We have established predominance of homozygote genotype of more rare DD allele in patients with SSNS (28%) compared with subjects of control group (8.99%). We have found predominance of heterozygote genotype 4a/4b in patients with SSNS compared with subjects of control group (41.8 and 25.39%, respectively). The data obtained allow to suggest that eNOS gene polymorphism might be associated with SSNS.

  11. Alanine scanning of cucumber mosaic virus (CMV 2b protein identifies different positions for cell-to-cell movement and gene silencing suppressor activity.

    Directory of Open Access Journals (Sweden)

    Katalin Nemes

    Full Text Available The multifunctional 2b protein of CMV has a role in the long distance and local movement of the virus, in symptom formation, in evasion of defense mediated by salicylic acid as well as in suppression of RNA silencing. The role of conserved amino acid sequence domains were analyzed previously in the protein function, but comprehensive analysis of this protein was not carried out until recently. We have analyzed all over the 2b protein by alanine scanning mutagenesis changing three consecutive amino acids (aa to alanine. We have identified eight aa triplets as key determinants of the 2b protein function in virus infection. Four of them (KKQ/22-24/AAA, QNR/31-33/AAA, RER/34-36/AAA, SPS/40-42/AAA overlap with previously determined regions indispensable in gene silencing suppressor function. We have identified two additional triplets necessary for the suppressor function of the 2b protein (LPF/55-57/AAA, NVE/10-12/AAA, and two other positions were required for cell-to-cell movement of the virus (MEL/1-3/AAA, RHV/70-72/AAA, which are not essential for suppressor activity.

  12. THE POLYMORPHISM OF Α2B-ADRENERGIC RECEPTOR GENE — A NEW GENETIC MARKER OF THE HEREDITARY SICK SINUS SYNDROME

    Directory of Open Access Journals (Sweden)

    S. Iu. Nikulina

    2010-01-01

    Full Text Available Aim. To study the association of the hereditary sick sinus syndrome (SSS with gene α2B-adrenergic receptor (ADRA2B polymorphism.Material and methods. 29 families with hereditary primary SSS from the database of the Chair of Therapy № 1 of Krasnoyarsk State Medical University named after prof. V.F. Voyno-Yasenetsky were included in the study. Group 1 included probands (20 women and 9 men, 58±0.15 y.o., group 2 – proband relatives of I, II and III degree (65 males and 68 females, 39±0.13 y.o., group 3 (control — 89 healthy volunteers. Clinical examination (physical examination, ECG, bicycle ergometry, ECG monitoring, atropine test, electrophysiological study, echocardiography was performed in all probands and their relatives. The diagnosis of SSS was confirmed by transesophageal left atrium stimulation in 75 individuals. Genotypic examination of gene ADRA2B I/D polymorphism was performed in 213 individuals: 75 SSS-patients, 49 their healthy relatives, 89 healthy volunteers.Results. 3 types of ADRA2B genotypes (II — homozygous wild, ID — heterozygous, DD — homozygous mutant were founded by allele-specific polymerase chain reaction. Significant prevalence of the homozygous genotype of more rare alleles DD in SSS-patients (28±5.2% compared to the control group (8.99±3.0% was found.Conclusion. Study of the genetic marker can be used to identify predisposition to hereditary SSS in the population and individual-family level. SSS due to mutations in genes that regulate cell function of sinus node and the sinoatrial conduct occurs, apparently, extremely rarely.

  13. Epigenetic mechanisms underlying the dynamic expression of cancer-testis genes, PAGE2, -2B and SPANX-B, during mesenchymal-to-epithelial transition.

    Directory of Open Access Journals (Sweden)

    Sinem Yilmaz-Ozcan

    Full Text Available Cancer-testis (CT genes are expressed in various cancers but not in normal tissues other than in cells of the germline. Although DNA demethylation of promoter-proximal CpGs of CT genes is linked to their expression in cancer, the mechanisms leading to demethylation are unknown. To elucidate such mechanisms we chose to study the Caco-2 colorectal cancer cell line during the course of its spontaneous differentiation in vitro, as we found CT genes, in particular PAGE2, -2B and SPANX-B, to be up-regulated during this process. Differentiation of these cells resulted in a mesenchymal-to-epithelial transition (MET as evidenced by the gain of epithelial markers CDX2, Claudin-4 and E-cadherin, and a concomitant loss of mesenchymal markers Vimentin, Fibronectin-1 and Transgelin. PAGE2 and SPAN-X up-regulation was accompanied by an increase in Ten-eleven translocation-2 (TET2 expression and cytosine 5-hydroxymethylation as well as the disassociation of heterochromatin protein 1 and the polycomb repressive complex 2 protein EZH2 from promoter-proximal regions of these genes. Reversal of differentiation resulted in down-regulation of PAGE2, -2B and SPANX-B, and induction of epithelial-to-mesenchymal transition (EMT markers, demonstrating the dynamic nature of CT gene regulation in this model.

  14. The Osr1 and Osr2 genes act in the pronephric anlage downstream of retinoic acid signaling and upstream of Wnt2b to maintain pectoral fin development.

    Science.gov (United States)

    Neto, Ana; Mercader, Nadia; Gómez-Skarmeta, José Luis

    2012-01-01

    Vertebrate odd-skipped related genes (Osr) have an essential function during the formation of the intermediate mesoderm (IM) and the kidney structures derived from it. Here, we show that these genes are also crucial for limb bud formation in the adjacent lateral plate mesoderm (LPM). Reduction of zebrafish Osr function impairs fin development by the failure of tbx5a maintenance in the developing pectoral fin bud. Osr morphant embryos show reduced wnt2b expression, and increasing Wnt signaling in Osr morphant embryos partially rescues tbx5a expression. Thus, Osr genes control limb bud development in a non-cell-autonomous manner, probably through the activation of Wnt2b. Finally, we demonstrate that Osr genes are downstream targets of retinoic acid (RA) signaling. Therefore, Osr genes act as a relay within the genetic cascade of fin bud formation: by controlling the expression of the signaling molecule Wnt2ba in the IM they play an essential function transmitting the RA signaling originated in the somites to the LPM.

  15. Nicotinic Acetylcholine Receptor α4 Subunit Gene Variation Associated with Attention Deficit Hyperactivity Disorder

    Institute of Scientific and Technical Information of China (English)

    HUANG Xuezhu; XU Yong; LI Qianqian; LIU Pozi; YANG Yuan; ZHANG Fuquan; GUO Tianyou; YANG Chuang; GUO Lanting

    2009-01-01

    Previous pharmacological, human genetics, and animal models have implicated the nicotinic ace-tylcholine receptor a4 subunit (CHRNA4) gene in the pathogenesis of attention deficit/hyperactivity disorder (ADHD). The objective of this study is to examine the genetic association between single nucleotide poly-morphisms in the CHRNA4 gene (rs2273502, rs1044396, rs1044397, and rs3827020 loci) and ADHD. Both case-control and family-based designs are used. Children aged 6 to 16 years were interviewed and as-sessed with the children behavior checklist and the revised conner' parent rating scale to identify probands. No significant differences in the frequency distribution of genotypes or alleles were found between the case and control groups. However, further haplotype analyses showed the CCGG haplotype on dsk for ADHD in 164 case-control samples and the standard transmission disequilibrium test analyses suggest that the allele C of rs2273502 was over-transferred in 98 ADHD parent-offspring tdos. These findings suggest that the CHRNA4 gene may play a role in the pathogenesis of ADHD.

  16. Molecular cloning and expression analysis of a novel BCCP subunit gene from Aleurites moluccana.

    Science.gov (United States)

    Xuan, W Y; Zhang, Y; Liu, Z Q; Feng, D; Luo, M Y

    2015-08-19

    Aleurites moluccana L. is grown as a roadside tree in southern China and the oil content of its seed is higher than other oil plants, such as Jatropha curcas and Camellia oleifera. A. moluccana is considered a promising energy plant because its seed oil could be used to produce biodiesel and bio-jet fuel. In addition, the bark, leaves, and kernels of A. moluccana have various medical and commercial uses. Here, a novel gene coding the biotin carboxyl carrier protein subunit (BCCP) was cloned from A. moluccana L. using the homology cloning method combined with rapid amplification of cDNA end (RACE) technology. The isolated full-length cDNA sequence (designated AM-accB) was 1188 bp, containing a 795-bp open reading frame coding for 265 amino acids. The deduced amino acid sequence of AM-accB contained a biotinylated domain located between amino acids 190 and 263. A. moluccana BCCP shows high identity at the amino acid level to its homologues in other higher plants, such as Vernicia fordii, J. curcas, and Ricinus communis (86, 77, and 70%, respectively), which all contain conserved domains for ACCase activity. The expression of the AM-accB gene during the middle stage of development and maturation in A. moluccana seeds was higher than that in early and later stages. The expression pattern of the AM-accB gene is very similar to that of the oil accumulation rate.

  17. Nuclear life of the voltage-gated Cacnb4 subunit and its role in gene transcription regulation.

    Science.gov (United States)

    Ronjat, Michel; Kiyonaka, Shigeki; Barbado, Maud; De Waard, Michel; Mori, Yasuo

    2013-01-01

    The pore-forming subunit of voltage-gated calcium channels is associated to auxiliary subunits among which the cytoplasmic β subunit. The different isoforms of this subunit control both the plasma membrane targeting and the biophysical properties of the channel moiety. In a recent study, we demonstrated that the Cacnb4 (β 4) isoform is at the center of a new signaling pathway that connects neuronal excitability and gene transcription. This mechanism relies on nuclear targeting of β 4 triggered by neuronal electrical stimulation. This re-localization of β 4 is promoted by its interaction with Ppp2r5d a regulatory subunit of PP2A in complex with PP2A itself. The formation, as well as the nuclear translocation, of the β 4/ Ppp2r5d/ PP2A complex is totally impaired by the premature R482X stops mutation of β 4 that has been previously associated with juvenile epilepsy. Taking as a case study the tyrosine hydroxylase gene that is strongly upregulated in brain of lethargic mice, deficient for β 4 expression, we deciphered the molecular steps presiding to this signaling pathway. Here we show that expression of wild-type β 4 in HEK293 cells results in the regulation of several genes, while expression of the mutated β 4 (β 1-481) produces a different set of gene regulation. Several genes regulated by β 4 in HEK293 cells were also regulated upon neuronal differentiation of NG108-15 cells that induces nuclear translocation of β 4 suggesting a link between β 4 nuclear targeting and gene regulation.

  18. Frequent germ-line succinate dehydrogenase subunit D gene mutations in patients with apparently sporadic parasympathetic paraganglioma

    NARCIS (Netherlands)

    H. Dannenberg (Hilde); W.N.M. Dinjens (Winand); M. Abbou; H. van Urk (Hero); B.K. Pauw; D. Mouwen; W.J. Mooi (Wolter); R.R. de Krijger (Ronald)

    2002-01-01

    textabstractPURPOSE: Recently, familial paraganglioma (PGL) was shown to be caused bymutations in the gene encoding succinate dehydrogenase subunit D (SDHD). However, the prevalence of SDHD mutations in apparently sporadic PGL is unknown. We studied the frequency and spectrum of ge

  19. A novel mutation in the succinate dehydrogenase subunit D gene in siblings with the hereditary paraganglioma-pheochromocytoma syndrome.

    Science.gov (United States)

    Prasad, Chaithra; Oakley, Gerard J; Yip, Linwah; Coyne, Christopher; Rangaswamy, Balasubramanya; Dixit, Sanjay B

    2014-01-01

    Germline mutations in the succinate dehydrogenase complex subunit D gene are now known to be associated with hereditary paraganglioma-pheochromocytoma syndromes. Since the initial succinate dehydrogenase complex subunit D gene mutation was identified about a decade ago, more than 131 unique variants have been reported. We report the case of two siblings presenting with multiple paragangliomas and pheochromocytomas; they were both found to carry a mutation in the succinate dehydrogenase complex subunit D gene involving a substitution of thymine to guanine at nucleotide 236 in exon 3. This particular mutation of the succinate dehydrogenase complex subunit D gene has only been reported in one previous patient in Japan; this is, therefore, the first report of this pathogenic mutation in siblings and the first report of this mutation in North America. With continued screening of more individuals, we will be able to create a robust mutation database that can help us understand disease patterns associated with particular variants and may be a starting point in the development of new therapies for familial paraganglioma syndromes.

  20. A novel mutation in the succinate dehydrogenase subunit D gene in siblings with the hereditary paraganglioma–pheochromocytoma syndrome

    Directory of Open Access Journals (Sweden)

    Chaithra Prasad

    2014-10-01

    Full Text Available Germline mutations in the succinate dehydrogenase complex subunit D gene are now known to be associated with hereditary paraganglioma–pheochromocytoma syndromes. Since the initial succinate dehydrogenase complex subunit D gene mutation was identified about a decade ago, more than 131 unique variants have been reported. We report the case of two siblings presenting with multiple paragangliomas and pheochromocytomas; they were both found to carry a mutation in the succinate dehydrogenase complex subunit D gene involving a substitution of thymine to guanine at nucleotide 236 in exon 3. This particular mutation of the succinate dehydrogenase complex subunit D gene has only been reported in one previous patient in Japan; this is, therefore, the first report of this pathogenic mutation in siblings and the first report of this mutation in North America. With continued screening of more individuals, we will be able to create a robust mutation database that can help us understand disease patterns associated with particular variants and may be a starting point in the development of new therapies for familial paraganglioma syndromes.

  1. A novel mutation in the succinate dehydrogenase subunit D gene in siblings with the hereditary paraganglioma–pheochromocytoma syndrome

    Science.gov (United States)

    Oakley, Gerard J; Yip, Linwah; Coyne, Christopher; Rangaswamy, Balasubramanya; Dixit, Sanjay B

    2014-01-01

    Germline mutations in the succinate dehydrogenase complex subunit D gene are now known to be associated with hereditary paraganglioma–pheochromocytoma syndromes. Since the initial succinate dehydrogenase complex subunit D gene mutation was identified about a decade ago, more than 131 unique variants have been reported. We report the case of two siblings presenting with multiple paragangliomas and pheochromocytomas; they were both found to carry a mutation in the succinate dehydrogenase complex subunit D gene involving a substitution of thymine to guanine at nucleotide 236 in exon 3. This particular mutation of the succinate dehydrogenase complex subunit D gene has only been reported in one previous patient in Japan; this is, therefore, the first report of this pathogenic mutation in siblings and the first report of this mutation in North America. With continued screening of more individuals, we will be able to create a robust mutation database that can help us understand disease patterns associated with particular variants and may be a starting point in the development of new therapies for familial paraganglioma syndromes. PMID:27489656

  2. Characterization of the low-molecular-weight glutenin subunit gene family members using a PCR-based marker approach

    Science.gov (United States)

    Low-molecular-weight glutenin subunits (LMW-GS) are a class of seed storage proteins that play a major role in the determination of the processing quality of wheat flour. The LMW-GS are encoded by multi-gene families located on the short arms of the homoeologous group 1 chromosomes, at the Glu-A3, G...

  3. Genes involved in lactose catabolism and organic acid production during growth of Lactobacillus delbrueckii UFV H2b20 in skimmed milk.

    Science.gov (United States)

    Do Carmo, A P; De Oliveira, M N V; Da Silva, D F; Castro, S B; Borges, A C; De Carvalho, A F; De Moraes, C A

    2012-03-01

    There are three main reasons for using lactic acid bacteria (LAB) as starter cultures in industrial food fermentation processes: food preservation due to lactic acid production; flavour formation due to a range of organic molecules derived from sugar, lipid and protein catabolism; and probiotic properties attributed to some strains of LAB, mainly of lactobacilli. The aim of this study was to identify some genes involved in lactose metabolism of the probiotic Lactobacillus delbrueckii UFV H2b20, and analyse its organic acid production during growth in skimmed milk. The following genes were identified, encoding the respective enzymes: ldh - lactate dehydrogenase, adhE - Ldb1707 acetaldehyde dehydrogenase, and ccpA-pepR1 - catabolite control protein A. It was observed that L. delbrueckii UFV H2b20 cultivated in different media has the unexpected ability to catabolyse galactose, and to produce high amounts of succinic acid, which was absent in the beginning, raising doubts about the subspecies in question. The phylogenetic analyses showed that this strain can be compared physiologically to L. delbrueckii subsp. bulgaricus and L. delbrueckii subsp. lactis, which are able to degrade lactose and can grow in milk. L. delbrueckii UFV H2b20 sequences have grouped with L. delbrueckii subsp. bulgaricus ATCC 11842 and L. delbrueckii subsp. bulgaricus ATCC BAA-365, strengthening the classification of this probiotic strain in the NCFM group proposed by a previous study. Additionally, L. delbrueckii UFV H2b20 presented an evolutionary pattern closer to that of probiotic Lactobacillus acidophilus NCFM, corroborating the suggestion that this strain might be considered as a new and unusual subspecies among L. delbrueckii subspecies, the first one identified as a probiotic. In addition, its unusual ability to metabolise galactose, which was significantly consumed in the fermentation medium, might be exploited to produce low-browning probiotic Mozzarella cheeses, a desirable property

  4. The roles of co-chaperone CCRP/DNAJC7 in Cyp2b10 gene activation and steatosis development in mouse livers.

    Directory of Open Access Journals (Sweden)

    Marumi Ohno

    Full Text Available Cytoplasmic constitutive active/androstane receptor (CAR retention protein (CCRP and also known as DNAJC7 is a co-chaperone previously characterized to retain nuclear receptor CAR in the cytoplasm of HepG2 cells. Here we have produced CCRP knockout (KO mice and demonstrated that CCRP regulates CAR at multiple steps in activation of the cytochrome (Cyp 2b10 gene in liver: nuclear accumulation, RNA polymerase II recruitment and epigenetic modifications. Phenobarbital treatment greatly increased nuclear CAR accumulation in the livers of KO males as compared to those of wild type (WT males. Despite this accumulation, phenobarbital-induced activation of the Cyp2b10 gene was significantly attenuated. In ChIP assays, a CAR/retinoid X receptor-α (RXRα heterodimer binding to the Cyp2b10 promoter was already increased before phenobarbital treatment and further pronounced after treatment. However, RNA polymerase II was barely recruited to the promoter even after phenobarbital treatment. Histone H3K27 on the Cyp2b10 promoter was de-methylated only after phenobarbital treatment in WT but was fully de-methylated before treatment in KO males. Thus, CCRP confers phenobarbital-induced de-methylation capability to the promoter as well as the phenobarbital responsiveness of recruiting RNA polymerase II, but is not responsible for the binding between CAR and its cognate sequence, phenobarbital responsive element module. In addition, KO males developed steatotic livers and increased serum levels of total cholesterol and high density lipoprotein in response to fasting. CCRP appears to be involved in various hepatic regulations far beyond CAR-mediated drug metabolism.

  5. Molecular evolution of the cytochrome c oxidase subunit 5A gene in primates

    Directory of Open Access Journals (Sweden)

    Wildman Derek E

    2008-01-01

    Full Text Available Abstract Background Many electron transport chain (ETC genes show accelerated rates of nonsynonymous nucleotide substitutions in anthropoid primate lineages, yet in non-anthropoid lineages the ETC proteins are typically highly conserved. Here, we test the hypothesis that COX5A, the ETC gene that encodes cytochrome c oxidase subunit 5A, shows a pattern of anthropoid-specific adaptive evolution, and investigate the distribution of this protein in catarrhine brains. Results In a dataset comprising 29 vertebrate taxa, including representatives from all major groups of primates, there is nearly 100% conservation of the COX5A amino acid sequence among extant, non-anthropoid placental mammals. The most recent common ancestor of these species lived about 100 million years (MY ago. In contrast, anthropoid primates show markedly elevated rates of nonsynonymous evolution. In particular, branch site tests identify five positively selected codons in anthropoids, and ancestral reconstructions infer that substitutions in these codons occurred predominantly on stem lineages (anthropoid, ape and New World monkey and on the human terminal branch. Examination of catarrhine brain samples by immunohistochemistry characterizes for the first time COX5A protein distribution in the primate neocortex, and suggests that the protein is most abundant in the mitochondria of large-size projection neurons. Real time quantitative PCR supports previous microarray results showing COX5A is expressed in cerebral cortical tissue at a higher level in human than in chimpanzee or gorilla. Conclusion Taken together, these results suggest that both protein structural and gene regulatory changes contributed to COX5A evolution during humankind's ancestry. Furthermore, these findings are consistent with the hypothesis that adaptations in ETC genes contributed to the emergence of the energetically expensive anthropoid neocortex.

  6. An ancient repeat sequence in the ATP synthase beta-subunit gene of forcipulate sea stars.

    Science.gov (United States)

    Foltz, David W

    2007-11-01

    A novel repeat sequence with a conserved secondary structure is described from two nonadjacent introns of the ATP synthase beta-subunit gene in sea stars of the order Forcipulatida (Echinodermata: Asteroidea). The repeat is present in both introns of all forcipulate sea stars examined, which suggests that it is an ancient feature of this gene (with an approximate age of 200 Mya). Both stem and loop regions show high levels of sequence constraint when compared to flanking nonrepetitive intronic regions. The repeat was also detected in (1) the family Pterasteridae, order Velatida and (2) the family Korethrasteridae, order Velatida. The repeat was not detected in (1) the family Echinasteridae, order Spinulosida, (2) the family Astropectinidae, order Paxillosida, (3) the family Solasteridae, order Velatida, or (4) the family Goniasteridae, order Valvatida. The repeat lacks similarity to published sequences in unrestricted GenBank searches, and there are no significant open reading frames in the repeat or in the flanking intron sequences. Comparison via parametric bootstrapping to a published phylogeny based on 4.2 kb of nuclear and mitochondrial sequence for a subset of these species allowed the null hypothesis of a congruent phylogeny to be rejected for each repeat, when compared separately to the published phylogeny. In contrast, the flanking nonrepetitive sequences in each intron yielded separate phylogenies that were each congruent with the published phylogeny. In four species, the repeat in one or both introns has apparently experienced gene conversion. The two introns also show a correlated pattern of nucleotide substitutions, even after excluding the putative cases of gene conversion.

  7. Function of Partially Duplicated Human α7 Nicotinic Receptor Subunit CHRFAM7A Gene

    Science.gov (United States)

    de Lucas-Cerrillo, Ana M.; Maldifassi, M. Constanza; Arnalich, Francisco; Renart, Jaime; Atienza, Gema; Serantes, Rocío; Cruces, Jesús; Sánchez-Pacheco, Aurora; Andrés-Mateos, Eva; Montiel, Carmen

    2011-01-01

    The neuronal α7 nicotinic receptor subunit gene (CHRNA7) is partially duplicated in the human genome forming a hybrid gene (CHRFAM7A) with the novel FAM7A gene. The hybrid gene transcript, dupα7, has been identified in brain, immune cells, and the HL-60 cell line, although its translation and function are still unknown. In this study, dupα7 cDNA has been cloned and expressed in GH4C1 cells and Xenopus oocytes to study the pattern and functional role of the expressed protein. Our results reveal that dupα7 transcript was natively translated in HL-60 cells and heterologously expressed in GH4C1 cells and oocytes. Injection of dupα7 mRNA into oocytes failed to generate functional receptors, but when co-injected with α7 mRNA at α7/dupα7 ratios of 5:1, 2:1, 1:1, 1:5, and 1:10, it reduced the nicotine-elicited α7 current generated in control oocytes (α7 alone) by 26, 53, 75, 93, and 94%, respectively. This effect is mainly due to a reduction in the number of functional α7 receptors reaching the oocyte membrane, as deduced from α-bungarotoxin binding and fluorescent confocal assays. Two additional findings open the possibility that the dominant negative effect of dupα7 on α7 receptor activity observed in vitro could be extrapolated to in vivo situations. (i) Compared with α7 mRNA, basal dupα7 mRNA levels are substantial in human cerebral cortex and higher in macrophages. (ii) dupα7 mRNA levels in macrophages are down-regulated by IL-1β, LPS, and nicotine. Thus, dupα7 could modulate α7 receptor-mediated synaptic transmission and cholinergic anti-inflammatory response. PMID:21047781

  8. ASSIGNMENT OF THE GENE CODING FOR HUMAN CYTOCHROME-C-OXIDASE SUBUNIT-VIB TO CHROMOSOME-19, BAND-Q13.1, BY FLUORESCENCE INSITU HYBRIDIZATION

    NARCIS (Netherlands)

    TAANMAN, JW; VANDERVEEN, AY; SCHRAGE, C; DEVRIES, H; BUYS, CHCM

    1991-01-01

    A cloned, 40 kb, genomic DNA fragment, containing the last exon of the gene for human cytochrome c oxidase subunit VIb and its flanking sequences, was used as a probe to localize the subunit VIb gene on human metaphase chromosomes. The probe was labelled with Bio-11-dUTP and detected by fluorescence

  9. Gene for the catalytic subunit of mouse DNA-dependent protein kinase maps to the scid locus.

    Science.gov (United States)

    Miller, R D; Hogg, J; Ozaki, J H; Gell, D; Jackson, S P; Riblet, R

    1995-01-01

    The gene encoding the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs) has been proposed recently as a candidate gene for the mouse severe combined immune deficiency (scid) locus. We have used a partial cDNA clone for human DNA-PKcs to map the mouse homologue using a large interspecific backcross panel. We found that the mouse gene for DNA-PKcs does not recombine with scid, consistent with the hypothesis that scid is a mutation in the mouse gene for DNA-PKcs. Images Fig. 3 PMID:7479885

  10. Polymorphisms of estrogen metabolism-related genes ESR1 , UGT2B17 , and UGT1A1 are not associated with osteoporosis in artificial menopausal Japanese women

    Directory of Open Access Journals (Sweden)

    Megumi Yokota

    2015-09-01

    Full Text Available Introduction : Bilateral salpingo-oophorectomy (BSO is a risk factor for osteoporosis. Previous studies have reported an association between genetic polymorphisms and the risk of developing osteoporosis. However, the relationship between osteoporosis and genetic polymorphisms in Japanese women treated with BSO is not well understood. To improve the quality of life for post-BSO patients, it is important to determine the genetic factors that influence their risk for osteoporosis. The aim of this study was to investigate the association between gene variations of estrogen metabolism-related genes and osteoporosis in surgically menopausal patients, which may improve the quality of life for surgically menopausal patients. Material and methods : This study included 203 menopausal women treated with BSO because of gynecologic disorders. One hundred and twenty-six women with artificial (surgical menopause, who had undergone BSO in the premenopausal period, were compared with 77 women with natural menopause, who had undergone BSO in the postmenopausal period. The women were tested for bone mineral density to diagnose osteoporosis. Polymorphisms of estrogen receptor 1 ( ESR1 and UDP-glucuronosyl transferase (UGT genes UGT2B17 and UGT1A1 were analyzed, and their association with bone mass and osteoporosis was statistically evaluated. Results : No significant association was found between osteoporosis and polymorphisms in ESR1 , UGT2B17 , or UGT1A1 in both groups, suggesting that BSO might be a more significant physiological factor in influencing bone mass density compared to genetic variations. Conclusions : These results suggest that the ESR1 , UGT2B17 , and UGT1A1 polymorphisms are not genetic factors affecting osteoporosis in postmenopausal Japanese women.

  11. Phylogenetic relationships of Mongolian Babesia bovis isolates based on the merozoite surface antigen (MSA)-1, MSA-2b, and MSA-2c genes.

    Science.gov (United States)

    Altangerel, Khukhuu; Sivakumar, Thillaiampalam; Battsetseg, Badgar; Battur, Banzragch; Ueno, Akio; Igarashi, Ikuo; Yokoyama, Naoaki

    2012-03-23

    We conducted a molecular epidemiological study on Babesia bovis in Mongolia. Three hundred blood samples collected from cattle grazed in seven different districts were initially screened using a previously established diagnostic polymerase chain reaction (PCR) assay for the detection of B. bovis-specific DNA. Positive samples were then used to amplify and sequence the hyper-variable regions of three B. bovis genes encoding the merozoite surface antigen (MSA)-1, MSA-2b, and MSA-2c. The diagnostic PCR assay detected B. bovis among cattle populations of all districts surveyed (4.4-26.0%). Sequences of each of the three genes were highly homologous among the Mongolian isolates, and found in a single phylogenetic cluster. In particular, a separate branch was formed only by the Mongolian isolates in the MSA-2b gene-based phylogenetic tree. Our findings indicate that effective preventative and control strategies are essential to control B. bovis infection in Mongolian cattle populations, and suggest that a careful approach must be adopted when using immunization techniques.

  12. Molecular evolution at the cytochrome oxidase subunit 2 gene among divergent populations of the intertidal copepod, Tigriopus californicus.

    Science.gov (United States)

    Rawson, Paul D; Burton, Ronald S

    2006-06-01

    The cytochrome c oxidase subunit 2 gene (COII) encodes a highly conserved protein that is directly responsible for the initial transfer of electrons from cytochrome c to cytochrome c oxidase (COX) crucial to the production of ATP during cellular respiration. Despite its integral role in electron transport, we have observed extensive intraspecific nucleotide and amino acid variation among 26 full-length COII sequences sampled from seven populations of the marine copepod, Tigriopus californicus. Although intrapopulation divergence was virtually nonexistent, interpopulation divergence at the COII locus was nearly 20% at the nucleotide level, including 38 nonsynonymous substitutions. Given the high degree of interaction between the cytochrome c oxidase subunit 2 protein (COX2) and the nuclear-encoded subunits of COX and cytochrome c (CYC), we hypothesized that some codons in the COII gene are likely to be under positive selection in order to compensate for amino acid substitutions in other subunits. Estimates of the ratio of nonsynonymous to synonymous substitution (omega), obtained using a series of maximum likelihood models of codon substitution, indicated that the majority of codons in T. californicus COII are under strong purifying selection (omega < 1), while approximately 4% of the sites in this gene appear to evolve under relaxed selective constraint (omega = 1). A branch-site maximum likelihood model identified three sites that may have experienced positive selection within the central California sequence clade in our COII phylogeny; these results are consistent with previous studies showing functional and fitness consequences among interpopulation hybrids between central and northern California populations.

  13. Phosphatidylinositol 3-kinase p85 regulatory subunit gene and spinal muscular atrophy disease

    Directory of Open Access Journals (Sweden)

    Monica STAVARACHI

    2009-11-01

    Full Text Available Spinal muscular atrophy (SMA is a frequent neuromuscular disorder caused by motoneuronal apoptosis, as a result of SMN (Survival Motor Neuron protein deficiency. Although the SMA determining gene was identified, the molecular mechanism of the disease is not clearly understood, due to the heterogeneity of clinical manifestations. Trying to complete the molecular describing SMA picture, by identifying potential modulators factors, we investigated the relationship between phosphatidylinositol 3-kinase p85 regulatory subunit gene (PIK3R1 and SMA pathology. As IGF signaling pathway has been reported to play an important role in motoneurons survival and PIK3 is a key element of this cascade signaling, we focused on the relationship between PIK3R1 gene Met326Ile polymorphism and SMA type I, the most severe form of the disease. A total of 80 subjects (40 SMA type I patients and 40 unrelated healthy controls were included in the study. The statistical analyzes performed consequently to the genotyping by mismatch PCR-RFLP method, revealed that Met326Ile polymorphism is not associated with SMA type I disease: ORMet/Met = 0.398 with a p = 0.072 meanwhile ORMet = 0.495, p = 0.063. However, the Cochrane – Armitage test indicated that there is a statistically association trend between the analyzed polymorphism and SMA type I pathology: ORMet = 0.438, p = 0.032. We concluded that additional researches with an increased subjects number and replicates studies in other populations will clarify the investigated relationship and it may contribute to the SMA molecular mechanism understanding.

  14. Comparison of osteogenic potentials of human rat BMP4 and BMP6 gene therapy using [E1-] and [E1-,E2b-] adenoviral vectors

    Directory of Open Access Journals (Sweden)

    Hongwei Li, Jin Zhong Li, Debra D. Pittman, Andy Amalfitano, Gerald R. Hankins, Gregory A. Helm

    2006-01-01

    Full Text Available Osteogenic potentials of some recombinant human bone morphogenetic protein (BMP first-generation adenoviral vectors (ADhBMPs are significantly limited in immunocompetent animals. It is unclear what role expression of viral proteins and foreign proteins transduced by adenoviral vectors play in the host immune response and in ectopic bone formation. In this study two sets of experiments were designed and performed. First, rat BMP6 cDNA were amplified, sequenced, and recombined in first-generation adenoviral vector (ADrBMP6. A comparison of human and rat BMP6 adenoviral vectors demonstrated identical osteogenic activities in both immunodeficient and immunocompetent rats. Second, the activities of recombinant human BMP6 in E1- (ADhBMP6 and [E1-,E2b-] ( [E1-,E2b-]ADGFP&hBMP6, and [E1-,E2b-]ADhBMP6 adenoviral vectors were compared in both in vitro and in vivo models. Similar activities of these two generations of BMP adenoviral vectors were found in all models. These results indicate that the amount of viral gene expression and the source of the BMP cDNA are not major factors in the interruption of osteogenic potentials of recombinant BMP6 adenoviral vectors in immunocompetent animals.

  15. Spt10 and Swi4 Control the Timing of Histone H2A/H2B Gene Activation in Budding Yeast ▿

    OpenAIRE

    Eriksson, Peter R.; Ganguli, Dwaipayan; Clark, David J.

    2010-01-01

    The expression of the histone genes is regulated during the cell cycle to provide histones for nucleosome assembly during DNA replication. In budding yeast, histones H2A and H2B are expressed from divergent promoters at the HTA1-HTB1 and HTA2-HTB2 loci. Here, we show that the major activator of HTA1-HTB1 is Spt10, a sequence-specific DNA binding protein with a putative histone acetyltransferase (HAT) domain. Spt10 binds to two pairs of upstream activation sequence (UAS) elements in the HTA1-H...

  16. [The association between the GRIN2B gene and verbal fluency and impairment of abstract thinking in schizophrenia].

    Science.gov (United States)

    Alfimova, M V; Golimbet, V E; Korovaitseva, G I; Abramova, L I; Lezheiko, T V; Aksenova, E V

    2016-01-01

    Цель исследования. Исследование было направлено на поиск ассоциаций между геном GRIN2B и признаками нарушения мышления и речи при шизофрении, в основе которых может лежать снижение доступа к ментальному лексикону. Материал и методы. В группе из 552 пациентов с расстройствами шизофренического спектра определяли связи между полиморфизмом rs7301328 в гене GRIN2B и семантической вербальной беглостью и пятью симптомами нарушений мышления и речи, входящими в шкалу позитивных и негативных симптомов (PANSS). Результаты и обсуждение. Обнаружена ассоциация гена GRIN2B как с вербальной беглостью (p=0,013), так и нарушением абстрактного мышления (p=0,012). При этом не выявлено опосредующей роли вербальной беглости в ассоциации между геном и нарушением мышления. Результаты позволяют предположить, что ген GRIN2B оказывает модифицирующее действие на лингвистические процессы, обеспечивающие извлечение информации из ментального лексикона на основе семантических признаков, и, кроме того, вносит вклад в вариативность клинически выраженных нарушений абстрактного мышления у больных шизофренией. При этом гете

  17. Differential expression of genes encoding neuronal ion-channel subunits in major depression, bipolar disorder and schizophrenia: implications for pathophysiology.

    Science.gov (United States)

    Smolin, Bella; Karry, Rachel; Gal-Ben-Ari, Shunit; Ben-Shachar, Dorit

    2012-08-01

    Evidence concerning ion-channel abnormalities in the pathophysiology of common psychiatric disorders is still limited. Given the significance of ion channels in neuronal activity, neurotransmission and neuronal plasticity we hypothesized that the expression patterns of genes encoding different ion channels may be altered in schizophrenia, bipolar and unipolar disorders. Frozen samples of striatum including the nucleus accumbens (Str-NAc) and the lateral cerebellar hemisphere of 60 brains from depressed (MDD), bipolar (BD), schizophrenic and normal subjects, obtained from the Stanley Foundation Brain Collection, were assayed. mRNA of 72 different ion-channel subunits were determined by qRT-PCR and alteration in four genes were verified by immunoblotting. In the Str-NAc the prominent change was observed in the MDD group, in which there was a significant up-regulation in genes encoding voltage-gated potassium-channel subunits. However, in the lateral cerebellar hemisphere (cerebellum), the main change was observed in schizophrenia specimens, as multiple genes encoding various ion-channel subunits were significantly down-regulated. The impaired expression of genes encoding ion channels demonstrates a disease-related neuroanatomical pattern. The alterations observed in Str-NAc of MDD may imply electrical hypo-activity of this region that could be of relevance to MDD symptoms and treatment. The robust unidirectional alteration of both excitatory and inhibitory ion channels in the cerebellum may suggests cerebellar general hypo-transcriptional activity in schizophrenia.

  18. Familial Congenital Hypothyroidism Caused by Abnormal and Bioinactive TSH due to Mutations in the beta-Subunit Gene.

    Science.gov (United States)

    Medeiros-Neto, G; de Lacerda, L; Wondisford, F E

    1997-01-01

    Hereditary TSH deficiency is a rare autosomal recessive disease described in inbred Japanese families and in Greek and Brazilian kindreds. The TSH-beta-subunit gene has been shown to be the site of mutations that will give rise to truncated proteins that cannot dimerize with the alpha subunit or, alternatively, will produce a mutated TSH that is present in the circulation of the affected patients, but it is biologically inactive. Characteristically, the patients with TSH-beta-subunit-defects are born with congenital hypothyroidism, with very low levels of serum thyroid hormones and serum thyroglobulin and, paradoxically, with serum TSH levels that are consistently undetectable or at very low levels. Goiter is not present at birth, but the low radioactive thyroid uptake will increase after bovine TSH stimulation. Other pituitary hormones responses to provocative tests are normal. The subunit levels are at high concentration and are significantly increased following TRH stimulation. In two kindreds, molecular biological studies have indicated mutations in two different sites of exon 2, generating a peptide that would not dimerize with subunits to synthesize TSH molecules. In one kindred, a truncated TSH-beta protein was translated that generated a biologically inactive but detectable serum TSH molecule. (c) 1997, Elsevier Science Inc. (Trends Endocrinol Metab 1997;8:15-20).

  19. Mutation analysis of genes that control the G1/S cell cycle in melanoma: TP53, CDKN1A, CDKN2A, and CDKN2B

    Directory of Open Access Journals (Sweden)

    López-Nevot Miguel

    2005-04-01

    Full Text Available Abstract Background The role of genes involved in the control of progression from the G1 to the S phase of the cell cycle in melanoma tumors in not fully known. The aim of our study was to analyse mutations in TP53, CDKN1A, CDKN2A, and CDKN2B genes in melanoma tumors and melanoma cell lines Methods We analysed 39 primary and metastatic melanomas and 9 melanoma cell lines by single-stranded conformational polymorphism (SSCP. Results The single-stranded technique showed heterozygous defects in the TP53 gene in 8 of 39 (20.5% melanoma tumors: three new single point mutations in intronic sequences (introns 1 and 2 and exon 10, and three new single nucleotide polymorphisms located in introns 1 and 2 (C to T transition at position 11701 in intron 1; C insertion at position 11818 in intron 2; and C insertion at position 11875 in intron 2. One melanoma tumor exhibited two heterozygous alterations in the CDKN2A exon 1 one of which was novel (stop codon, and missense mutation. No defects were found in the remaining genes. Conclusion These results suggest that these genes are involved in melanoma tumorigenesis, although they may be not the major targets. Other suppressor genes that may be informative of the mechanism of tumorigenesis in skin melanomas should be studied.

  20. PHYLOGENY OF ANGIOSTRONGYLUS CANTONENSIS IN THAILAND BASED ON CYTOCHROME C OXIDASE SUBUNIT I GENE SEQUENCE.

    Science.gov (United States)

    Apichat, Vitta; Narongrit, Srisongcram; Jittranuch, Thiproaj; Anucha, Wongma; Wilaiwan, Polsut; Chamaiporn, Fukruksa; Thatcha, Yimthin; Bandid, Mangkit; Aunchalee, Thanwisai; Paron, Dekumyoy

    2016-05-01

    Angiostrongylus cantonensis is an emerging infectious agent causing eosinophilic meningitis or meningoencephalitis in humans with clinical manifestation of severe headache. Molecular genetic studies on classification and phylogeny of A. cantonensis in Thailand are limited. This study surveyed A. cantonensis larvae prevalence in natural intermediate hosts across Thailand and analyzed their phylogenetic relationships. A total of 14,032 freshwater and land snails were collected from 19 provinces of Thailand. None of Filopaludina sp, Pomacea sp, and Cyclophorus sp were infected with Angiostrongylus larvae, whereas Achatina fulica, Cryptozona siamensis, and Megaustenia siamensis collected from Kalasin, Kamphaeng Phet, Phetchabun, Phitsanulok, and Tak Provinces were infected, with C. siamensis being the common intermediate host. Based on morphology, larvae isolated from 11 samples of these naturally infected snails preliminarily were identified as A. cantonensis. Comparison of partial nucleotide sequences of cytochrome c oxidase subunit I gene revealed that four sequences are identical to A. cantonensis haplotype ac4 from Bangkok and the other seven to that of A. cantonensis isolate AC Thai, indicating two independent lineages of A. cantonensis in Thailand.

  1. Cloning of the Gene Encoding Urease Subunit A in Helicobacter Pylori

    Institute of Scientific and Technical Information of China (English)

    施理; 张宜俊; 陈劼; 候晓华

    2004-01-01

    Summary: The gene encoding urease subunit A (ureA) of Helicobacter pylori (H. pylori) was cloned from H. pylori isolate by polymerase chain reaction (PCR). Sterile distilled water instead of DNA served as negative control. The nucleotide sequence of the amplified product was determined.Homologous analysis of the ureA against that reported by Clayton CL and the GenBank and SwissProt databases were performed with the BLAST program at the Genome Net through the Internet.0.8 kb PCR product was amplified from all H. pylori clinical isolators. The nucleotide sequence of the ureA was determined. The nucleotide sequence of the ureA began with ATG as the initiation codon and terminated in TAA as stop codon. The coding regions had a 44 % G+ C content. The DNA sequence was 98 % homologous to that reported by Clayton CL (688 out of 702 residues were identical). The derived amino-acid sequences of the ureA were 99 % homologous to that reported by Clayton CL (232 out of 234 residues were identical). The nucleotide sequence and the predicted protein showed significant homology to ureA of H. pylori in the NCBI Entrez database.

  2. Genetic diversity of the VP1/VP2 gene of canine parvovirus type 2b amplified from clinical specimens in Brazil Diversidade genética no gene VP1/VP2 do parvovirus canino tipo 2b amplificado de material clínico no Brasil

    Directory of Open Access Journals (Sweden)

    Cesar A. D. Pereira

    2000-10-01

    Full Text Available We evaluated the genetic diversity in the VP1/VP2 gene of CPV type 2b isolates from symptomatic dogs in Brazil. A total of 21 isolates collected from 1990 through 1995 previously typed as CPV2b by PCR assay were studied. Overall we found a high degree of similarity among sequences from different CPV clinical isolates collected. Genetic analysis of this selected region gave no indication of a specific Brazilian parvovirus lineage.Neste estudo foi avaliada a diversidade genética no gene VP1/VP2 do parvovírus canino tipo 2b a partir de amostras isoladas de cães sintomáticos no Brasil. Foram estudadas 21 amostras coletadas no período de 1990 à 1995, previamente caracterizadas como CPV 2b pela técnica de PCR. Observou-se alto grau de similaridade entre as seqüências estudadas e a análise genética da região selecionada não indicou a presença de uma linhagem brasileira específica.

  3. The role of GluN2A and GluN2B NMDA receptor subunits in AgRP and POMC neurons on body weight and glucose homeostasis

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    Aykut Üner

    2015-10-01

    Conclusions: GluN2B-containing NMDA receptors in AgRP neurons play a critical role in central control of body weight homeostasis and blood glucose balance via mechanisms that likely involve regulation of AgRP neuronal survival and structure, and modulation of hypothalamic leptin action.

  4. Amplification of TLO Mediator Subunit Genes Facilitate Filamentous Growth in Candida Spp.

    Science.gov (United States)

    Liu, Zhongle; Moran, Gary P.; Myers, Lawrence C.

    2016-01-01

    Filamentous growth is a hallmark of C. albicans pathogenicity compared to less-virulent ascomycetes. A multitude of transcription factors regulate filamentous growth in response to specific environmental cues. Our work, however, suggests the evolutionary history of C. albicans that resulted in its filamentous growth plasticity may be tied to a change in the general transcription machinery rather than transcription factors and their specific targets. A key genomic difference between C. albicans and its less-virulent relatives, including its closest relative C. dubliniensis, is the unique expansion of the TLO (TeLOmere-associated) gene family in C. albicans. Individual Tlo proteins are fungal-specific subunits of Mediator, a large multi-subunit eukaryotic transcriptional co-activator complex. This amplification results in a large pool of ‘free,’ non-Mediator associated, Tlo protein present in C. albicans, but not in C. dubliniensis or other ascomycetes with attenuated virulence. We show that engineering a large ‘free’ pool of the C. dubliniensis Tlo2 (CdTlo2) protein in C. dubliniensis, through overexpression, results in a number of filamentation phenotypes typically associated only with C. albicans. The amplitude of these phenotypes is proportional to the amount of overexpressed CdTlo2 protein. Overexpression of other C. dubliniensis and C. albicans Tlo proteins do result in these phenotypes. Tlo proteins and their orthologs contain a Mediator interaction domain, and a potent transcriptional activation domain. Nuclear localization of the CdTlo2 activation domain, facilitated naturally by the Tlo Mediator binding domain or artificially through an appended nuclear localization signal, is sufficient for the CdTlo2 overexpression phenotypes. A C. albicans med3 null mutant causes multiple defects including the inability to localize Tlo proteins to the nucleus and reduced virulence in a murine systemic infection model. Our data supports a model in which the

  5. Sequential mutations in the interleukin-3 (IL3)/granulocyte-macrophage colony-stimulating factor/IL5 receptor beta-subunit genes are necessary for the complete conversion to growth autonomy mediated by a truncated beta C subunit.

    Science.gov (United States)

    Hannemann, J; Hara, T; Kawai, M; Miyajima, A; Ostertag, W; Stocking, C

    1995-05-01

    An amino-terminally truncated beta C receptor (beta C-R) subunit of the interleukin-3 (IL3)/granulocyte-macrophage colony-stimulating factor/IL5 receptor complex mediates factor-independent and tumorigenic growth in two spontaneous mutants of a promyelocytic cell line. The constitutive activation of the JAK2 protein kinase in these mutants confirms that signaling occurs through the truncated receptor protein. Noteworthily, in addition to a 10-kb deletion in the beta C-R subunit gene encoding the truncated receptor, several secondary and independent mutations that result in the deletion or functional inactivation of the allelic beta C-R subunit and the closely related beta IL3-R subunit genes were observed in both mutants, suggesting that such mutations are necessary for the full oncogenic penetrance of the truncated beta C-R subunit. Reversion of these mutations by the expression of the wild-type beta C-R in the two mutants resulted in a fivefold decrease in cloning efficiency of the mutants in the absence of IL3, confirming a functional interaction between the wild-type and truncated proteins. Furthermore, expression of the truncated beta C-R subunit in factor-dependent myeloid cells did not immediately render the cells autonomous but increased the spontaneous frequency to factor-independent growth by 4 orders of magnitude. Implications for both leukemogenic progression and receptor-subunit interaction and signaling are discussed.

  6. Analysis of the expression and polymorphism of APOE, HSP, BDNF, and GRIN2B genes associated with the neurodegeneration process in the pathogenesis of primary open angle glaucoma.

    Science.gov (United States)

    Nowak, Alicja; Majsterek, Ireneusz; Przybyłowska-Sygut, Karolina; Pytel, Dariusz; Szymanek, Katarzyna; Szaflik, Jerzy; Szaflik, Jacek P

    2015-01-01

    Glaucoma is characterized by optic neuropathy of the RGC or retinal nerve fiber. The aim of this study was to evaluate a relationship between the neurodegenerative genes' polymorphisms of the APOE (rs449647), BDNF (rs2030324), GRIN2B (rs3764028), and HSP70-1 (rs1043618) and the occurrence risk of POAG and to investigate its effect on allele-specific gene expression. Genomic DNA was extracted from peripheral blood. Analysis of the genes' polymorphisms was performed using PCR-RFLP. The level of mRNA expression was determined by QRT-PCR. We showed a statistically significant association of BDNF and APOE genes' polymorphisms with a risk of POAG occurrence. There was a statistically significant association of the rs2030324 polymorphism with progression of POAG based on cup disc ratio value and rs1043618 polymorphism based on nerve fiber index and rim area. Furthermore, we found that mean HSP70-1 mRNA expression was significantly lower in the case of individuals with the G/G genotype than in the case of minor allele carriers, that is, G/C and C/C. We also found that BDNF and HSP70-1 expression level are associated with the progression of POAG based on rim area value. In conclusion, our results suggest that BDNF, APOE, and HSP70-1 genes might be associated with a risk of POAG occurrence in the Polish population.

  7. Structure and expression analysis of genes encoding ADP-glucose pyrophosphorylase large subunit in wheat and its relatives.

    Science.gov (United States)

    Zhang, Xiao-Wei; Li, Si-Yu; Zhang, Ling-Ling; Yang, Qiang; Jiang, Qian-Tao; Ma, Jian; Qi, Peng-Fei; Li, Wei; Chen, Guo-Yue; Lan, Xiu-Jin; Deng, Mei; Lu, Zhen-Xiang; Liu, Chunji; Wei, Yu-Ming; Zheng, You-Liang

    2016-07-01

    ADP-glucose pyrophosphorylase (AGP), which consists of two large subunits (AGP-L) and two small subunits (AGP-S), controls the rate-limiting step in the starch biosynthetic pathway. In this study, a full-length open reading frame (ORF) of AGP-L gene (named as Agp2) in wheat and a series of Agp2 gene sequences in wheat relatives were isolated. The coding region of Agp2 contained 15 exons and 14 introns including a full-length ORF of 1566 nucleotides, and the deduced protein contained 522 amino acids (57.8 kDa). Generally, the phylogenetic tree of Agp2 indicated that sequences from A- and D-genome donor species were most similar to each other and sequences from B-genome donor species contained more variation. Starch accumulation and Agp2 expression in wheat grains reached their peak at 21 and 15 days post anthesis (DPA), respectively.

  8. Evolution, expression differentiation and interaction specificity of heterotrimeric G-protein subunit gene family in the mesohexaploid Brassica rapa.

    Directory of Open Access Journals (Sweden)

    Gulab C Arya

    Full Text Available Heterotrimeric G-proteins, comprising of Gα, Gβ, and Gγ subunits, are important signal transducers which regulate many aspects of fundamental growth and developmental processes in all eukaryotes. Initial studies in model plants Arabidopsis and rice suggest that the repertoire of plant G-protein is much simpler than that observed in metazoans. In order to assess the consequence of whole genome triplication events within Brassicaceae family, we investigated the multiplicity of G-protein subunit genes in mesohexaploid Brassica rapa, a globally important vegetable and oilseed crop. We identified one Gα (BraA.Gα1, three Gβ (BraA.Gβ1, BraA.Gβ2, and BraA.Gβ3, and five Gγ (BraA.Gγ1, BraA.Gγ2, BraA.Gγ3, BraA.Gγ4, and BraA.Gγ5 genes from B. rapa, with a possibility of 15 Gαβγ heterotrimer combinations. Our analysis suggested that the process of genome triplication coupled with gene-loss (gene-fractionation phenomenon have shaped the quantitative and sequence diversity of G-protein subunit genes in the extant B. rapa genome. Detailed expression analysis using qRT-PCR assays revealed that the G-protein genes have retained ubiquitous but distinct expression profiles across plant development. The expression of multiple G-protein genes was differentially regulated during seed-maturation and germination stages, and in response to various phytohormone treatments and stress conditions. Yeast-based interaction analysis showed that G-protein subunits interacted in most of the possible combinations, with some degree of subunit-specific interaction specificity, to control the functional selectivity of G-protein heterotrimer in different cell and tissue-types or in response to different environmental conditions. Taken together, this research identifies a highly diverse G-protein signaling network known to date from B. rapa, and provides a clue about the possible complexity of G-protein signaling networks present across globally important Brassica

  9. Cloning and expression in Escherichia coli of a new gene of Schistosoma japonicum encoding casein kinase Ⅱ beta subunit

    Institute of Scientific and Technical Information of China (English)

    彭寨玉; 余新炳; 吴忠道; 徐劲; 吴德; 李孜

    2004-01-01

    Background Nowadays it is now a focus topic in schistosomiasis research to find ideal vaccine candidates and new drug targets for developing anti-schistosomiasis vaccine. We cloned a new gene, casein kinase Ⅱ beta subunit, of Schistosoma japonicum (S. japonicum) and express it in Escherichia coli (E.coli).Methods The ESTs obtained in our laboratory were analyzed by homologous searching, and a new gene was recognized. The full-length cDNA of the new gene was obtained by joining the 3'RACE PCR fragment and the EST clone. To express the new gene, the cDNA was cloned into pGEX-4T-1 vector and then transformed into E.coli JM109. The recombinant protein was analyzed by SDS-PAGE and Western-blot. Results A 908 bp cDNA was isolated from S. japonicum and identified to be casein kinase Ⅱ beta subunit gene by sequence analysis. The open reading frame of the gene encodes a protein of 217 amino acids exhibiting 75.8%, 75.8%, 73.9%, 68.2%, 51.6% identity to the amino acids sequence of the corresponding genes of Homo sapiens (H. sapiens), Xenopus laevi (X. laevi), Drosophila melanogaster (D. melanogaster), Caenorhabditis elegan (C. elegan), and Schizosaccharomyces pombe (S. promber) respectively. The predicted molecular weight of the protein was 24.921 kDa. The new cDNA sequence had been submitted to GenBank, and its accession number is AY241391. This cDNA was subcloned into the pGEX-4T-1 vector and expressed in E.coli JM109.The recombinant protein could be recognized by the S. japonicum infected rabbit serum. Conclusion The full-length cDNA sequences encoding S. japonicum casein kinase Ⅱ beta subunit were firstly sequenced, cloned, and expressed in E.coli.

  10. Rational design of glycerol dehydratase: Swapping the genes encoding the subunits of glycerol dehydratase to improve enzymatic properties

    Institute of Scientific and Technical Information of China (English)

    QI Xianghui; SUN Liang; LUO Zhaofei; WU Jiequn; MENG Xiaolei; TANG Yue; WEI Yutuo; HUANG Ribo

    2006-01-01

    1,3-propanediol (1,3-PD) is an important material for chemical industry, and there has been always much interest in the production of 1,3-PD using all possible routes. The genes encoding glycerol dehydratase (GDHt) from Citrobacter freundii,Klebsiella pneumoniae and metagenome were cloned and expressed in E. coli. All glycerol dehydratases but the one from metagenome could be detected to show enzyme activities. In order to improve the enzymatic properties of GDHts, the genes encoding α and β-γ subunits were cloned, and the enzyme characteristics were evolved by rational design based on their 3D structures which were constructed by homology modeling. Six heteroenzymes were obtained by swapping the α subunit genes of these three different-source-derived GDHts. The pH,thermal stability and Vmax of some heteroenzymes were dramatically improved by 2-5 times compared with the wild one (GDHtKP). The GDHt cloned from metagenome, originally proved to be with no enzyme activity, was converted into active enzyme by swapping its subunits with other different GDHts. In addition, the effect of subtle 3D structural changes on the properties of the enzyme was also observed.

  11. The catalytic subunit of cAMP-dependent protein kinase induces expression of genes containing cAMP-responsive enhancer elements.

    Science.gov (United States)

    Riabowol, K T; Fink, J S; Gilman, M Z; Walsh, D A; Goodman, R H; Feramisco, J R

    1988-11-03

    Transcriptional regulation of eukaryotic genes by cyclic AMP requires a cAMP-dependent protein kinase (A kinase). Two hypotheses have been proposed to explain how the holoenzyme of the A kinase induces transcription. The regulatory subunits of the A kinase, which bind cAMP and DNA, and have amino-acid homology with the Escherichia coli catabolite activator protein could directly stimulate gene expression. Alternatively, phosphorylation by the catalytic subunits could induce transcription by activating proteins involved in gene transcription. To distinguish between these models, we microinjected purified preparations of the catalytic and regulatory subunits of A kinase into tissue culture cells and monitored expression of a stably integrated fusion gene containing a cAMP-responsive human promoter fused to a bacterial reporter gene, or of the endogenous c-fos gene. The catalytic subunit stimulated expression of these genes, whereas the regulatory subunit did not. These results indicate that the catalytic subunit of A kinase is sufficient to induce expression of two cAMP-responsive genes, without increasing levels of cAMP.

  12. Transgene inheritance and quality improvement by expressing novel HMW glutenin subunit (HMW-GS) genes in winter wheat

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    The expression vector pBPC30, which carries the high molecular weight glutenin subunit (HMW-GS) 1Dx5 and 1Dy10 genes, was transferred into hexaploid winter wheat cv. Jinghua No. 1, Jing411 and Jingdong No. 6 explants of immature embryos and immature inflorescence by particle bombardment. A large number of resistant transgenic plants were obtained under the selection of herbicide bialaphos or phosphinothricin (PPT). Confirmed transgenic plants of T0 generation showed successful integration of HMW-GS genes and bar gene into the wheat genome. T1 generation of transgenic plants can resist 20-150 mg/L PPT. Protein analysis of T2 seed by SDS-PAGE showed that HMW-GS 1Dx5 and 1Dy10 genes were well expressed in offspring seed of transgenic lines by co-expression with or substitution of endogenous 1Dx2 or 1Dy10. In one transgenic line, TG3-74, a new protein band between endogenous protein subunits 7 and 8 (marked as 8*) of glutenin appeared, but endogenous subunit 8 (encoded by 1By8 gene) was absent. Analysis of gluten rheological quality on seed proteins of 102 T3 plants showed that the sedimentation value of 5 transgenic lines (44.2-49.0 mL) was remarkably improved, 59.6%-64.3% higher than that of wild type Jinghua No. 1 and Jingdong No. 6, similar to bread wheat Cheyenne (48.0 mL). Analysis of dough rheological properties of transgenic lines showed that the dough stable time of 5 transgenic lines range from 16 to 30 min, whereas the dough stable time of wild type was only between 3-7 min. Our research suggests that introducing novel HMW-GS genes into wheat is an efficient way to improve its bread-making quality.

  13. Isolation and Sequence Analysis of HMW Glutenin Subunit 1Dy10.1 Ecoding Gene from Xinjiang Wheat (Triticum petropavlovskyi Udacz.et Migusch)

    Institute of Scientific and Technical Information of China (English)

    JIANG Qian-tao; WEI Yu-ming; WANG Ji-rui; YAN Ze-hong; ZHENG You-liang

    2006-01-01

    A novel HMW glutenin subunit gene 1Dy10.1 was isolated and characterized from Xinjiang wheat (Triticum petropavlovskyi. Udacz. et Migusch) accession Daomai 2. The complete open reading frame (ORF) of 1Dy10.1 was 1965 bp, encoding 655 amino acids. The numbers and distribution of cysteines in 1Dy10.1 were similar to those of 1Dy10 and other y-type subunits. In the N-terminal of 1Dy10.1, an amino acid was changed from L (leucine) to P (proline) at position 55. The repetitive domain of 1Dy10.1 differed from those of known HMW subunits by substitutions, insertions or/and deletions involving single or more amino acid residues. In the repetitive domain of subunit 1Dy10.1, the deletion of tripeptide GQQ in the consensus unit PGQGQQ resulted in the appearance of the motif PGQ that have not been observed in other known y-type HMW subunits. In comparison with the subunit 1Dy12, a deletion of dipeptide GQ, which occurred in subunit 1Dy10, was also observed in subunit 1Dy10.1. The cloned 1Dyl0.1 gene had been successfully expressed in Escherichia coli, and the expressed protein had the identical mobility with the endogenous subunit 1Dyl0.1 from seed.

  14. Ribosomal protein S3: a KH domain subunit in NF-kappaB complexes that mediates selective gene regulation.

    Science.gov (United States)

    Wan, Fengyi; Anderson, D Eric; Barnitz, Robert A; Snow, Andrew; Bidere, Nicolas; Zheng, Lixin; Hegde, Vijay; Lam, Lloyd T; Staudt, Louis M; Levens, David; Deutsch, Walter A; Lenardo, Michael J

    2007-11-30

    NF-kappaB is a DNA-binding protein complex that transduces a variety of activating signals from the cytoplasm to specific sets of target genes. To understand the preferential recruitment of NF-kappaB to specific gene regulatory sites, we used NF-kappaB p65 in a tandem affinity purification and mass spectrometry proteomic screen. We identified ribosomal protein S3 (RPS3), a KH domain protein, as a non-Rel subunit of p65 homodimer and p65-p50 heterodimer DNA-binding complexes that synergistically enhances DNA binding. RPS3 knockdown impaired NF-kappaB-mediated transcription of selected p65 target genes but not nuclear shuttling or global protein translation. Rather, lymphocyte-activating stimuli caused nuclear translocation of RPS3, parallel to p65, to form part of NF-kappaB bound to specific regulatory sites in chromatin. Thus, RPS3 is an essential but previously unknown subunit of NF-kappaB involved in the regulation of key genes in rapid cellular activation responses. Our observations provide insight into how NF-kappaB selectively controls gene expression.

  15. Switch in glutamate receptor subunit gene expression in CA1 subfield of hippocampus following global ischemia in rats.

    Science.gov (United States)

    Pellegrini-Giampietro, D E; Zukin, R S; Bennett, M V; Cho, S; Pulsinelli, W A

    1992-11-01

    Severe, transient global ischemia of the brain induces delayed damage to specific neuronal populations. Sustained Ca2+ influx through glutamate receptor channels is thought to play a critical role in postischemic cell death. Although most kainate-type glutamate receptors are Ca(2+)-impermeable, Ca(2+)-permeable kainate receptors have been reported in specific kinds of neurons and glia. Recombinant receptors assembled from GluR1 and/or GluR3 subunits in exogenous expression systems are permeable to Ca2+; heteromeric channels containing GluR2 subunits are Ca(2+)-impermeable. Thus, altered expression of GluR2 in development or following a neurological insult or injury to the brain can act as a switch to modify Ca2+ permeability. To investigate the molecular mechanism underlying delayed postischemic cell death, GluR1, GluR2, and GluR3 gene expression was examined by in situ hybridization in postischemic rats. Following severe, transient forebrain ischemia GluR2 gene expression was preferentially reduced in CA1 hippocampal neurons at a time point that preceded their degeneration. The switch in expression of kainate/AMPA receptor subunits coincided with the previously reported increase in Ca2+ influx into CA1 cells. Timing of the switch indicates that it may play a causal role in postischemic cell death.

  16. B2B marketing

    OpenAIRE

    Pospíšilová, Lucie

    2010-01-01

    The main goal of this bachelor thesis is to apply theoretical knowledge in B2B marketing to the example of marketing processes in a particular company, to evaluate the current situation of its activities with regard to B2B principles and to suggest relevant recommendations. The theoretical part focuses on specific characteristics of B2B marketing, describes its differences from marketing on consumer markets, deals with buying behaviour of organizations and specifies particular features of mar...

  17. Domain mapping of the retinal cyclic GMP phosphodiesterase gamma-subunit. Function of the domains encoded by the three exons of the gamma-subunit gene.

    Science.gov (United States)

    Takemoto, D J; Hurt, D; Oppert, B; Cunnick, J

    1992-02-01

    Retinal rod-outer-segment phosphodiesterase (PDE) is a heterotetramer consisting of two similar, but not identical, catalytic subunits (alpha and beta) and two identical inhibitory subunits (gamma 2). Previously, we have reported that the site of PDE alpha/beta interaction with PDE gamma is located within residues 54-87 [Cunnick, Hurt, Oppert, Sakamoto & Takemoto (1990) Biochem. J. 271, 721-727]. The site for PDE gamma interaction with transducin alpha (T alpha) was found to encompass residues 24-45 of PDE gamma [Morrison, Cunnick, Oppert & Takemoto (1989) J. Biol. Chem. 264, 11671-11681]. In order to identify binding sites and other functional domains of PDE gamma, the three peptides which are encoded by the three exons of the PDE gamma gene were synthesized chemically. These exons encode for residues 1-49, 50-62 and 63-87 of bovine PDE gamma [Piriev, Purishko, Khramtsov & Lipkin (1990) Dokl. Akad. Nauk. SSSR 315, 229-230]. The peptide encompassing residues 63-87 was inhibitory in a PDE assay, whereas peptides 1-49 and 50-62 had no effect. However, both peptides 1-49 and 63-87 bound to PDE alpha/beta in a solid-phase binding assay. Only peptide 1-49 bound to T alpha.GTP[S] (GTP[S] is guanosine 5'-[gamma-thio]triphosphate). These data confirm that the inhibitory region of PDE gamma is encoded by exon 3 (residues 63-87), whereas a separate binding site for PDE alpha/beta and for T alpha.GTP[S] is encoded by exon 1 (residues 1-49). To study further the structure-function relationship of PDE gamma, this entire protein and two mutants were chemically synthesized. One mutant (-CT) lacked residues 78-87, whereas another replaced tyrosine-84 with glycine (TYR-84). Whereas the synthetic PDE gamma inhibited PDE alpha/beta catalytic activity, the -CT and TVR-84 mutants did not. All three synthetic proteins bound to both PDE alpha/beta and and T alpha.GTP[S]. These data confirm the presence of an alternative binding site on PDE gamma and demonstrate the importance of tyrosine

  18. Nuclear factor YY1 activates the mammalian F0F1 ATP synthase alpha-subunit gene.

    Science.gov (United States)

    Breen, G A; Vander Zee, C A; Jordan, E M

    1996-01-01

    Analysis of the promoters of the bovine and human nuclear-encoded mitochondrial F0F1 ATP synthase alpha-subunit genes (ATPA) has identified several positive cis-acting regulatory regions that are important for basal promoter activity in human HeLa cells. We have previously determined that the binding of a protein factor, termed ATPF1, to an E-box sequence (CANNTG) located within one of these cis-acting regions is critical for transcriptional activation of the ATPA gene. In this article, we describe a second positive cis-acting regulatory element of the ATPA gene that is important for expression of the ATPA gene. We show that this cis-acting element also contains a binding site for a protein present in HeLa cells. On the basis of electrophoretic mobility shift patterns, oligonucleotide competition assays, and immunological cross-reactivity, we conclude that this protein factor is Yin-Yang 1 (YY1). Experiments carried out to examine the functional role of YY1 within the context of the ATPA promoter demonstrated that YY1 acts as a positive regulator of the ATPA gene. For example, when the YY1 binding site of the ATPA promoter was placed upstream of a reporter gene it was found to activate transcription in transient transfection assays. In addition, disruption of the YY1 binding site in the ATPA gene resulted in a loss of transcriptional activity. Furthermore, in cotransfection experiments overexpression of YY1 in trans was found to activate transcription of ATPA promoter-CAT constructs. Thus, at least two positive trans-acting regulatory factors, ATPF1 and YY1, are important for expression of the bovine and human F0F1 ATP synthase alpha-subunit genes.

  19. Analysis of the Expression and Polymorphism of APOE, HSP, BDNF, and GRIN2B Genes Associated with the Neurodegeneration Process in the Pathogenesis of Primary Open Angle Glaucoma

    Directory of Open Access Journals (Sweden)

    Alicja Nowak

    2015-01-01

    Full Text Available Glaucoma is characterized by optic neuropathy of the RGC or retinal nerve fiber. The aim of this study was to evaluate a relationship between the neurodegenerative genes’ polymorphisms of the APOE (rs449647, BDNF (rs2030324, GRIN2B (rs3764028, and HSP70-1 (rs1043618 and the occurrence risk of POAG and to investigate its effect on allele-specific gene expression. Genomic DNA was extracted from peripheral blood. Analysis of the genes’ polymorphisms was performed using PCR-RFLP. The level of mRNA expression was determined by QRT-PCR. We showed a statistically significant association of BDNF and APOE genes’ polymorphisms with a risk of POAG occurrence. There was a statistically significant association of the rs2030324 polymorphism with progression of POAG based on cup disc ratio value and rs1043618 polymorphism based on nerve fiber index and rim area. Furthermore, we found that mean HSP70-1 mRNA expression was significantly lower in the case of individuals with the G/G genotype than in the case of minor allele carriers, that is, G/C and C/C. We also found that BDNF and HSP70-1 expression level are associated with the progression of POAG based on rim area value. In conclusion, our results suggest that BDNF, APOE, and HSP70-1 genes might be associated with a risk of POAG occurrence in the Polish population.

  20. Enhancement of lipid productivity in oleaginous Colletotrichum fungus through genetic transformation using the yeast CtDGAT2b gene under model-optimized growth condition.

    Science.gov (United States)

    Dey, Prabuddha; Mall, Nikunj; Chattopadhyay, Atrayee; Chakraborty, Monami; Maiti, Mrinal K

    2014-01-01

    Oleaginous fungi are of special interest among microorganisms for the production of lipid feedstocks as they can be cultured on a variety of substrates, particularly waste lingocellulosic materials, and few fungal strains are reported to accumulate inherently higher neutral lipid than bacteria or microalgae. Previously, we have characterized an endophytic filamentous fungus Colletotrichum sp. DM06 that can produce total lipid ranging from 34% to 49% of its dry cell weight (DCW) upon growing with various carbon sources and nutrient-stress conditions. In the present study, we report on the genetic transformation of this fungal strain with the CtDGAT2b gene, which encodes for a catalytically efficient isozyme of type-2 diacylglycerol acyltransferase (DGAT) from oleaginous yeast Candida troplicalis SY005. Besides the increase in size of lipid bodies, total lipid titer by the transformed Colletotrichum (lipid content ∼73% DCW) was found to be ∼1.7-fold more than the wild type (lipid content ∼38% DCW) due to functional activity of the CtDGAT2b transgene when grown under standard condition of growth without imposition of any nutrient-stress. Analysis of lipid fractionation revealed that the neutral lipid titer in transformants increased up to 1.8-, 1.6- and 1.5-fold compared to the wild type when grown under standard, nitrogen stress and phosphorus stress conditions, respectively. Lipid titer of transformed cells was further increased to 1.7-fold following model-based optimization of culture conditions. Taken together, ∼2.9-fold higher lipid titer was achieved in Colletotrichum fungus due to overexpression of a rate-limiting crucial enzyme of lipid biosynthesis coupled with prediction-based bioprocess optimization.

  1. Enhancement of lipid productivity in oleaginous Colletotrichum fungus through genetic transformation using the yeast CtDGAT2b gene under model-optimized growth condition.

    Directory of Open Access Journals (Sweden)

    Prabuddha Dey

    Full Text Available Oleaginous fungi are of special interest among microorganisms for the production of lipid feedstocks as they can be cultured on a variety of substrates, particularly waste lingocellulosic materials, and few fungal strains are reported to accumulate inherently higher neutral lipid than bacteria or microalgae. Previously, we have characterized an endophytic filamentous fungus Colletotrichum sp. DM06 that can produce total lipid ranging from 34% to 49% of its dry cell weight (DCW upon growing with various carbon sources and nutrient-stress conditions. In the present study, we report on the genetic transformation of this fungal strain with the CtDGAT2b gene, which encodes for a catalytically efficient isozyme of type-2 diacylglycerol acyltransferase (DGAT from oleaginous yeast Candida troplicalis SY005. Besides the increase in size of lipid bodies, total lipid titer by the transformed Colletotrichum (lipid content ∼73% DCW was found to be ∼1.7-fold more than the wild type (lipid content ∼38% DCW due to functional activity of the CtDGAT2b transgene when grown under standard condition of growth without imposition of any nutrient-stress. Analysis of lipid fractionation revealed that the neutral lipid titer in transformants increased up to 1.8-, 1.6- and 1.5-fold compared to the wild type when grown under standard, nitrogen stress and phosphorus stress conditions, respectively. Lipid titer of transformed cells was further increased to 1.7-fold following model-based optimization of culture conditions. Taken together, ∼2.9-fold higher lipid titer was achieved in Colletotrichum fungus due to overexpression of a rate-limiting crucial enzyme of lipid biosynthesis coupled with prediction-based bioprocess optimization.

  2. Antibody responses elicited in mice immunized with Bacillus subtilis vaccine strains expressing Stx2B subunit of enterohaemorragic Escherichia coli O157:H7 Resposta de anticorpos obtidas em camundongos imunizados com linhagens vacinais de Bacillus subtilis expressando a subunidade B da Stx2 de Escherichia coli O157:H7 enterohemorrágica

    Directory of Open Access Journals (Sweden)

    P.A.D.P. Gomes

    2009-06-01

    Full Text Available No effective vaccine or immunotherapy is presently available for patients with the hemolytic uremic syndrome (HUS induced by Shiga-like toxin (Stx producedbyenterohaemorragic Escherichia coli (EHEC strains, such as those belonging to the O157:H7 serotype. In this work we evaluated the performance of Bacillus subtilis strains, a harmless spore former gram-positive bacterium species, as a vaccine vehicle for the expression of Stx2B subunit (Stx2B. A recombinant B. subtilis vaccine strain expressing Stx2B under the control of a stress inducible promoter was delivered to BALB/c mice via oral, nasal or subcutaneous routes using both vegetative cells and spores. Mice immunized with vegetative cells by the oral route developed low but specific anti-Stx2B serum IgG and fecal IgA responses while mice immunized with recombinant spores developed anti-Stx2B responses only after administration via the parenteral route. Nonetheless, serum anti-Stx2B antibodies raised in mice immunized with the recombinant B. subtilis strain did not inhibit the toxic effects of the native toxin, both under in vitro and in vivo conditions, suggesting that either the quantity or the quality of the induced immune response did not support an effective neutralization of Stx2 produced by EHEC strains.Até o presente o momento, não há vacina ou imunoterapia disponível para pacientes com Síndrome Hemolítica Urêmica (SHU induzida pela toxina Shiga-like (Stx produzida por linhagens de Escherichia coli entero-hemorragica (EHEC, tais como as pertencentes ao sorotipo O157:H7. Neste trabalho, avaliamos a performance de Bacillus subtilis, uma espécie bacteriana gram-positiva não-patogênica formadora de esporos, como veículo vacinal para a expressão da subunidade B da Stx2B (Stx2B. Uma linhagem vacinal recombinante de B. subtilis expressando Stx2B, sob o controle de um promoter induzível por estresse, foi administrada a camundongos BALB/c por via oral, nasal ou subcutânea usando

  3. Conserved cis-regulatory modules in promoters of genes encoding wheat high-molecular-weight glutenin subunits.

    Science.gov (United States)

    Ravel, Catherine; Fiquet, Samuel; Boudet, Julie; Dardevet, Mireille; Vincent, Jonathan; Merlino, Marielle; Michard, Robin; Martre, Pierre

    2014-01-01

    The concentration and composition of the gliadin and glutenin seed storage proteins (SSPs) in wheat flour are the most important determinants of its end-use value. In cereals, the synthesis of SSPs is predominantly regulated at the transcriptional level by a complex network involving at least five cis-elements in gene promoters. The high-molecular-weight glutenin subunits (HMW-GS) are encoded by two tightly linked genes located on the long arms of group 1 chromosomes. Here, we sequenced and annotated the HMW-GS gene promoters of 22 electrophoretic wheat alleles to identify putative cis-regulatory motifs. We focused on 24 motifs known to be involved in SSP gene regulation. Most of them were identified in at least one HMW-GS gene promoter sequence. A common regulatory framework was observed in all the HMW-GS gene promoters, as they shared conserved cis-regulatory modules (CCRMs) including all the five motifs known to regulate the transcription of SSP genes. This common regulatory framework comprises a composite box made of the GATA motifs and GCN4-like Motifs (GLMs) and was shown to be functional as the GLMs are able to bind a bZIP transcriptional factor SPA (Storage Protein Activator). In addition to this regulatory framework, each HMW-GS gene promoter had additional motifs organized differently. The promoters of most highly expressed x-type HMW-GS genes contain an additional box predicted to bind R2R3-MYB transcriptional factors. However, the differences in annotation between promoter alleles could not be related to their level of expression. In summary, we identified a common modular organization of HMW-GS gene promoters but the lack of correlation between the cis-motifs of each HMW-GS gene promoter and their level of expression suggests that other cis-elements or other mechanisms regulate HMW-GS gene expression.

  4. Nucleotide sequence of a crustacean 18S ribosomal RNA gene and secondary structure of eukaryotic small subunit ribosomal RNAs.

    Science.gov (United States)

    Nelles, L; Fang, B L; Volckaert, G; Vandenberghe, A; De Wachter, R

    1984-12-11

    The primary structure of the gene for 18 S rRNA of the crustacean Artemia salina was determined. The sequence has been aligned with 13 other small ribosomal subunit RNA sequences of eukaryotic, archaebacterial, eubacterial, chloroplastic and plant mitochondrial origin. Secondary structure models for these RNAs were derived on the basis of previously proposed models and additional comparative evidence found in the alignment. Although there is a general similarity in the secondary structure models for eukaryotes and prokaryotes, the evidence seems to indicate a different topology in a central area of the structures.

  5. Repeated nebulisation of non-viral CFTR gene therapy in patients with cystic fibrosis: a randomised, double-blind, placebo-controlled, phase 2b trial

    Science.gov (United States)

    Alton, Eric W F W; Armstrong, David K; Ashby, Deborah; Bayfield, Katie J; Bilton, Diana; Bloomfield, Emily V; Boyd, A Christopher; Brand, June; Buchan, Ruaridh; Calcedo, Roberto; Carvelli, Paula; Chan, Mario; Cheng, Seng H; Collie, D David S; Cunningham, Steve; Davidson, Heather E; Davies, Gwyneth; Davies, Jane C; Davies, Lee A; Dewar, Maria H; Doherty, Ann; Donovan, Jackie; Dwyer, Natalie S; Elgmati, Hala I; Featherstone, Rosanna F; Gavino, Jemyr; Gea-Sorli, Sabrina; Geddes, Duncan M; Gibson, James S R; Gill, Deborah R; Greening, Andrew P; Griesenbach, Uta; Hansell, David M; Harman, Katharine; Higgins, Tracy E; Hodges, Samantha L; Hyde, Stephen C; Hyndman, Laura; Innes, J Alastair; Jacob, Joseph; Jones, Nancy; Keogh, Brian F; Limberis, Maria P; Lloyd-Evans, Paul; Maclean, Alan W; Manvell, Michelle C; McCormick, Dominique; McGovern, Michael; McLachlan, Gerry; Meng, Cuixiang; Montero, M Angeles; Milligan, Hazel; Moyce, Laura J; Murray, Gordon D; Nicholson, Andrew G; Osadolor, Tina; Parra-Leiton, Javier; Porteous, David J; Pringle, Ian A; Punch, Emma K; Pytel, Kamila M; Quittner, Alexandra L; Rivellini, Gina; Saunders, Clare J; Scheule, Ronald K; Sheard, Sarah; Simmonds, Nicholas J; Smith, Keith; Smith, Stephen N; Soussi, Najwa; Soussi, Samia; Spearing, Emma J; Stevenson, Barbara J; Sumner-Jones, Stephanie G; Turkkila, Minna; Ureta, Rosa P; Waller, Michael D; Wasowicz, Marguerite Y; Wilson, James M; Wolstenholme-Hogg, Paul

    2015-01-01

    Summary Background Lung delivery of plasmid DNA encoding the CFTR gene complexed with a cationic liposome is a potential treatment option for patients with cystic fibrosis. We aimed to assess the efficacy of non-viral CFTR gene therapy in patients with cystic fibrosis. Methods We did this randomised, double-blind, placebo-controlled, phase 2b trial in two cystic fibrosis centres with patients recruited from 18 sites in the UK. Patients (aged ≥12 years) with a forced expiratory volume in 1 s (FEV1) of 50–90% predicted and any combination of CFTR mutations, were randomly assigned, via a computer-based randomisation system, to receive 5 mL of either nebulised pGM169/GL67A gene–liposome complex or 0·9% saline (placebo) every 28 days (plus or minus 5 days) for 1 year. Randomisation was stratified by % predicted FEV1 (<70 vs ≥70%), age (<18 vs ≥18 years), inclusion in the mechanistic substudy, and dosing site (London or Edinburgh). Participants and investigators were masked to treatment allocation. The primary endpoint was the relative change in % predicted FEV1. The primary analysis was per protocol. This trial is registered with ClinicalTrials.gov, number NCT01621867. Findings Between June 12, 2012, and June 24, 2013, we randomly assigned 140 patients to receive placebo (n=62) or pGM169/GL67A (n=78), of whom 116 (83%) patients comprised the per-protocol population. We noted a significant, albeit modest, treatment effect in the pGM169/GL67A group versus placebo at 12 months' follow-up (3·7%, 95% CI 0·1–7·3; p=0·046). This outcome was associated with a stabilisation of lung function in the pGM169/GL67A group compared with a decline in the placebo group. We recorded no significant difference in treatment-attributable adverse events between groups. Interpretation Monthly application of the pGM169/GL67A gene therapy formulation was associated with a significant, albeit modest, benefit in FEV1 compared with placebo at 1 year, indicating a stabilisation of

  6. Cloning, expression and molecular analysis of Iranian Brucella melitensis Omp25 gene for designing a subunit vaccine

    Science.gov (United States)

    Yousefi, Soheil; Tahmoorespur, Mojtaba; Sekhavati, Mohammad Hadi

    2016-01-01

    Brucellosis is a well-known domestic animal infectious disease, which is caused by Brucella bacterium. The outer membrane protein 25 kDa (Omp25) gene plays an important role in simulating of TNF-α, IFN-α, macrophage, and cytokines cells. In the current study molecular cloning and expression analysis of Omp25 gene for designing a subunit vaccine against Brucella was investigated. Amplifying the full length of candidate gene was performed using specific primers. Sub-cloning of this gene conducted using pTZ57R/T vector in TOP10F strain of Escherichia coli(E.coli) as the host. Also, pET32(a)+ vector used for expression in BL21 (DE3) strain of E.coli. Omp25 gene with 642 bp size was amplified and cloned successfully. The expression results were confirmed by sequencing and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analyses which showed 42 kDa protein band correctly. Also, phylogenic analysis showed this gene has a near genetic relation with other Brucella strains. According to our results we can propose this gene as a candidate useful for stimulation of cell-mediated and humoral immunity system in future study. PMID:27920824

  7. hELP3 Subunit of the Elongator Complex Regulates the Transcription of HSP70 Gene in Human Cells

    Institute of Scientific and Technical Information of China (English)

    Qiuju HAN; Xiaozhe HOU; Dongmei SU; Lina PAN; Jizhou DUAN; Liguo CUI; Baiqu HUANG; Jun LU

    2007-01-01

    The human Elongator complex is remarkably similar to its yeast counterpart in several aspects.In a previous study, we analyzed the functions of the human elongation protein 3 (hELP3) subunit of the human Elongator by using an in vivo yeast complementation system. However, direct evidence for hELP3 functions in regulating gene expression in human cells was not obtained. In this study, we used hELP3 antisense oligonucleotide inhibitors to knock down hELP3 gene expression to investigate its function in human 293T cells. The results showed that specific reduction of hELP3 mRNA and protein caused a significant suppression of HSP70-2 gene expression, and this was accompanied by histone H3 hypoacetylation and decreased RNA polymerase Ⅱ density at the HSP70-2 gene. Moreover, the data also showed that hELP3 exerted the transcriptional regulatory function directly through its presence on the HSP70-2 gene. Data presented in this report provide further insight and direct evidence of the functions of hELP3 in HSP70-2 gene transcriptional elongation in human cells.

  8. Structural organization, sequence, and expression of the mouse HEXA gene encoding the alpha subunit of hexosaminidase A.

    Science.gov (United States)

    Wakamatsu, N; Benoit, G; Lamhonwah, A M; Zhang, Z X; Trasler, J M; Triggs-Raine, B L; Gravel, R A

    1994-11-01

    Genomic clones of the mouse HEXA gene encoding the alpha subunit of lysosomal beta-hexosaminidase A have been isolated, analyzed, and sequenced. The HEXA gene spans approximately 26 kb and consists of 14 exons and 13 introns. The 5' flanking region of the gene has three candidate GC boxes and a number of potential promoter and regulatory elements. Promoter analysis using deletion constructs of 5' flanking sequence fused to the bacterial chloramphenicol acetyltransferase (CAT) gene showed that 150 bp of 5' sequence was sufficient for expression in transfected monkey kidney COS cells. Determination of the sequence of the 5' end of the Hex alpha mRNA by an "anchor-ligation PCR" procedure showed that transcription is initiated from a cluster of sites centered -42, -32, and -21 bp from the first in-frame ATG. Northern blot analysis from 11 different tissues showed over five times the steady-state level of Hex alpha mRNA in testis as compared to that found in three different brain regions; the lowest level (about 1/3 of brain) was found in liver. Comparison of the 5' flanking sequence with that of the human HEXA gene revealed 78% identity within the first 100 bp. These data suggest that the mouse HEXA gene is controlled mainly by sequences located within 150 bp of the 5' flanking region, and we speculate that it may have a role, not only in brain and other tissues, but also in reproductive function in the adult male mouse.

  9. Changes in the protein expression of NR2B subunit and CaMK Ⅱ in hippocampus CA1 of rats with recurrence of CPP induced by morphine%吗啡点燃条件位置性偏爱重现大鼠海马CA1区NR2B、CaMKⅡ的变化

    Institute of Scientific and Technical Information of China (English)

    邵晓霞; 赵永娜; 李树清; 方正梅; 杨贞永

    2011-01-01

    目的:探讨吗啡(morphine,Mor)点燃条件位置性偏爱(conditioned place preference,CPP)重现大鼠海马CA1区N甲基-D-天冬氨酸受体(NMDA受体)调节亚基NR2B,钙依赖钙调蛋白激Ⅱ(Ca2+/calmodulin dependent protein kinaseⅡ,CaMKⅡ)表达的变化.方法:用恒量法(10 mg·kg-1)给大鼠连续颈背部皮下注射(subcutaneous,sc)吗啡8d建立CPP模型;用生理盐水替代吗啡训练大鼠10d,使形成的CPP逐渐消退;单次sc2.5 mg·kg-1吗啡点燃已消退的CPP.用免疫组化法检测吗啡点燃CPP重现大鼠海马CA1区NR2B、CaMKⅡ表达,图像分析系统测定阳性反应产物的平均灰度值的变化.结果:sc 10m·kg-1吗啡8d建立CPP,生理盐水训练10d使已形成的CPP消退,小剂量吗啡(2.5mg·kg-1)使消退的CPP重现;吗啡点燃CPP重现大鼠海马CA1区NA2B、CaMKⅡ表达,与对照组比较呈显著增加(P<0.05).结论:小剂量吗啡诱发大鼠CPP重现行为可能与海马CA1区NR2B、CAMK Ⅱ表达增加有关.%OBJECTIVE To study the changes of the protein expression of NR2B and CaMK Ⅱ in hippocampus CA1 area of rats with recurrence of conditioned place preference (CPP) induced by morphine. METHODS Morphine was administered via subcutaneous injection at constant dose (10 mg·kg-1 ) for 8 days to establish morphine CPP. The rats were administered saline instead of morphine to induce CPP extinction for 10 days. CPP was reinstated following a single priming injection of morphine (2. 5 mg·kg-1 ). The morphine induced rat CPP acquisition, extinction and reinstatement model was established. The protein levels of NR2B subunit and CaMK Ⅱ in hippocampus CA1 of rats with CPP recurrence of rats were measured by streptavidin-perosidase immunohistochemical method and the mean gray values of immunoreactive product was detected by image analysis system. RESULTS Injecting morphine (10 mg·kg-1) for 8 days succeeded to induce CPP. After 10 days training with saline, CPP failed to be induced. A single

  10. Serotonin Transporter (5-HTT) and gamma-Aminobutyric Acid Receptor Subunit beta3 (GABRB3) Gene Polymorphisms are not Associated with Autism in the IMGSA Families

    DEFF Research Database (Denmark)

    Maestrini, E.; Lai, C.; Marlow, A.;

    1999-01-01

    Previous studies have suggested that the serotonin transporter (5-HTT) gene and the gamma-aminobutyric acid receptor subunit beta3 (GABRB3) gene, or other genes in the 15q11-q13 region, are possibly involved in susceptibility to autism. To test this hypothesis we performed an association study...... and the GABRB3 genes are unlikely to play a major role in the aetiology of autism in our family data set....

  11. CLONING, SEQUENCING AND EXPRESSION STUDIES OF THE GENES ENCODING AMICYANIN AND THE BETA-SUBUNIT OF METHYLAMINE DEHYDROGENASE FROM THIOBACILLUS-VERSUTUS

    NARCIS (Netherlands)

    UBBINK, M; VANKLEEF, MAG; KLEINJAN, DJ; HOITINK, CWG; HUITEMA, F; BEINTEMA, JJ; DUINE, JA; CANTERS, GW

    1991-01-01

    The genes encoding amicyanin and the beta-subunit of methylamine dehydrogenase (MADH) from Thiobacillus versutus have been cloned and sequenced. The organization of these genes makes it likely that they are coordinately expressed and it supports earlier findings that the blue copper protein amicyani

  12. Sequence analysis of β-subunit genes of the 20S proteasome in patients with relapsed multiple myeloma treated with bortezomib or dexamethasone

    NARCIS (Netherlands)

    D.I. Lichter (David); H. Danaee (Hadi); M.D. Pickard (Michael); O. Tayber (Olga); M. Sintchak (Michael); H. Shi (Hongliang); P.G. Richardson (Paul Gerard); J. Cavenagh (Jamie); J. Bladé (Joan); T. Facon (Thierry); R. Niesvizky; M. Alsina (Melissa); W. Dalton (William); P. Sonneveld (Pieter); S. Lonial (Sagar); H. van de Velde (Helgi); D. Ricci (Deborah); D.-L. Esseltine (Dixie-Lee); W.L. Trepicchio (William); G. Mulligan (George); K.C. Anderson (Kenneth Carl)

    2012-01-01

    textabstractVariations within proteasome β (PSMB) genes, which encode the β subunits of the 20S proteasome, may affect proteasome function, assembly, and/or binding of proteasome inhibitors. To investigate the potential association between PSMB gene variants and treatment-emergent resistance to bort

  13. Mouse mutants for the nicotinic acetylcholine receptor ß2 subunit display changes in cell adhesion and neurodegeneration response genes.

    Directory of Open Access Journals (Sweden)

    Carol M Rubin

    Full Text Available Mice lacking expression of the ß2 subunit of the neuronal nicotinic acetylcholine receptor (CHRNB2 display abnormal retinal waves and a dispersed projection of retinal ganglion cell (RGC axons to their dorsal lateral geniculate nuclei (dLGNs. Transcriptomes of LGN tissue from two independently generated Chrnb2-/- mutants and from wildtype mice were obtained at postnatal day 4 (P4, during the normal period of segregation of eye-specific afferents to the LGN. Microarray analysis reveals reduced expression of genes located on the cell membrane or in extracellular space, and of genes active in cell adhesion and calcium signaling. In particular, mRNA for cadherin 1 (Cdh1, a known axon growth regulator, is reduced to nearly undetectable levels in the LGN of P4 mutant mice and Lypd2 mRNA is similarly suppressed. Similar analysis of retinal tissue shows increased expression of crumbs 1 (Crb1 and chemokine (C-C motif ligand 21 (Ccl21 mRNAs in Chrnb2-/- mutant animals. Mutations in these genes are associated with retinal neuronal degeneration. The retinas of Chrnb2-/- mutants are normal in appearance, but the increased expression of these genes may also be involved in the abnormal projection patterns of RGC to the LGN. These data may provide the tools to distinguish the interplay between neural activity and molecular expression. Finally, comparison of the transcriptomes of the two different Chrnb2-/- mutant strains reveals the effects of genetic background upon gene expression.

  14. Transcritption regulation of soybean ribulose-1,5-bisphos-phate carboxylase small sub-unit gene by external factors

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Ribulose-1,5-bisphosphate carboxylase small subunit gene (rbcS) is present with multi-gene family in plant genome. In Glycine max, the rbcS polypeptide (EC4.1.1.39) is encoded by a gene family containing 4-8 members. Three full-length rbcS cDNA clones were isolated and characterized from soybean seedlings, and both of their nucleotide and amino acid sequences showed high similarity. Differential accumulation of the rbcS mRNA was observed among roots, hypocotyls, cotyledons, epicotyls and leaves. The rbcS genes were up-regulated by various external factors such as salicylic acid (SA), salt stress and drought stress. The expression level of rbcS genes after being treated by 2.0 mmol/L SA and 0.4% NaCl, respectively, is 2.5-3.0-fold as high as that of control sample. Moreover, soybean rbcS mRNA was accumulated with diurnal variation but easily influenced by light and low temperature.

  15. Intronic Polymorphisms in the CDKN2B-AS1 Gene Are Strongly Associated with the Risk of Myocardial Infarction and Coronary Artery Disease in the Saudi Population

    Directory of Open Access Journals (Sweden)

    Sayed AbdulAzeez

    2016-03-01

    Full Text Available Recent genome-wide association studies identified single nucleotide polymorphisms (SNPs on the chromosome 9p21.3 conferring the risk for CAD (coronary artery disease in individuals of Caucasian ancestry. We performed a genetic association study to investigate the effect of 12 candidate SNPs within 9p21.3 locus on the risk of CAD in the Saudi population of the Eastern Province of Saudi Arabia. A total of 250 Saudi CAD patients who had experienced an myocardial infarction (MI and 252 Saudi age-matched healthy controls were genotyped using TaqMan assay. Controls with evidenced lack of CAD provided 90% of statistical power at the type I error rate of 0.05. Five percent of the results were rechecked for quality control using Sanger sequencing, the results of which concurred with the TaqMan genotyping results. Association analysis of 12 SNPs indicated a significant difference in the genotype distribution for four SNPs between cases and controls (rs564398 p = 0.0315, χ2 = 4.6, odds ratio (OD = 1.5; rs4977574 p = 0.0336, χ2 = 4.5, OD = 1.4; rs2891168 p = 1.85 × 10 − 10, χ2 = 40.6, OD = 2.1 and rs1333042 p = 5.14 × 10 − 9, χ2 = 34.1, OD = 2.2. The study identified three protective haplotypes (TAAG p = 1.00 × 10 − 4; AGTA p = 0.022 and GGGCC p = 0.0175 and a risk haplotype (TGGA p = 2.86 × 10 − 10 for the development of CAD. This study is in line with others that indicated that the SNPs located in the intronic region of the CDKN2B-AS1 gene are associated with CAD.

  16. Fit 2-B FATHERS.

    Science.gov (United States)

    Maiorano, Joseph J.

    2001-01-01

    Fit 2-B FATHERS is a parenting-skills education program for incarcerated adult males. The goals of this program are for participants to have reduced recidivism rates and a reduced risk of their children acquiring criminal records. These goals are accomplished by helping participants become physically, practically, and socially fit for the demands…

  17. Z-360, a novel therapeutic agent for pancreatic cancer, prevents up-regulation of ephrin B1 gene expression and phosphorylation of NR2B via suppression of interleukin-1 β production in a cancer-induced pain model in mice

    Directory of Open Access Journals (Sweden)

    Hori Yuko

    2010-10-01

    Full Text Available Abstract Background Z-360 is an orally active cholecystokinin-2 (CCK2/gastrin receptor antagonist currently under development as a therapeutic drug for pancreatic cancer. It was previously reported that Z-360 treatment in combination with gemcitabine prolonged the survival period in a lethal pancreatic cancer xenograft model in mice. In a phase Ib/IIa clinical study, Z-360 treatment displayed a trend of reduced pain in patients with advanced pancreatic cancer in combination with gemcitabine including analgesics such as opioids. Here, we investigated the mechanism of analgesic action of Z-360 in a severe cancer-induced pain model in mice, which is considered to be opioid-resistant, by examining ephrin B1 gene expression, N-methyl-D-aspartate receptor NR2B subunit phosphorylation, and interleukin-1β (IL-1β production. Results In a mouse model of cancer-induced pain, ephrin B1 gene expression in dorsal root ganglia (DRGs and the phosphorylation of NR2B in the spinal cord were induced. Z-360 treatment inhibited both ephrin B1 gene expression and the phosphorylation of NR2B. In addition, IL-1β production increased in the cancer-inoculated hind paw of mice, but could be suppressed by treatment with Z-360. Moreover, we observed that the CCK1 receptor antagonist devazepide similarly suppressed up-regulation of ephrin B1 gene expression and IL-1β production, and that the intraperitoneal injection of sulfated CCK-8 induced the production of IL-1β in the cancer-inoculated region. Conclusions We have identified a novel pain cascade, in which IL-1β production in cancer-inoculated regions induces ephrin B1 gene expression in DRGs and then ephrin B1 enhances the tyrosine phosphorylation of NR2B via Eph B receptor in the spinal cord. Notably, Z-360 relieves cancer-induced pain by preventing this pain cascade through the suppression of IL-1β production, likely via the blockade of CCK1 receptor. The pre-clinical results presented here support the analgesic

  18. Targeted deletion of the mouse α2 nicotinic acetylcholine receptor subunit gene (Chrna2) potentiates nicotine-modulated behaviors.

    Science.gov (United States)

    Lotfipour, Shahrdad; Byun, Janet S; Leach, Prescott; Fowler, Christie D; Murphy, Niall P; Kenny, Paul J; Gould, Thomas J; Boulter, Jim

    2013-05-01

    Baseline and nicotine-modulated behaviors were assessed in mice harboring a null mutant allele of the nicotinic acetylcholine receptor (nAChR) subunit gene α2 (Chrna2). Homozygous Chrna2(-/-) mice are viable, show expected sex and Mendelian genotype ratios, and exhibit no gross neuroanatomical abnormalities. A broad range of behavioral tests designed to assess genotype-dependent effects on anxiety (elevated plus maze and light/dark box), motor coordination (narrow bean traverse and gait), and locomotor activity revealed no significant differences between mutant mice and age-matched wild-type littermates. Furthermore, a panel of tests measuring traits, such as body position, spontaneous activity, respiration, tremors, body tone, and startle response, revealed normal responses for Chrna2-null mutant mice. However, Chrna2(-/-) mice do exhibit a mild motor or coordination phenotype (a decreased latency to fall during the accelerating rotarod test) and possess an increased sensitivity to nicotine-induced analgesia in the hotplate assay. Relative to wild-type, Chrna2(-/-) mice show potentiated nicotine self-administration and withdrawal behaviors and exhibit a sex-dependent enhancement of nicotine-facilitated cued, but not trace or contextual, fear conditioning. Overall, our results suggest that loss of the mouse nAChR α2 subunit has very limited effects on baseline behavior but does lead to the potentiation of several nicotine-modulated behaviors.

  19. Preparation of Polyclonal Antibodies of Rubisco Large and Small Subunits and Their Application in the Functional Analysis of the Genes

    Institute of Scientific and Technical Information of China (English)

    Peng-Da MA; Tian-Cheng LU; Xiao-Fu ZHOU; Xiao-Juan ZHU; Xing-Zhi WANG

    2004-01-01

    Spinach Rubisco (ribulose-l,5-bisphosphate carboxylase/oxygenase) large (rbcL) and small (rbcS) subunits were separated by SDS-PAGE, and protein amount and purity were determined by Bradford assay. Polyclonal antibodies against rbcL and rbcS subunit were generated in female BALB/c mice and had no cross-reaction with each other. A total of 81 μg antigens were used and 0.3 ml anti-sera with titer of 1:5000were yielded. The antibodies were also applicable to study rbcL and rbcS in tobacco plant Nicotiana benthamiana. Potato virus X vector pGR107 induced silencing of rbcS gene by Agrobacterium in Nicotiana benthaniana was performed. The expression level ofrbcL and rbcS was lower in rbcS silenced plants than that in control plants as detected by the corresponding antibodies. This implied that the expression of rbcL was regulated by rbcS.

  20. Analysis of IL-12 p40 subunit gene and IFN-γ G5644A polymorphisms in Idiopathic Pulmonary Fibrosis

    Directory of Open Access Journals (Sweden)

    Welsh Kenneth I

    2003-06-01

    Full Text Available Abstract Background Genes encoding cytokine mediators are prime candidates for genetic analysis in conditions with T-helper (Th cell disease driven imbalance. Idiopathic Pulmonary Fibrosis (IPF is a predominantly Th2 mediated disease associated with a paucity of interferon-gamma (IFN-γ. The paucity of IFN-γ may favor the development of progressive fibrosis in IPF. Interleukin-12 (IL-12 plays a key role in inducing IFN-γ production. The aim of the current study was to assess whether the 1188 (A/C 3'UTR single nucleotide polymorphism (SNP in the IL-12 p40 subunit gene which was recently found to be functional and the 5644 (G/A 3' UTR SNP of the IFN-γ gene were associated with susceptibility to IPF. Methods We investigated the allelic distribution in these loci in UK white Caucasoid subjects comprising 73 patients with IPF and 157 healthy controls. The SNPs were determined using the polymerase chain reaction in association with sequence-specific primers incorporating mismatches at the 3'-end. Results Our results showed that these polymorphisms were distributed similarly in the IPF and control groups Conclusion We conclude that these two potentially important candidate gene single nucleotide polymorphisms are not associated with susceptibility to IPF.

  1. Phylogenetic relationships between Vorticella convallaria and other species inferred from small subunit rRNA gene sequences.

    Science.gov (United States)

    Itabashi, Takeshi; Mikami, Kazuyuki; Fang, Jie; Asai, Hiroshi

    2002-08-01

    Vorticellid ciliates generally dwell in freshwater. In nature, the species have up until now been identified by comparison with previous descriptions. It is difficult to identify between species of the genus Vorticella, because the morphological markers of vorticellid ciliates described in reports are limited and variable. Unfortunately, culturing them has only succeeded with certain species such as Vorticella convallaria, but many others have been impossible to culture. To find out whether the sequence of a small subunit rRNA gene was an appropriate marker to identify vorticellid ciliates, the gene was aligned and compared. Finding a new convenient method will contribute to research on vorticellid ciliates. In strains of V. convallaria, classified morphologically, some varieties of the SSrRNA gene sequences were recognized, but there were large variations within the same species. According to the phylogenetic tree, these strains are closely related. However, the difference was not as big as between Vorticella and Carchesium. In addition, Carchesium constructed a distinct clade from the genus Vorticella and Epistylis. These results show the possibility that the SSrRNA gene is one of the important markers to identify species of Vorticella. This study is first to approach and clarify the complicated taxa in the genus Vorticella.

  2. Molecular evolution and nucleotide sequences of the maize plastid genes for the alpha subunit of CF1 (atpA) and the proteolipid subunit of CF0 (atpH).

    Science.gov (United States)

    Rodermel, S R; Bogorad, L

    1987-05-01

    The nucleotide sequences of the maize plastid genes for the alpha subunit of CF1 (atpA) and the proteolipid subunit of CF0 (atpH) are presented. The evolution of these genes among higher plants is characterized by a transition mutation bias of about 2:1 and by rates of synonymous and nonsynonymous substitution which are much lower than similar rates for genes from other sources. This is consistent with the notion that the plastid genome is evolving conservatively in primary sequence. Yet, the mode and tempo of sequence evolution of these and other plastid-encoded coupling factor genes are not the same. In particular, higher rates of nonsynonymous substitution in atpE (the gene for the epsilon subunit of CF1) and higher rates of synonymous substitution in atpH in the dicot vs. monocot lineages of higher plants indicate that these sequences are likely subject to different evolutionary constraints in these two lineages. The 5'- and 3'-transcribed flanking regions of atpA and atpH from maize, wheat and tobacco are conserved in size, but contain few putative regulatory elements which are conserved either in their spatial arrangement or sequence complexity. However, these regions likely contain variable numbers of "species-specific" regulatory elements. The present studies thus suggest that the plastid genome is not a passive participant in an evolutionary process governed by a more rapidly changing, readily adaptive, nuclear compartment, but that novel strategies for the coordinate expression of genes in the plastid genome may arise through rapid evolution of the flanking sequences of these genes.

  3. The C. elegans nuclear receptor gene fax-1 and homeobox gene unc-42 coordinate interneuron identity by regulating the expression of glutamate receptor subunits and other neuron-specific genes.

    Science.gov (United States)

    Wightman, Bruce; Ebert, Bryan; Carmean, Nicole; Weber, Katherine; Clever, Sheila

    2005-11-01

    The fax-1 gene of the nematode C. elegans encodes a conserved nuclear receptor that is the ortholog of the human PNR gene and functions in the specification of neuron identities. Mutations in fax-1 result in locomotion defects. FAX-1 protein accumulates in the nuclei of 18 neurons, among them the AVA, AVB, and AVE interneuron pairs that coordinate body movements. The identities of AVA and AVE interneurons are defective in fax-1 mutants; neither neuron expresses the NMDA receptor subunits nmr-1 and nmr-2. Other ionotropic glutamate receptor subunits are expressed normally in the AVA and AVE neurons. The unc-42 homeobox gene also regulates AVA and AVE identity; however, unc-42 mutants display the complementary phenotype: NMDA receptor subunit expression is normal, but some non-NMDA glutamate receptor subunits are not expressed. These observations support a combinatorial role for fax-1 and unc-42 in specifying AVA and AVE identity. However, in four other neuron types, fax-1 is regulated by unc-42, and both transcriptional regulators function in the regulation of the opt-3 gene in the AVE neurons and the flp-1 and ncs-1 genes in the AVK neurons. Therefore, while fax-1 and unc-42 act in complementary parallel pathways in some cells, they function in overlapping or linear pathways in other cellular contexts, suggesting that combinatorial relationships among transcriptional regulators are complex and cannot be generalized from one neuron type to another.

  4. Tyrosine Phosphorylation of NR2B Contributes to Chronic Migraines via Increased Expression of CGRP in Rats

    Directory of Open Access Journals (Sweden)

    Xiping Liang

    2017-01-01

    Full Text Available Tyrosine phosphorylation of NR2B (NR2B-pTyr, a subunit of the N-methyl-D-aspartate (NMDA receptor, has been reported to develop central sensitization and persistent pain in the spine, but its effect in chronic migraines has not been examined. We hypothesized that tyrosine phosphorylation of NR2B contributes to chronic migraines (CM through calcitonin gene-related peptide (CGRP in rats. Ninety-four male Sprague-Dawley rats were subjected to seven inflammatory soup (IS injections. In a subset of animals, the time course and location of NR2B tyrosine phosphorylation were detected by western blot and immunofluorescence double staining. Another set of animals were given either genistein, vehicle, or genistein and recombinant CGRP. The mechanical threshold was measured, the expressions of NR2B-pTyr, NR2B, and CGRP were quantified using western blot, and nitric oxide (NO was measured with the nitric acid reductase method. NR2B-pTyr expression, in neurons, peaked at 24 hours after CM. Genistein improved the mechanical threshold and reduced migraine attacks 24 and 72 hours after CM. Tyrosine phosphorylation of NR2B decreased the mechanical threshold and increased migraine attacks via upregulated CGRP expression in the rat model of CM. Thus, tyrosine phosphorylation of NR2B may be a potential therapeutic target for treatment of CM.

  5. Tyrosine Phosphorylation of NR2B Contributes to Chronic Migraines via Increased Expression of CGRP in Rats

    Science.gov (United States)

    Liang, Xiping; Wang, Sha; Qin, Guangcheng; Xie, Jingmei; Tan, Ge; Zhou, Jiying; McBride, Devin W.

    2017-01-01

    Tyrosine phosphorylation of NR2B (NR2B-pTyr), a subunit of the N-methyl-D-aspartate (NMDA) receptor, has been reported to develop central sensitization and persistent pain in the spine, but its effect in chronic migraines has not been examined. We hypothesized that tyrosine phosphorylation of NR2B contributes to chronic migraines (CM) through calcitonin gene-related peptide (CGRP) in rats. Ninety-four male Sprague-Dawley rats were subjected to seven inflammatory soup (IS) injections. In a subset of animals, the time course and location of NR2B tyrosine phosphorylation were detected by western blot and immunofluorescence double staining. Another set of animals were given either genistein, vehicle, or genistein and recombinant CGRP. The mechanical threshold was measured, the expressions of NR2B-pTyr, NR2B, and CGRP were quantified using western blot, and nitric oxide (NO) was measured with the nitric acid reductase method. NR2B-pTyr expression, in neurons, peaked at 24 hours after CM. Genistein improved the mechanical threshold and reduced migraine attacks 24 and 72 hours after CM. Tyrosine phosphorylation of NR2B decreased the mechanical threshold and increased migraine attacks via upregulated CGRP expression in the rat model of CM. Thus, tyrosine phosphorylation of NR2B may be a potential therapeutic target for treatment of CM.

  6. Human natural killer cell microRNA: differential expression of MIR181A1B1 and MIR181A2B2 genes encoding identical mature microRNAs.

    Science.gov (United States)

    Presnell, S R; Al-Attar, A; Cichocki, F; Miller, J S; Lutz, C T

    2015-01-01

    Natural killer (NK) and T lymphocytes share many properties, yet only NK cells respond rapidly to infection and cancer without pre-activation. We found that few microRNAs (miRNAs) differed significantly between human NK and T cells. Among those miRNAs, miR-181a and miR-181b levels rose during NK cell differentiation. Prior studies indicate that miR-181a and miR-181b are critical for human NK cell development and are co-transcribed from genes on chromosome 1 (MIR181A1B1) and on chromosome 9 (MIR181A2B2). We mapped human MIR181A1B1 and MIR181A2B2 transcription start sites to 78.3 kb and 34.0 kb upstream of the mature miRNAs, generating predominantly unspliced transcripts of 80-127 kb and ~60 kb, respectively. Unlike mouse thymocytes, human T cells expressed both MIR181A1B1 and MIR181A2B2. We tested the hypothesis that NK cells differentially transcribe the two genes during development and in response to immune regulatory cytokines. During NK-cell differentiation, MIR181A2B2 expression rose markedly and exceeded that of MIR181A1B1. TGF-β treatment increased NK-cell MIR181A2B2 transcription, whereas IL-2, IL-15 and IL-12/IL-18 treatments upregulated MIR181A1B1. The MIR181A2B2 promoter was strongly transactivated by SMAD3 and SMAD4 transcription factors, suggesting that TGF-β signaling upregulates MIR181A2B2 expression, at least in part, through SMAD-dependent promoter activation.

  7. Mitochondrial bioenergetics and redox state are unaltered in Trypanosoma cruzi isolates with compromised mitochondrial complex I subunit genes.

    Science.gov (United States)

    Carranza, Julio César; Kowaltowski, Alicia J; Mendonça, Marco Aurélio G; de Oliveira, Thays C; Gadelha, Fernanda R; Zingales, Bianca

    2009-06-01

    In trypanosomatids the involvement of mitochondrial complex I in NADH oxidation has long been debated. Here, we took advantage of natural Trypanosoma cruzi mutants which present conspicuous deletions in ND4, ND5 and ND7 genes coding for complex I subunits to further investigate its functionality. Mitochondrial bioenergetics of wild type and complex I mutants showed no significant differences in oxygen consumption or respiratory control ratios in the presence of NADH-linked substrates or FADH(2)-generating succinate. No correlation could be established between mitochondrial membrane potentials and ND deletions. Since release of reactive oxygen species occurs at complex I, we measured mitochondrial H(2)O(2) formation induced by different substrates. Significant differences not associated to ND deletions were observed among the parasite isolates, demonstrating that these mutations are not important for the control of oxidant production. Our data support the notion that complex I has a limited function in T. cruzi.

  8. Molecular phylogeny of silk-producing insects based on 16S ribosomal RNA and cytochrome oxidase subunit I genes

    Indian Academy of Sciences (India)

    B. Mahendran; S. K. Ghosh; S. C. Kundu

    2006-04-01

    We have examined the molecular-phylogenetic relationships between nonmulberry and mulberry silkwormspecies that belong to the families Saturniidae, Bombycidae and Lasiocampidae using 16S ribosomal RNA (16S rRNA) and cytochrome oxidase subunit I (coxI) gene sequences. Aligned nucleotide sequences of 16S rRNA and coxI from 14 silk-producing species were used for construction of phylogenetic trees by maximum likelihood and maximum parsimony methods. The tree topology on the basis of 16S rRNA supports monophyly for members of Saturniidae and Bombycidae. Weighted parsimony analysis weighted towards transversions relative to transitions (ts, tv4) for coxI resulted in more robust bootstrap support over unweighted parsimony and favours the 16S rRNA tree topology. Combined analysis reflected clear biogeographic pattern, and agrees with morphological and cytological data.

  9. Functional characterization of the promoter for the mouse SPTLC2 gene, which encodes subunit 2 of serine palmitoyltransferase

    Science.gov (United States)

    Linn, Stephen C.; Andras, Lindsay M.; Kim, Hee-Sook; Wei, Jia; Nagiec, M. Marek; Dickson, Robert C.; Merrill, Alfred H.

    2006-01-01

    A series of luciferase reporter constructs was prepared from a 1035-bp fragment of mouse genomic DNA flanking the 5′ -coding sequence for the SPTLC2 subunit of serine palmitoyltransferase, the initial enzyme of de novo sphingolipid biosynthesis. The full-length DNA fragment promoted strong reporter gene expression in NIH3T3 cells while deletion and site-directed mutagenesis indicated that the proximal 335 bp contain initiator and downstream promoter elements, two proximal GC boxes that appear to stimulate transcription in a cooperative manner, and several additional elements whose activity cannot be accounted for by known factor binding sites. These findings provide insight into the control mechanisms for transcription of mammalian SPTLC2. PMID:17070807

  10. Phylogenetic position of Dysteria derouxi (Ciliophora:Phyllopharyngea: Dysteriida) inferred from the small subunit ribosomal RNA gene sequence

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    The complete small subunit rRNA (SSrRNA) gene sequence of a marine ciliate, Dysteria derouxi Gong and Song, 2004, was determined to be of 1 708 nucleotides. The phylogenetic position of this species within the class Phyllopharyngea was deduced using distance matrix, maximum parsimony and maximum likelihood methods. Dysteria derouxi, together with other available ciliates of the class Phyllopharyngea, forms a monophyletic clade with strong bootstrap support in the distance matrix, maximum parsimony and likelihood tree construction methods, while the dysterids are, as a monophyletic group, phylogenetically close to the clade of chlamydodontids [values of 100% LS(least-squares), 100% NJ(neighbor-joining)]. In addition, the trees indicate that dysteriids may be a higher or specialized group within the class, which corresponds well to the morphology and infraciliature.

  11. Phylogenetic position of three Condylostoma species(Protozoa,Ciliophora,Heterotricheal inferred from the small subunit rRNA gene sequence

    Institute of Scientific and Technical Information of China (English)

    Wenbo Guo; Shan Gao; Weibo Song; Khaled A.S.A1-Rasheid; Chen Shao; Miao Miao; Saleh A.A1-Farraj; Saleh A.A1-Qurishy; Zigui Chen; Zhenzhen Yi

    2008-01-01

    The systematically poorly known ciliate genus Conadylostoma was erected by Vincent in 1826.About 10 morphotypes have been reported,but any molecular investigations concerning this group SO far are lacking.In this work,the small subunit ribosomal RNA (SS rRNA)gene of three marine Conaylostoma species was sequenced,by which the phylogenetic trees were constructed by distance-matrix,maximum parsimony and Bayesian inference methods.The results show that(1)all the trees have similar topologies with high supports;(2)Condylostoma is mostly related to the genus Condylostentor;and(3)three Condylostoma species as well as Conadylostentor auriculatus cluster together and form a sister group with other heterotrichs.This is moderately consistent with the assessment of phylo-genetic relationships of Conaylostoma-related heterotrichs from the morphological information.The phylogenetic relationship of some other related heterotrichs,Peritromus,Fotlictllina,Stentor and Blepharisma,has been also discussed.

  12. Identification of a new human mtDNA polymorphism (A14290G in the NADH dehydrogenase subunit 6 gene

    Directory of Open Access Journals (Sweden)

    M. Houshmand

    2006-06-01

    Full Text Available Leber's hereditary optic neuropathy (LHON is a maternally inherited form of retinal ganglion cell degeneration leading to optic atrophy in young adults. Several mutations in different genes can cause LHON (heterogeneity. The ND6 gene is one of the mitochondrial genes that encodes subunit 6 of complex I of the respiratory chain. This gene is a hot spot gene. Fourteen Persian LHON patients were analyzed with single-strand conformational polymorphism and DNA sequencing techniques. None of these patients had four primary mutations, G3460A, G11788A, T14484C, and G14459A, related to this disease. We identified twelve nucleotide substitutions, G13702C, T13879C, T14110C, C14167T, G14199T, A14233G, G14272C, A14290G, G14365C, G14368C, T14766C, and T14798C. Eleven of twelve nucleotide substitutions had already been reported as polymorphism. One of the nucleotide substitutions (A14290G has not been reported. The A14290G nucleotide substitution does not change its amino acid (glutamic acid. We looked for base conservation using DNA star software (MEGALIGN program as a criterion for pathogenic or nonpathogenic nucleotide substitution in A14290G. The results of ND6 gene alignment in humans and in other species (mouse, cow, elegans worm, and Neurospora crassa mold revealed that the 14290th base was not conserved. Fifty normal controls were also investigated for this polymorphism in the Iranian population and two had A14290G polymorphism (4%. This study provides evidence that the mtDNA A14290G allele is a new nonpathogenic polymorphism. We suggest follow-up studies regarding this polymorphism in different populations.

  13. Implication of human UGT2B7, 2B15 and 2B17 in 19-norandrosterone metabolism

    Directory of Open Access Journals (Sweden)

    Emmanuel eStrahm

    2013-06-01

    Full Text Available Nandrolone (19-nortestosterone is an anabolic androgenic steroid commonly abused for doping purposes. Nandrolone is mainly metabolized in the liver into 19-norandrosterone prior to glucuronidation and excretion through urine over an extended period of time. Several UGTs (i.e. UGT2B7, UGT2B15 and UGT2B17 are thought to be the major enzymes responsible for conjugation of androgens in human. An in-vitro study using recombinant enzymes expressed in insect cells showed that UGT1A4 and UGT2B7 are the two main enzymes responsible of 19-norandrosterone glucuronidation. However, the identity of the enzyme involved in nandrolone metabolism in vivo together with their relative contribution and regulation remain unknown.Inhibition assays using human liver microsomes incubated with 19-norandrosterone and selective inhibitors confirmed that UGT2B7 and UGT2B15 are involved in 19-norandrosterone glucuronidation, since the presence of the specific UGT2B7 and UGT2B15 inhibitors gemfibrosil and valproic acid inhibited the 19-norandrosterone glucuronidation by 35 and 45 %, respectively. Human liver microsomes were genotyped for UGT2B15 D85Y, UGT2B7 H268Y and the UGT2B17 deletion polymorphism. The glucuronidation activity on 19 norandrosterone was significantly higher in UGT2B15 DD than in the other UGT2B15 genotypes (p<0.05. Moreover, human liver cancer HepG2 cells were exposed to androgens to determine if the transcriptional activity of the genes of interest was affected. Only UGT2B7 mRNA expression was significantly increased (1.8-folds after incubation with nandrolone decanoate.These results show that the UGT2B7 and UGT2B15 are involved in 19-norandrosterone glucuronidation and that the UGT2B15 polymorphism (D85Y is the only UGT genetic variation that influences the glucuronidation activity. This could partly explain the inter-individual variation in 19 norandrosterone excretion.

  14. The RFC2 gene encoding a subunit of replication factor C of Saccharomyces cerevisiae.

    OpenAIRE

    Noskov, V; Maki, S.; Kawasaki, Y.; Leem, S H; Ono, B; Araki, H; Pavlov, Y; Sugino, A

    1994-01-01

    Replication Factor C (RF-C) of Saccharomyces cerevisiae is a complex that consists of several different polypeptides ranging from 120- to 37 kDa (Yoder and Burgers, 1991; Fien and Stillman, 1992), similar to human RF-C. We have isolated a gene, RFC2, that appears to be a component of the yeast RF-C. The RFC2 gene is located on chromosome X of S. cerevisiae and is essential for cell growth. Disruption of the RFC2 gene led to a dumbbell-shaped terminal morphology, common to mutants having a def...

  15. Pyrosequencing method for the detection of UGT1A3 and UGT2B7 gene polymorphism in Chinese Han population%焦磷酸测序技术检测UGT1A3和UGT2B7在中国汉族人群中的基因多态性

    Institute of Scientific and Technical Information of China (English)

    许景峰; 赵刚涛; 许茜; 丁媛媛; 杨凡; 阎晨霞

    2011-01-01

    目的 建立焦磷酸测序技术(pyrosequencing)研究二相代谢酶UGT1 A3和UGT2B7基因多态性在中国汉族人群中的分布.方法 应用带生物素标记扩增引物并经PCR扩增和Beads分离,制备UGT1 A3和UGT2B7焦磷酸测序单倍摸板.在PYroMarkID焦磷酸测序上进行焦磷酸测序,检测233血样的DNA标本的17个SNP位点,以确定血样DNA标本的的基因型.结果 233例血样的DNA标本中,UGTA3等位基因有9种表型,分别为UGT1A3*1*1、UGT1A3*1*2、UGT1A3*1*3、UGT1A3*1*4、UGT1A3* 1*5、UGT1A3*2*3、UGT1A3*2*4、UGT1A3*3*3和UGT1A3*3*5.UGT2B7-1和UGT2B7-2各有3种基因型,分别为G/G型、G/T型、T/T和C/C型、C/T型、T/T型.结论 我国汉族人群中UGT1A3和UGT2B7基因突变较高.%Objective To establish a pyrosequencing-based method to detect UGT1A3 and UGT2B7 gene polymorphism in Chinese population. Methods Biotin-labeled primers, amplification of PCR, separation of Beads, and preparation of UGT1A3 and UGT2B7 pyrosequencing single-stranded template of mutational sites were done to determine the allelic frequency of the UGT1A3 and UGT2B7 mutant in 233 Chinese subjects by pyrosequencing apparatus. Re-sults There were 9 genetypes of the allele of UGT1A3: UGT1A3 * 1 * 1, UGT1A3 * 1 * 2, UGT1A3 * 1 * 3, UGT1A3*1*4, UGT1A3*1*5, UGT1A3*2*3, UGT1A3*2*4, UGT1A3*3*3, and UGT1 A3 * 3 * 5 respectively. There were 3 genetypes based on the presence of UGT2B7-1: genetypes G/G, G/T, and T/T; and 3 genetypes of the allele of UGT2B7-2: C/C, C/T, and T/T. Conclusion The frequency of the UGT1A3 and UGT2B7 mutant allele is high in Chinese population.

  16. The Vacuolar ATPase from Entamoeba histolytica: Molecular cloning of the gene encoding for the B subunit and subcellular localization of the protein

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    Luna-Arias Juan

    2008-12-01

    Full Text Available Abstract Background Entamoeba histolytica is a professional phagocytic cell where the vacuolar ATPase plays a key role. This enzyme is a multisubunit complex that regulates pH in many subcellular compartments, even in those that are not measurably acidic. It participates in a wide variety of cellular processes such as endocytosis, intracellular transport and membrane fusion. The presence of a vacuolar type H+-ATPase in E. histolytica trophozoites has been inferred previously from inhibition assays of its activity, the isolation of the Ehvma1 and Ehvma3 genes, and by proteomic analysis of purified phagosomes. Results We report the isolation and characterization of the Ehvma2 gene, which encodes for the subunit B of the vacuolar ATPase. This polypeptide is a 55.3 kDa highly conserved protein with 34 to 80% identity to orthologous proteins from other species. Particularly, in silico studies showed that EhV-ATPase subunit B displays 78% identity and 90% similarity to its Dictyostelium ortholog. A 462 bp DNA fragment of the Ehvma2 gene was expressed in bacteria and recombinant polypeptide was used to raise mouse polyclonal antibodies. EhV-ATPase subunit B antibodies detected a 55 kDa band in whole cell extracts and in an enriched fraction of DNA-containing organelles named EhkOs. The V-ATPase subunit B was located by immunofluorescence and confocal microscopy in many vesicles, in phagosomes, plasma membrane and in EhkOs. We also identified the genes encoding for the majority of the V-ATPase subunits in the E. histolytica genome, and proposed a putative model for this proton pump. Conclusion We have isolated the Ehvma2 gene which encodes for the V-ATPase subunit B from the E. histolytica clone A. This gene has a 154 bp intron and encodes for a highly conserved polypeptide. Specific antibodies localized EhV-ATPase subunit B in many vesicles, phagosomes, plasma membrane and in EhkOs. Most of the orthologous genes encoding for the EhV-ATPase subunits

  17. Detection of the enzymatically-active polyhydroxyalkanoate synthase subunit gene, phaC, in cyanobacteria via colony PCR.

    Science.gov (United States)

    Lane, Courtney E; Benton, Michael G

    2015-12-01

    A colony PCR-based assay was developed to rapidly determine if a cyanobacterium of interest contains the requisite genetic material, the PHA synthase PhaC subunit, to produce polyhydroxyalkanoates (PHAs). The test is both high throughput and robust, owing to an extensive sequence analysis of cyanobacteria PHA synthases. The assay uses a single detection primer set and a single reaction condition across multiple cyanobacteria strains to produce an easily detectable positive result - amplification via PCR as evidenced by a band in electrophoresis. In order to demonstrate the potential of the presence of phaC as an indicator of a cyanobacteria's PHA accumulation capabilities, the ability to produce PHA was assessed for five cyanobacteria with a traditional in vivo PHA granule staining using an oxazine dye. The confirmed in vivo staining results were then compared to the PCR-based assay results and found to be in agreement. The colony PCR assay was capable of successfully detecting the phaC gene in all six of the diverse cyanobacteria tested which possessed the gene, while exhibiting no undesired product formation across the nine total cyanobacteria strains tested. The colony PCR quick prep provides sufficient usable DNA template such that this assay could be readily expanded to assess multiple genes of interest simultaneously.

  18. Gene aberrations of RRM1 and RRM2B and outcome of advanced breast cancer after treatment with docetaxel with or without gemcitabine

    DEFF Research Database (Denmark)

    Jørgensen, Charlotte Lt; Ejlertsen, Bent; Bjerre, Karsten D

    2013-01-01

    of the hypothesis that aberrations of RRM1 or RRM2B, neither individually nor in combination, are associated with an altered clinical outcome following chemotherapy with gemcitabine in combination with docetaxel compared to docetaxel alone in advanced breast cancer patients....... agent docetaxel in advanced breast cancer patients. Methods Primary tumor samples from patients randomly assigned to gemcitabine plus docetaxel or docetaxel alone were analyzed for RRM1 and RRM2B copy number changes using Fluorescence In Situ Hybridization (FISH) technology with probes covering...... endpoint. Overall survival (OS) and response rate (RR) were secondary endpoints. Associations between RRM1/CEN-11 and/or RRM2B/CEN-8 ratios and time-to-event endpoints were analyzed by unadjusted and adjusted Cox proportional hazards regression models. Heterogeneity of treatment effects on TTP and OS...

  19. Inefficiency in GM2 ganglioside elimination by human lysosomal beta-hexosaminidase beta-subunit gene transfer to fibroblastic cell line derived from Sandhoff disease model mice.

    Science.gov (United States)

    Itakura, Tomohiro; Kuroki, Aya; Ishibashi, Yasuhiro; Tsuji, Daisuke; Kawashita, Eri; Higashine, Yukari; Sakuraba, Hitoshi; Yamanaka, Shoji; Itoh, Kohji

    2006-08-01

    Sandhoff disease (SD) is an autosomal recessive GM2 gangliosidosis caused by the defect of lysosomal beta-hexosaminidase (Hex) beta-subunit gene associated with neurosomatic manifestations. Therapeutic effects of Hex subunit gene transduction have been examined on Sandhoff disease model mice (SD mice) produced by the allelic disruption of Hexb gene encoding the murine beta-subunit. We demonstrate here that elimination of GM2 ganglioside (GM2) accumulated in the fibroblastic cell line derived from SD mice (FSD) did not occur when the HEXB gene only was transfected. In contrast, a significant increase in the HexB (betabeta homodimer) activity toward neutral substrates, including GA2 (asialo-GM2) and oligosaccharides carrying the terminal N-acetylglucosamine residues at their non-reducing ends (GlcNAc-oligosaccharides) was observed. Immunoblotting with anti-human HexA (alphabeta heterodimer) serum after native polyacrylamide gel electrophoresis (Native-PAGE) revealed that the human HEXB gene product could hardly form the chimeric HexA through associating with the murine alpha-subunit. However, co-introduction of the HEXA encoding the human alpha-subunit and HEXB genes caused significant corrective effect on the GM2 degradation by producing the human HexA. These results indicate that the recombinant human HexA could interspeciesly associate with the murine GM2 activator protein to degrade GM2 accumulated in the FSD cells. Thus, therapeutic effects of the recombinant human HexA isozyme but not human HEXB gene product could be evaluated by using the SD mice.

  20. Molecular characterization of Fasciola hepatica and phylogenetic analysis based on mitochondrial (nicotiamide adenine dinucleotide dehydrogenase subunit I and cytochrome oxidase subunit I) genes from the North-East of Iran

    Science.gov (United States)

    Reaghi, Saber; Haghighi, Ali; Harandi, Majid Fasihi; Spotin, Adel; Arzamani, Kourosh; Rouhani, Soheila

    2016-01-01

    Aim: Fascioliasis is one of the most zoonotic diseases with global extension. As the epidemiological distribution of Fasciola may lead to various genetic patterns of the parasite, the aim of this study is to identify Fasciola hepatica based on spermatogenesis, and phylogenetic analysis using mitochondrial (nicotiamide adenine dinucleotide dehydrogenase subunit I [ND1] and cytochrome oxidase subunit I) gene marker. Materials and Methods: In this study, 90 F. hepatica collected from 30 cattle at slaughterhouse located in three different geographical locations in the North-East of Iran were evaluated based on spermatogenetic ability and internal transcribed spacer 1 gene restriction fragment length polymorphism pattern. Genetic diversity and phylogenetic relationship using mtDNA gene marker for the isolates from the North-East of Iran, and other countries were then analyzed. Results: Partial sequences of mtDNA showed eight haplotypes in both genes. The phylogenic analysis using neighbor joining as well as maximum likelihood methods showed similar topologies of trees. Pairwise fixation index between different F. hepatica populations calculated from the nucleotide data set of ND1 gene are statistically significant and show the genetic difference. Conclusion: F. hepatica found in this region of Iran has different genetic structures through the other Fasciola populations in the world. PMID:27733809

  1. Design of a molecular method for subspecies specific identification of Klebsiella pneumoniae by using the 16S ribosomal subunit gene

    Directory of Open Access Journals (Sweden)

    Nelson Enrique Arenas

    2009-12-01

    Full Text Available Introduction: Rhinoscleroma is caused by Klebsiella pneumoniae subsp. rhinoscleromatis and the ozena infections caused by K. pneumoniae subsp. ozaenae, both infections affect the upper respiratory tract. In the first clinical phases the symptoms are unspecific, and the disease can be misdiagnosed as a common cold, therefore antimicrobial therapy cannot reach effective results and patients must be following up for several years since the infection became chronic. Objective: To identify Klebsiella subspecies using a specific assay based on amplicons restriction of a gene which encodes 16S subunit ribosomal (rDNA16S. Methodology: Specific restriction patterns were generated; using reported sequences from rDNA16S gene and bioinformatics programs MACAW, PFE, GENEDOC and GENE RUNNER. Amplification and restriction assays were standardized. Results: Predictions in silico allowed us to propose an algorithm for Klebsiella species and subspecies identification. Two reference strains were included and two clinical isolates which were biotyped and identified by the proposed method. rDNA16S gene restriction patterns showed differences regarding the initially identified species for conventional methods. Additionally two patterns of bands were observed for K. pneumoniae subsp. rhinoscleromatis, indicating the polymorphisms presence in the rDNA16S gene. Conclusions: We confirmed the difficulty to identify K. pneumoniae subspecies by conventional methods. Implementation of this technique could allow accurate and rapid differentiation among K. pneumoniae subsp. ozaenae and K. pneumoniae subsp. rhinoscleromatis the aetiological agents of two frequently misdiagnosed infections. Antimicrobial therapy usually could be ineffective, especially in chronic patients. Finally we consider very important to enlarge the study by using more clinical and reference strains.

  2. Design of a molecular method for subspecies specific identification of Klebsiella pneumoniae by using the 16S ribosomal subunit gene.

    Directory of Open Access Journals (Sweden)

    Nelson Enrique Arenas

    2009-12-01

    Full Text Available Introduction: Rhinoscleroma is caused by Klebsiella pneumoniae rhinoscleromatis and the ozena infections caused by K. pneumoniae ozaenae, both infections affect the upper respiratory tract. In the first clinical phases the symptoms are unspecific, and the disease can be misdiagnosed as a common cold, therefore antimicrobial therapy cannot reach effective results and patients must be following up for several years since the infection became chronic. Objective: To identify Klebsiella subspecies using a specific assay based on amplicons restriction of a gene which encodes 16S subunit ribosomal (rDNA16S.Methodology: Specific restriction patterns were generated; using reported sequences from rDNA16S gene and bioinformatics programs MACAW, PFE, GENEDOC and GENE RUNNER. Amplification and restriction assays were standardized. Results: Predictions in silico allowed to propose an algorithm for Klebsiella species and subspecies identification. Two reference strains were included and two clinical isolates which were biotyped and identified by the proposed method. rDNA16S gene restriction patterns showed differences regarding the initially identified species for conventional methods. Additionally two patterns of bands were observed for K. pneumoniae rhinoscleromatis, indicating the polymorphisms presence in the rDNA16S gene. Conclusions: It was confirmed the difficulty to identify K. pneumoniae subspecies by conventional methods. Implementation of this technique could allow an accurate and rapid differentiation among K. pneumoniae ozaenae and K. pneumoniae rhinoscleromatis aetiological agents of two frequently misdiagnosed infections. Antimicrobial therapy usually could be ineffective, especially in chronic patients. Finally it is considered very important to enlarge the study by using more clinical and reference strains.

  3. Cloning of a yeast gene coding for the glutamate synthase small subunit (GUS2) by complementation of Saccharomyces cerevisiae and Escherichia coli glutamate auxotrophs.

    Science.gov (United States)

    González, A; Membrillo-Hernández, J; Olivera, H; Aranda, C; Macino, G; Ballario, P

    1992-02-01

    A Saccharomyces cerevisiae glutamate auxotroph, lacking NADP-glutamate dehydrogenase (NADP-GDH) and glutamate synthase (GOGAT) activities, was complemented with a yeast genomic library. Clones were obtained which still lacked NADP-GDH but showed GOGAT activity. Northern analysis revealed that the DNA fragment present in the complementing plasmids coded for a 1.5kb mRNA. Since the only GOGAT enzyme so far purified from S. cerevisiae is made up of a small and a large subunit, the size of the mRNA suggested that the cloned DNA fragment could code for the GOGAT small subunit. Plasmids were purified and used to transform Escherichia coli glutamate auxotrophs. Transformants were only recovered when the recipient strain was an E. coli GDH-less mutant lacking the small GOGAT subunit. These data show that we have cloned the structural gene coding for the yeast small subunit (GUS2). Evidence is also presented indicating that the GOGAT enzyme which is synthesized in the E. coli transformants is a hybrid comprising the large E. coli subunit and the small S. cerevisiae subunit.

  4. Phenotypical Manifestations of Mutations in the Genes Encoding Subunits of the Cardiac Sodium Channel

    NARCIS (Netherlands)

    Wilde, Arthur A. M.; Brugada, Ramon

    2011-01-01

    Variations in the gene encoding for the major sodium channel (Na(v)1.5) in the heart, SCN5A, has been shown to cause a number of arrhythmia syndromes (with or without structural changes in the myocardium), including the long-QT syndrome (type 3), Brugada syndrome, (progressive) cardiac conduction di

  5. Genes encoding biotin carboxylase subunit of acetyl-CoA carboxylase from Brassica napus and parental species: cloning, expression patterns, and evolution

    Science.gov (United States)

    Comparative genomics is a useful tool to investigate gene and genome evolution. Biotin carboxylase (BC), an important subunit of heteromeric ACCase that is a rate-limiting enzyme in fatty acid biosynthesis in dicots, catalyzes ATP, biotin-carboxyl-carrier protein and CO2 to form carboxybiotin-carbo...

  6. Phylogeny, clinical associations, and diagnostic utility of the pilin subunit gene (sfpA) of sorbitol-fermenting, enterohemorrhagic Escherichia coli O157:H-

    NARCIS (Netherlands)

    Friedrich, Alexander W; Nierhoff, Katja V; Bielaszewska, Martina; Mellmann, Alexander; Karch, Helge

    2004-01-01

    The plasmid-borne sfpA gene encodes the pilin subunit in sorbitol-fermenting (SF) enterohemorrhagic Escherichia coli (EHEC) O157:H-. We investigated the distribution of sfpA among 600 E. coli isolates comprising the complete E. coli standard reference (ECOR) and diarrheagenic E. coli (DEC) strain co

  7. Gene expression profiles and phosphorylation patterns of AMP-activated protein kinase subunits in various mesenchymal cell types

    Institute of Scientific and Technical Information of China (English)

    Wang Yugang; Fan Qiming; Ma Rui; Lin Wentao; Tang Tingting

    2014-01-01

    Background Recent studies on bone have shown an endocrine role of the skeleton,which could be impaired in various human diseases,including osteoporosis,obesity,and diabetes-associated bone diseases.As a sensor and regulator of energy metabolism,AMP-activated protein kinase (AMPK) may also play an important role in the regulation of bone metabolism.The current study aimed to establish the expression profiles and phosphorylation patterns of AMPK subunits in several mesenchymal cell types.Methods Reverse transcription-polymerase chain reaction (PCR) for relative quantification,real-time PCR for absolute quantification,and Western blotting were used to investigate the gene expression profiles and phosphorylation patterns of AMPK subunits in several mesenchymal cell types,including primary human mesenchymal stem cells (hMSCs) and hFOB,Saos-2,C3H/10T1/2,MC3T3-E1,3T3-L1,and C2C12 cells.Results AMPKα1 and AMPKβ1 mRNAs were abundantly expressed in all cell types.AMPKY1 mRNA was abundantly expressed in C3H/10T1/2,MC3T3-E1,3T3-L1,and C2C12 but not detected in human-derived cell types.AMPKY2 mRNA was mildly expressed in all cell types.AMPKα1 protein was highly expressed in all cell types and AMPKα2 protein was highly expressed only in hFOB and Saos-2 cells.AMPKβ1 protein was abundantly expressed in all cell types except for Saos-2,in which AMPKβ2 protein overwhelmed AMPKβ1 expression.AMPKy1 and AMPKY2 proteins were expressed in C3H/10T1/2,MC3T3-E1,3T3-L1,and C2C12 cells and only AMPKY2 protein was expressed in hMSCs,hFOB and Saos2 cells.AMPKα was phosphorylated at Thr172 and Ser485 and AMPKβ1 was phosphorylated at Ser108 and Ser182 in all cell types with a specific pattern in each cell type.Conclusion The combination of AMPK α,β,and Y subunits and phosphorylation of AMPKα (Thr172 and Ser485) and AMPKβ1 (Ser108 and Ser182) showed a specific pattern in each cell type.

  8. Sequence Variation of the Pertussis Toxin S1 Subunit Encoding Gene in the Clinical Isolates of Bordetella pertussis in Iran

    Directory of Open Access Journals (Sweden)

    Hosseinpour

    2015-08-01

    Full Text Available Background Whooping cough (pertussis is an acute respiratory disease caused by Bordetella pertussis (B. pertussis. Pertussis toxin is an important virulence factor of B. pertussis and plays a major role in the immune and inflammatory responses. Likewise, allelic variations in the genes of virulence factors have led to the non-responsiveness of the new strains to both whole-cell and acellular vaccines. Given the importance of pertussis vaccine, we sought to address the lack of fundamental studies on the polymorphisms of the virulence genes of B. pertussis in Iran. Objectives The aim of this study was to identify the polymorphisms of the pertussis toxin S1 subunit (ptxS1 gene in the circulating strains and compare them to the vaccine strain. Patients and Methods In this study, 50 strains of B. pertussis isolated from patients with pertussis were investigated in the pertussis reference laboratory of Pasteur institute of Iran. Cultivation, biochemical tests, and the specific antisera were used to confirm B. pertussis. The sequencing of the polymerase chain reaction products was performed to determine the ptxS1 alleles, and B. pertussis 134 was studied as the vaccine strain. Results The results showed that all the strains had the dominant allele ptxS1A. There were differences between the alleles of the clinical strains and the vaccine strain. Conclusions In recent years, a significant increase in the incidence of pertussis has been reported worldwide. Our findings regarding the allelic shift of the ptxS1 gene are similar to those reported in many European and American countries showing the difference of the dominant allele of ptxS1 between the circulating isolates and the vaccine strains.

  9. Establishment of a continuous culture system for Entamoeba muris and analysis of the small subunit rRNA gene

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    Kobayashi S.

    2009-06-01

    Full Text Available We established a culture system for Entamoeba muris (MG-EM-01 strain isolated from a Mongolian gerbil using a modified Balamuth’s egg yolk infusion medium supplemented with 4% adult bovine serum and Bacteroides fragilis cocultured with Escherichia coli. Further, encystation was observed in the culture medium. The morphological characteristics of E. muris are similar to those of Entamoeba coli (E. coli; moreover, the malic isoenzyme electrophoretic band, which shows species-specific electrophoretic mobility, of E. muris had almost the same mobility as that observed with the malic isoenzyme electrophorectic band of E. coli (UZG-EC-01 strain isolated from a gorilla. We determined the small subunit rRNA (SSU-rRNA gene sequence of the MG-EM-01 strain, and this sequence was observed to show 82.7% homology with that of the UZG-EC-01 strain. Further, the resultant phylogenetic tree for molecular taxonomy based on the SSU-rRNA genes of the 21 strains of the intestinal parasitic amoeba species indicated that the MG-EM-01 strain was most closely related to E. coli.

  10. Genetic diversity of Echinococcus granulosus in southwest China determined by the mitochondrial NADH dehydrogenase subunit 2 gene.

    Science.gov (United States)

    Wang, Jiahai; Wang, Ning; Hu, Dandan; Zhong, Xiuqin; Wang, Shuxian; Gu, Xiaobin; Peng, Xuerong; Yang, Guangyou

    2014-01-01

    We evaluated genetic diversity and structure of Echinococcus granulosus by analyzing the complete mitochondrial NADH dehydrogenase subunit 2 (ND2) gene in 51 isolates of E. granulosus sensu stricto metacestodes collected at three locations in this region. We detected 19 haplotypes, which formed a distinct clade with the standard sheep strain (G1). Hence, all 51 isolates were identified as E. granulosus sensu stricto (G1-G3). Genetic relationships among haplotypes were not associated with geographical divisions, and fixation indices (Fst) among sampling localities were low. Hence, regional populations of E. granulosus in the southwest China are not differentiated, as gene flow among them remains high. This information is important for formulating unified region-wide prevention and control measures. We found large negative Fu's Fs and Tajima's D values and a unimodal mismatch distribution, indicating that the population has undergone a demographic expansion. We observed high genetic diversity among the E. granulosus s. s. isolates, indicating that the parasite population in this important bioregion is genetically robust and likely to survive and spread. The data from this study will prove valuable for future studies focusing on improving diagnosis and prevention methods and developing robust control strategies.

  11. Apparent selection intensity for the cytochrome oxidase subunit I gene varies with mode of reproduction in echinoderms.

    Science.gov (United States)

    Foltz, David W; Hrincevich, Adam W; Rocha-Olivares, Axayácatl

    2004-10-01

    When most amino acid substitutions in protein-coding genes are slightly deleterious rather than selectively neutral, life history differences can potentially modify the effective population size or the selective regime, resulting in altered ratios of non-synonymous to synonymous substitutions among taxa. We studied substitution patterns for the mitochondrial cytochrome oxidase subunit I (COI) gene in a sea star genus (Leptasterias spp.) with an obligate brood-protecting mode of reproduction and small-scale population genetic subdivision, and compared the results to available COI sequences in nine other genera of echinoderms with pelagic larvae: three sea stars, five sea urchins and one brittle star. We predicted that this life history difference would be associated with differences in the ratio of non-synonymous (dN) to synonymous (dS) substitution rates. Leptasterias had a significantly greater dN/dS ratio (both between species and within species), a significantly smaller transition/transversion rate ratio, and a significantly lower average nucleotide diversity within species, than did the non-brooding genera. Other explanations for the results, such as altered mutation rates or selective sweeps, were not supported by the data analysis. These findings highlight the potential influence of reproductive traits and other life history factors on patterns of nucleotide substitution within and between species.

  12. A split and rearranged nuclear gene encoding the iron-sulfur subunit of mitochondrial succinate dehydrogenase in Euglenozoa

    Directory of Open Access Journals (Sweden)

    Gray Michael W

    2009-02-01

    Full Text Available Abstract Background Analyses based on phylogenetic and ultrastructural data have suggested that euglenids (such as Euglena gracilis, trypanosomatids and diplonemids are members of a monophyletic lineage termed Euglenozoa. However, many uncertainties are associated with phylogenetic reconstructions for ancient and rapidly evolving groups; thus, rare genomic characters become increasingly important in reinforcing inferred phylogenetic relationships. Findings We discovered that the iron-sulfur subunit (SdhB of mitochondrial succinate dehydrogenase is encoded by a split and rearranged nuclear gene in Euglena gracilis and trypanosomatids, an example of a rare genomic character. The two subgenic modules are transcribed independently and the resulting mRNAs appear to be independently translated, with the two protein products imported into mitochondria, based on the presence of predicted mitochondrial targeting peptides. Although the inferred protein sequences are in general very divergent from those of other organisms, all of the required iron-sulfur cluster-coordinating residues are present. Moreover, the discontinuity in the euglenozoan SdhB sequence occurs between the two domains of a typical, covalently continuous SdhB, consistent with the inference that the euglenozoan 'half' proteins are functional. Conclusion The discovery of this unique molecular marker provides evidence for the monophyly of Euglenozoa that is independent of evolutionary models. Our results pose questions about the origin and timing of this novel gene arrangement and the structure and function of euglenozoan SdhB.

  13. The effects of clobazam treatment in rats on the expression of genes and proteins encoding glucronosyltransferase 1A/2B (UGT1A/2B) and multidrug resistance‐associated protein-2 (MRP2), and development of thyroid follicular cell hypertrophy

    Energy Technology Data Exchange (ETDEWEB)

    Miyawaki, Izuru, E-mail: izuru-miyawaki@ds-pharma.co.jp; Tamura, Akitoshi; Matsumoto, Izumi; Inada, Hiroshi; Kunimatsu, Takeshi; Kimura, Juki; Funabashi, Hitoshi

    2012-12-15

    Clobazam (CLB) is known to increase hepatobiliary thyroxine (T4) clearance in Sprague–Dawley (SD) rats, which results in hypothyroidism followed by thyroid follicular cell hypertrophy. However, the mechanism of the acceleration of T4-clearance has not been fully investigated. In the present study, we tried to clarify the roles of hepatic UDP-glucronosyltransferase (UGT) isoenzymes (UGT1A and UGT2B) and efflux transporter (multidrug resistance–associated protein-2; MRP2) in the CLB-induced acceleration of T4-clearance using two mutant rat strains, UGT1A-deficient mutant (Gunn) and MRP2-deficient mutant (EHBR) rats, especially focusing on thyroid morphology, levels of circulating hormones (T4 and triiodothyronine (T3)) and thyroid-stimulating hormone (TSH), and mRNA or protein expressions of UGTs (Ugt1a1, Ugt1a6, and Ugt2b1/2) and MRP2 (Mrp). CLB induced thyroid morphological changes with increases in TSH in SD and Gunn rats, but not in EHBR rats. T4 was slightly decreased in SD and Gunn rats, and T3 was decreased in Gunn rats, whereas these hormones were maintained in EHBR rats. Hepatic Ugt1a1, Ugt1a6, Ugt2b1/2, and Mrp2 mRNAs were upregulated in SD rats. In Gunn rats, UGT1A mRNAs (Ugt1a1/6) and protein levels were quite low, but UGT2B mRNAs (Ugt2b1/2) and protein were prominently upregulated. In SD and Gunn rats, MRP2 mRNA and protein were upregulated to the same degree. These results suggest that MRP2 is an important contributor in development of the thyroid cellular hypertrophy in CLB-treated rats, and that UGT1A and UGT2B work in concert with MRP2 in the presence of MRP2 function to enable the effective elimination of thyroid hormones. -- Highlights: ► Role of UGT and MRP2 in thyroid pathology was investigated in clobazam-treated rats. ► Clobazam induced thyroid cellular hypertrophy in SD and Gunn rats, but not EHBR rats. ► Hepatic Mrp2 gene and protein were upregulated in SD and Gunn rats, but not EHBR rats. ► Neither serum thyroid hormones (T3/T4

  14. A Novel Approach to Functional Analysis of the Ribulose Bisphosphate Carboxylase Small Subunit Gene by Agrobacterium-Mediated Gene Silencing

    Institute of Scientific and Technical Information of China (English)

    Xiao-Fu Zhou; Peng-Da Ma; Ren-Hou Wang; Bo Liu; Xing-Zhi Wang

    2006-01-01

    A novel approach to virus-induced post-transcriptional gene silencing for studying the function of the ribulose bisphosphate carboxylase small subunlt (rbcS) gene was established and optimized using potato virus X vector and Nicotiana benthamiana as experimental material. The analysis of silencing phenomena,transcriptional level, protein expression, and pigment measurement showed that the expression of the rbcS endogenous gene was inactivated by the expression of a 500-bp homologous cDNA fragment carried in the virus vector.

  15. Initial analysis of the hemocyanin subunit type 1 (Hc1 gene) from Locusta migratoria manilensis.

    Science.gov (United States)

    Yin, Hong; Guan, Ni; Dong, Lijun; Yue, Qiaoyun; Yin, Xiangchu; Zhang, Daochuan

    2012-03-01

    Hemocyanins are copper-containing (Cu(+)) proteins that transport oxygen in many arthropods hemolymph. We characterized Hc1 gene from the grasshopper species Locusta migratoria manilensis. In particular, we cloned and sequenced the corresponding cDNAs and studied their expression at different developmental stages. The cDNA of Hc1 gene (GenBank accession no.:HQ213937) is 2271 bp in length and the open reading frame is 2016 bp, which encodes a 672 amino acids protein with a calculated molecular mass of 77.9 kD and the isoelectric point of 6.06. Sequence alignment analysis result showed that this gene shares 94.7% identity with Schistocerca americana EHP. In addition, analysis of quantitative RT-PCR indicated that, LmiHc1 was expressed in the embyro (24, 39, 62, 86, 144, and 193 h after hatch), nymphs (1st instar, 2nd instar, 3rd instar, 4th instar and 5th instar) and in adult. These results showed that Hc1 plays an important role in grasshopper, which may be related to an enhanced oxygen supply. Phylogenetic analysis of insecta based on Hc1 are basically consistent with the morphology.

  16. Identification and characterization of human neuronal voltage-gated calcium channel gamma 3 subunit gene

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    By homologous expressed sequence tag (EST) searching,one EST (GenBank: W29095) was obtained,which shows 75% identity in 435 bp overlap with the coding sequence of mouse Cacng2 gene. A 1 545 bp cDNA fragment was obtained from the nested polymerase chain reaction (PCR) and rapid applification of cDNA end (RACE) reaction in the human brain prefrontal cortex cDNA library and the human brain Ready cDNA with the primers designed on W29095. The fragment contained a 948-bp open reading frame (ORF) encoding 315 amino acids,and was named CACNG3. As it was identical to a BAC clone (GenBank: AC004125) from chromosome 16p12-p13.1,the CACNG3 gene was mapped to human chromosome 16p12-p13.1,and the coding region was composed of 4 exons. Reverse transcription PCR (RT-PCR) analysis showed that the CACNG3 gene expressed in human adult brain and fetal brain. Single strand comformation polymorphism (SSCP) analysis was performed in 3 pedigrees with autosomal recessive retinitis pigmentosa,8 pedigrees with autosomal recessive retinitis pigmentosa accompanied by deafness and 2 pedigrees with epilepsy,but no mutation was detected.

  17. New insight into the role of the β3 subunit of the GABAA-R in development, behavior, body weight regulation, and anesthesia revealed by conditional gene knockout

    Directory of Open Access Journals (Sweden)

    Hileman Stanley M

    2007-10-01

    Full Text Available Abstract Background The β3 subunit of the γ-aminobutyric acid type A receptor (GABAA-R has been reported to be important for palate formation, anesthetic action, and normal nervous system function. This subunit has also been implicated in the pathogenesis of Angelman syndrome and autism spectrum disorder. To further investigate involvement of this subunit, we previously produced mice with a global knockout of β3. However, developmental abnormalities, compensation, reduced viability, and numerous behavioral abnormalities limited the usefulness of that murine model. To overcome many of these limitations, a mouse line with a conditionally inactivated β3 gene was engineered. Results Gene targeting and embryonic stem cell technologies were used to create mice in which exon 3 of the β3 subunit was flanked by loxP sites (i.e., floxed. Crossing the floxed β3 mice to a cre general deleter mouse line reproduced the phenotype of the previously described global knockout. Pan-neuronal knockout of β3 was achieved by crossing floxed β3 mice to Synapsin I-cre transgenic mice. Palate development was normal in pan-neuronal β3 knockouts but ~61% died as neonates. Survivors were overtly normal, fertile, and were less sensitive to etomidate. Forebrain selective knockout of β3 was achieved using α CamKII-cre transgenic mice. Palate development was normal in forebrain selective β3 knockout mice. These knockouts survived the neonatal period, but ~30% died between 15–25 days of age. Survivors had reduced reproductive fitness, reduced sensitivity to etomidate, were hyperactive, and some became obese. Conclusion Conditional inactivation of the β3 gene revealed novel insight into the function of this GABAA-R subunit. The floxed β3 knockout mice described here will be very useful for conditional knockout studies to further investigate the role of the β3 subunit in development, ethanol and anesthetic action, normal physiology, and pathophysiologic processes.

  18. Nucleotide sequences of two fimbrial major subunit genes, pmpA and ucaA, from canine-uropathogenic Proteus mirabilis strains.

    Science.gov (United States)

    Bijlsma, I G; van Dijk, L; Kusters, J G; Gaastra, W

    1995-06-01

    Proteus mirabilis strains were isolated from dogs with urinary tract infection (UTI) and fimbriae were prepared from two strains. The N-terminal amino acid sequences of the major fimbrial subunits were determined and both sequences appeared identical to the N-terminal amino acid sequence of a urinary cell adhesin (UCA) (Wray, S. K., Hull, S. I., Cook, R. G., Barrish, J. & Hull, R. A., 1986, Infect Immun 54, 43-49). The genes of two different major fimbrial subunits were cloned using oligonucleotide probes that were designed on the basis of the N-terminal UCA sequence. Nucleotide sequencing revealed the complete ucaA gene of 540 bp (from strain IVB247) encoding a polypeptide of 180 amino acids, including a 22 amino acid signal sequence peptide, and the pmpA (P. mirabilis P-like pili) gene of 549 bp (from strain IVB219) encoding a polypeptide of 183 amino acids, including a 23 amino acid signal sequence. Hybridization experiments gave clear indications of the presence of both kinds of fimbriae in many UTI-related canine P. mirabilis isolates. However, the presence of these fimbriae could not be demonstrated in P. vulgaris or other Proteus-related species. Database analysis of amino acid sequences of major subunit proteins revealed that the UcaA protein shares about 56% amino acid identity with the F17A and F111A major fimbrial subunits from bovine enterotoxigenic Escherichia coli. In turn, the PmpA protein more closely resembled the pyelonephritis-associated pili (Pap)-like major subunit protein from UTI-related E. coli. The evolutionary relationship of UcaA, PmpA and various other fimbrial subunit proteins is presented in a phylogenetic tree.

  19. Synthesis, in vitro and in vivo pharmacology of a C-11 labeled analog of CP-101,606, ({+-})threo-1-(4-hydroxyphenyl)-2-[4-hydroxy-4-(p-[{sup 11}C]methoxyphenyl) peridino]-1-propanol, as a PET tracer for NR2B subunit-containing NMDA receptors

    Energy Technology Data Exchange (ETDEWEB)

    Haradahira, Terushi E-mail: terushi@nirs.go.jp; Maeda, Jun; Okauchi, Takashi; Zhang, Ming-Rong; Hojo, Junko; Kida, Takayo; Arai, Takuya; Yamamoto, Fumihiko; Sasaki, Shigeki; Maeda, Minoru; Suzuki, Kazutoshi; Suhara, Tetsuya

    2002-07-01

    A carbon-11 labeled methoxyl analog of CP-101,606, ({+-})threo-1-(4-hydroxyphenyl)-2-[4-hydroxy-4-(p-[{sup 11}C]methoxyphenyl) piperidino]-1-propanol [({+-})[{sup 11}C]1], was synthesized as a new subtype-selective PET radioligand for NMDA receptors. The in vitro binding studies using rat brain slices demonstrated that ({+-})[{sup 11}C]1 shows an extremely high-specific binding to the NR2B subunit of NMDA receptors. In contrast to the in vitro binding, the in vivo binding to mouse and monkey brains showed no apparent specific localization of the radioactivity in any of the brain regions. Metabolism and physicochemical properties such as the lipophilicity of ({+-})[{sup 11}C]1 seemed unlikely to affect the in vivo ({+-})[{sup 11}C]1 binding. Among the various endogenous ligands acting at the NMDA receptors, polyamines (spermine and spermidine) and divalent cations (Mg{sup 2+,} Zn{sup 2+,} and Ca{sup 2+}) strongly inhibited the in vitro ({+-})[{sup 11}C]1 binding. Thus, the present studies point to the possibility that the polyamines and cations behave as endogenous inhibitors for ({+-})[{sup 11}C]1 binding, leading to the loss of the specific binding in vivo.

  20. CK2(beta)tes gene encodes a testis-specific isoform of the regulatory subunit of casein kinase 2 in Drosophila melanogaster

    DEFF Research Database (Denmark)

    Kalmykova, Alla I; Shevelyov, Yuri Y; Polesskaya, Oksana O

    2002-01-01

    An earlier described CK2(beta)tes gene of Drosophila melanogaster is shown to encode a male germline specific isoform of regulatory beta subunit of casein kinase 2. Western-analysis using anti-CK2(beta)tes Ig revealed CK2(beta)tes protein in Drosophila testes extract. Expression of a CK2(beta...... and coimmunoprecipitation analysis of protein extract from Drosophila testes, we demonstrated an association between CK2(beta)tes and CK2alpha. Northern-analysis has shown that another regulatory (beta') subunit found recently in D. melanogaster genome is also testis-specific. Thus, we describe the first example of two...

  1. Cloning and characterization of genes encoding alpha and beta subunits of glutamate-gated chloride channel protein in Cylicocyclus nassatus.

    Science.gov (United States)

    Tandon, Ritesh; LePage, Keith T; Kaplan, Ray M

    2006-11-01

    The invertebrate glutamate-gated chloride channels (GluCls) are receptor molecules and targets for the avermectin-milbemycin (AM) group of anthelmintics. Mutations in GluCls are associated with ivermectin resistance in the soil dwelling nematode Caenorhabditis elegans and the parasitic nematode Cooperia oncophora. In this study, full-length cDNAs encoding alpha and beta subunits of GluCl were cloned and sequenced in Cylicocyclus nassatus, a common and important cyathostomin nematode parasite of horses. Both genes possess the sequence characteristics typical of GluCls, and phylogenetic analysis confirms that these genes are evolutionarily closely related to GluCls of other nematodes and flies. Complete coding sequences of C. nassatus GluCl-alpha and GluCl-beta were subcloned into pTL1 mammalian expression vector, and proteins were expressed in COS-7 cells. Ivermectin-binding characteristics were determined by incubating COS-7 cell membranes expressing C. nassatus GluCl-alpha and GluCl-beta proteins with [(3)H]ivermectin. In competitive binding experiments, fitting the data to a one site competition model, C. nassatus GluCl-alpha was found to bind [(3)H]ivermectin with a high amount of displaceable binding (IC(50)=208 pM). Compared to the mock-transfected COS-7 cells, the means of [(3)H]ivermectin binding were significantly different for C. nassatus GluCl-alpha and the Haemonchus contortus GluCl (HcGluCla) (p=0.018 and 0.023, respectively) but not for C. nassatus GluCl-beta (p=0.370). This is the first report of orthologs of GluCl genes and in vitro expression of an ivermectin-binding protein in a cyathostomin species. These data suggest the likelihood of a similar mechanism of action of AM drugs in these parasites, and suggest that mechanisms of resistance may also be similar.

  2. Cloning and expression analysis of GhDET3, a vacuolar H+-ATPase subunit C gene, from cotton

    Institute of Scientific and Technical Information of China (English)

    Zhongyi Xiao; Kunling Tan; Mingyu Hu; Peng Liao; Kuijun Chen; Ming Luo

    2008-01-01

    Vacuolar H+-ATPase was regarded as a key enzyme promoting the fiber cell elongation in cotton (Gossypium hirsuturm L.) through regulating turgot-driven pressure involved in polarity expansion of single cell fiber. The DET3, a V-ATPase subunit C, plays an impor-tant role in assembling subunits and regulating the enzyme activity, and is involved in Brassinosteroid-induced cell elongation. To ana-lyze the function of GhDET3 on the elongation of cotton fibers, seven candidates of ESTs were screened and contigged for a 5'-upstream sequence, and the 3'-RACE technique was used to clone the 3'-downstream sequence for the full length of GhDET3 gene. The full length of the target clone was 1,340 bp, including a 10 bp 5'-UTR, an ORF of 1,134 bp, and a 196 bp 3'-UTR. This cDNA sequence encoded a polypepide of 377 amino acid residues with a predicted molecular mass of 43 kDa and a basic isoelectric point of 5.58. Furthermore, a length of 3,410 bp sequence from genomic DNA of GhDET3 was also cloned by PCR. The deduced amino acid sequence had a high ho-mology with DET3 from Arabidopsis, rice, and maize. Quantitative real-time PCR (qRT-PCR) analysis showed that the GhDET3 expres-sion pattern was ubiquitous in all the tissues and organs detected. The result also revealed that the accumulation of GhDET3 mRNA reached the highest profile at the fiber elongation stage in 12 DPA (days post anthesis) fibers, compared with the lowest level at the fiber initiation stage in 0 DPA ovules (with fibers). The transcript accumulation in fibers and ovules shared the similar variation tendency. In addition, in vitro ovule culture experiment demonstrated that exogenous 24-EBL treatment to 4 DPA ovules (with fibers) was capable of increasing the expression level of GhDET3, and the mRNA accumulation of GhDET3 increased in transgenic FBP7::GhDET2 cotton fibers in vivo. These results indicate that GhDET3 gene plays a crucial role in cotton fiber elongation.

  3. The human mitochondrial NADH: Ubiquinone oxidoreductase 51-kDa subunit oxidoreductase 51-kDa subunit maps adjacent to the glutathione S-transferase P1-1 gene on chromosome 11q13

    Energy Technology Data Exchange (ETDEWEB)

    Spencer, S.R.; Taylor, J.B.; Cowell, I.G.; Xia, C.L.; Pemble, S.E.; Ketterer, B. (Univ. College and Middlesex School of Medicine, London (United Kingdom))

    1992-12-01

    The soluble glutathione transferases (GSTs) are a family of dimeric isoenymes catalyzing the conjugation of glutathione to hydrophobic electropiles. Their subunits can be grouped into four families, alpha, mu, pi, and theta, on the basis of their primary structures. In man, the pi class is represented by a single gene, GSTP1-1 (GST[pi]) localized to human chromosome 11, band q13. The oncogenes INT2, HSTF1, and PRAD1 are also localized at 11q13, and together with the GSTP1 locus and other gene loci mapped to 11q13, i.e., BCL1 and EMS1, they form a unit of DNA approximately 2000-2500 kb, known as the 11q13 amplicon, which is often amplified in a range of solid tumors. Any gene locus at 11q13 is of interest because it may influence tumorigenesis. 14 refs., 1 fig.

  4. Suggestive association between the C825T polymorphism of the G-protein beta3 subunit gene (GNB3) and clinical improvement with antipsychotics in schizophrenia.

    Science.gov (United States)

    Müller, Daniel J; De Luca, Vincenzo; Sicard, Tricia; King, Nicole; Hwang, Rudi; Volavka, Jan; Czobor, Pal; Sheitman, Brian B; Lindenmayer, Jean-Pierre; Citrome, Leslie; McEvoy, Joseph P; Lieberman, Jeffrey A; Meltzer, Herbert Y; Kennedy, James L

    2005-10-01

    G-proteins are composed of alpha, beta and gamma subunits. Once activated, these subunits play a major role in the conversion of external receptor activation into intracellular signals. The functional C825T polymorphism of the beta3 subunit gene (GNB3) has recently been shown to modulate antidepressant response, with the T-allele conferring an increased signaling and being associated with favorable antidepressant response. We hypothesized that this polymorphism may be associated with response to antipsychotics in a population of 145 chronic schizophrenic patients deriving from two study-samples and being mainly treated with clozapine for up to 6 months. Overall, the C/C genotype was significantly associated with relative clinical improvement as measured by Brief Psychiatric Rating Scale (BPRS) change scores after 6 and 12 weeks (ppoint to the role of intracellular mechanisms in antipsychotic response.

  5. B2B在线支付进化系统的信息基因测度模型%Measure Model for Information Gene of B2B Online Payment Evolutionary System

    Institute of Scientific and Technical Information of China (English)

    杨琦峰; 沈鑫; 宋平; 谢光萍; 王俊

    2011-01-01

    Each of the existing B2B online payment systems has its own limitation,and lack of an effective coordination mechanism for them. Based on the order parameter principle of synergy theory,participants' information genes were found as micro order parameter which decided the macro orderliness of B2B online payment evolution system. A participant information gene model was established on " identity characteristics degree + reputation degree" two - dimensional attribute; and order degree measurement algorithm was proposed. Then the cooperation mechanism of the evolutionary B2B online payment system was discussed following the information gene model.%针对现有B2B在线支付系统存在的局限性,缺乏有效协同运作机制的问题,基于协同论的序参量原理,发现参与者信息基因是决定B2B在线支付进化系统宏观有序性的微观序参量,构建了“身份特征值+信誉度”二维属性的参与者信息基因模型及有序度测度算法,讨论了B2B在线支付系统的协同进化问题.

  6. IDENTIFICATION OF 3 HUMAN PSEUDOGENES FOR SUBUNIT-VIB OF CYTOCHROME-C-OXIDASE - A MOLECULAR RECORD OF GENE EVOLUTION

    NARCIS (Netherlands)

    TAANMAN, JW; SCHRAGE, C; REUVEKAMP, P; BIJL, J; HARTOG, M; DEVRIES, H; AGSTERIBBE, E

    1991-01-01

    Three pseudogenes for the nuclear-encoded subunit VIb of cytochrome c oxidase (COX) were isolated by screening a human genomic library with cloned human cDNA coding for COX subunit VIb. The nucleotide sequences of the pseudogenes, designated PSI-COX6b-1, PSI-COX6b-2 and PSI-COX6b-3, were determined.

  7. Molecular characterization of Echinococcus granulosus from Peru by sequencing of the mitochondrial cytochrome C oxidase subunit 1 gene.

    Science.gov (United States)

    Sánchez, Elizabeth; Cáceres, Omar; Náquira, César; Garcia, David; Patiño, Gladys; Silvia, Herrera; Volotão, Aline C; Fernandes, Octavio

    2010-09-01

    Echinococcus granulosus, the etiologic agent of cystic echinococcosis (CE) in humans and other animal species, is distributed worldwide. Ten intra-specific variants, or genotypes (G1-G10), have been defined based on genetic diversity. To determine the genotypes present in endemic areas of Peru, samples were collected from cattle (44), sheep (41) and humans (14) from Junín, Puno Huancavelica, Cusco, Arequipa and Ayacucho. DNA was extracted from protoscolex and/or germinal layers derived from 99 E. granulosus isolates and used as templates to amplify the mitochondrial cytochrome C oxidase subunit 1 gene. The resulting polymerase chain reaction products were sequenced and further examined by sequence analysis. All isolates, independent of the host, exhibited the G1 genotype. Phylogenetic analysis showed that three isolates from Ayacucho shared the same cluster with microvariant G1(4). The G1 genotype is considered the most widespread and infectious form of E. granulosus worldwide and our results confirm that the same patterns apply to this country. Therefore, these findings should be taken into consideration in developing prevention strategies and control programs for CE in Peru.

  8. Mutations in Two Genes Encoding Different Subunits of a Receptor Signaling Complex Result in an Identical Disease Phenotype

    Science.gov (United States)

    Paloneva, Juha; Manninen, Tuula; Christman, Grant; Hovanes, Karine; Mandelin, Jami; Adolfsson, Rolf; Bianchin, Marino; Bird, Thomas; Miranda, Roxana; Salmaggi, Andrea; Tranebjærg, Lisbeth; Konttinen, Yrjö; Peltonen, Leena

    2002-01-01

    Polycystic lipomembranous osteodysplasia with sclerosing leukoencephalopathy (PLOSL), also known as “Nasu-Hakola disease,” is a globally distributed recessively inherited disease leading to death during the 5th decade of life and is characterized by early-onset progressive dementia and bone cysts. Elsewhere, we have identified PLOSL mutations in TYROBP (DAP12), which codes for a membrane receptor component in natural-killer and myeloid cells, and also have identified genetic heterogeneity in PLOSL, with some patients carrying no mutations in TYROBP. Here we complete the molecular pathology of PLOSL by identifying TREM2 as the second PLOSL gene. TREM2 forms a receptor signaling complex with TYROBP and triggers activation of the immune responses in macrophages and dendritic cells. Patients with PLOSL have no defects in cell-mediated immunity, suggesting a remarkable capacity of the human immune system to compensate for the inactive TYROBP-mediated activation pathway. Our data imply that the TYROBP-mediated signaling pathway plays a significant role in human brain and bone tissue and provide an interesting example of how mutations in two different subunits of a multisubunit receptor complex result in an identical human disease phenotype. PMID:12080485

  9. Strong inhibition of fimbrial 3 subunit gene transcription by a novel downstream repressive element in Bordetella pertussis.

    Science.gov (United States)

    Chen, Qing; Boulanger, Alice; Hinton, Deborah M; Stibitz, Scott

    2014-08-01

    The Bvg-regulated promoters for the fimbrial subunit genes fim2 and fim3 of Bordetella pertussis behave differently from each other both in vivo and in vitro. In vivo Pfim2 is significantly stronger than Pfim3 , even though predictions based on the DNA sequences of BvgA-binding motifs and core promoter elements would indicate the opposite. In vitro Pfim3 demonstrated robust BvgA∼P-dependent transcriptional activation, while none was seen with Pfim2 . This apparent contradiction was investigated further. By swapping sequence elements we created a number of hybrid promoters and assayed their strength in vivo. We found that, while Pfim3 promoter elements upstream of the +1 transcriptional start site do indeed direct Bvg-activated transcription more efficiently than those of Pfim2 , the overall promoter strength of Pfim3  in vivo is reduced due to sequences downstream of +1 that inhibit transcription more than 250-fold. This element, the DRE (downstream repressive element), was mapped to the 15 bp immediately downstream of the Pfim3 +1. Placing the DRE in different promoter contexts indicated that its activity was not specific to fim promoters, or even to Bvg-regulated promoters. However it does appear to be specific to Bordetella species in that it did not function in Escherichia coli.

  10. Molecular characterization of Echinococcus granulosusfrom Peru by sequencing of the mitochondrial cytochrome C oxidase subunit 1 gene

    Directory of Open Access Journals (Sweden)

    Elizabeth Sánchez

    2010-09-01

    Full Text Available Echinococcus granulosus, the etiologic agent of cystic echinococcosis (CE in humans and other animal species, is distributed worldwide. Ten intra-specific variants, or genotypes (G1-G10, have been defined based on genetic diversity. To determine the genotypes present in endemic areas of Peru, samples were collected from cattle (44, sheep (41 and humans (14 from Junín, Puno Huancavelica, Cusco, Arequipa and Ayacucho. DNA was extracted from protoscolex and/or germinal layers derived from 99 E. granulosus isolates and used as templates to amplify the mitochondrial cytochrome C oxidase subunit 1 gene. The resulting polymerase chain reaction products were sequenced and further examined by sequence analysis. All isolates, independent of the host, exhibited the G1 genotype. Phylogenetic analysis showed that three isolates from Ayacucho shared the same cluster with microvariant G1(4. The G1 genotype is considered the most widespread and infectious form of E. granulosusworldwide and our results confirm that the same patterns apply to this country. Therefore, these findings should be taken into consideration in developing prevention strategies and control programs for CE in Peru.

  11. Prevalent ciliate symbiosis on copepods: high genetic diversity and wide distribution detected using small subunit ribosomal RNA gene.

    Science.gov (United States)

    Guo, Zhiling; Liu, Sheng; Hu, Simin; Li, Tao; Huang, Yousong; Liu, Guangxing; Zhang, Huan; Lin, Senjie

    2012-01-01

    Toward understanding the genetic diversity and distribution of copepod-associated symbiotic ciliates and the evolutionary relationships with their hosts in the marine environment, we developed a small subunit ribosomal RNA gene (18S rDNA)-based molecular method and investigated the genetic diversity and genotype distribution of the symbiotic ciliates on copepods. Of the 10 copepod species representing six families collected from six locations of Pacific and Atlantic Oceans, 9 were found to harbor ciliate symbionts. Phylogenetic analysis of the 391 ciliate 18S rDNA sequences obtained revealed seven groups (ribogroups), six (containing 99% of all the sequences) belonging to subclass Apostomatida, the other clustered with peritrich ciliate Vorticella gracilis. Among the Apostomatida groups, Group III were essentially identical to Vampyrophrya pelagica, and the other five groups represented the undocumented ciliates that were close to Vampyrophrya/Gymnodinioides/Hyalophysa. Group VI ciliates were found in all copepod species but one (Calanus sinicus), and were most abundant among all ciliate sequences obtained, indicating that they are the dominant symbiotic ciliates universally associated with copepods. In contrast, some ciliate sequences were found only in some of the copepods examined, suggesting the host selectivity and geographic differentiation of ciliates, which requires further verification by more extensive sampling. Our results reveal the wide occurrence and high genetic diversity of symbiotic ciliates on marine copepods and highlight the need to systematically investigate the host- and geography-based genetic differentiation and ecological roles of these ciliates globally.

  12. Prevalent ciliate symbiosis on copepods: high genetic diversity and wide distribution detected using small subunit ribosomal RNA gene.

    Directory of Open Access Journals (Sweden)

    Zhiling Guo

    Full Text Available Toward understanding the genetic diversity and distribution of copepod-associated symbiotic ciliates and the evolutionary relationships with their hosts in the marine environment, we developed a small subunit ribosomal RNA gene (18S rDNA-based molecular method and investigated the genetic diversity and genotype distribution of the symbiotic ciliates on copepods. Of the 10 copepod species representing six families collected from six locations of Pacific and Atlantic Oceans, 9 were found to harbor ciliate symbionts. Phylogenetic analysis of the 391 ciliate 18S rDNA sequences obtained revealed seven groups (ribogroups, six (containing 99% of all the sequences belonging to subclass Apostomatida, the other clustered with peritrich ciliate Vorticella gracilis. Among the Apostomatida groups, Group III were essentially identical to Vampyrophrya pelagica, and the other five groups represented the undocumented ciliates that were close to Vampyrophrya/Gymnodinioides/Hyalophysa. Group VI ciliates were found in all copepod species but one (Calanus sinicus, and were most abundant among all ciliate sequences obtained, indicating that they are the dominant symbiotic ciliates universally associated with copepods. In contrast, some ciliate sequences were found only in some of the copepods examined, suggesting the host selectivity and geographic differentiation of ciliates, which requires further verification by more extensive sampling. Our results reveal the wide occurrence and high genetic diversity of symbiotic ciliates on marine copepods and highlight the need to systematically investigate the host- and geography-based genetic differentiation and ecological roles of these ciliates globally.

  13. siRNA干扰CLEC2B基因沉默对黑素细胞的影响%Effects of RNA Interference Inducing CLEC2B Gene Silence on Melanocytes

    Institute of Scientific and Technical Information of China (English)

    张峻岭; 柳君如; 徐士福; 程琳; ZHOU Youwen

    2013-01-01

    Objective To observe effect on that whether lymphocytes' supernatant could influence melanocytes proliferation or not when CLEC2B gene expression is silenced by establishing the lymphocytes transient silencing system, representing CLEC2B participation path in the immunity mechanism of vitiligo. Methods The lymphocytes lines Jurkat and melanocytes lines B16 were selected. CLEC2B gene was silenced by RNA interference technique, the proliferation of melanocytes and lymphocytes were detected by MTT. Results The lymphocyte proliferation had no significantly change, lymphocytes' supernatant could decrease the suppression to melanocytes proliferation (P<0.05), so as to create beneficial environment for melanocytes living rate. Conclusion Silencing CLEC2B gene eould influence melanocytes proliferation by the way of affecting lymphocytes'function. It shows that CLEC2B is participated in the immune-pathogenesis.%目的 探讨小干扰RNA(small interfering RNA,siRNA)诱导淋巴细胞中CLEC2B(C-type lectin domain family 2,member B)基因沉默后,其淋巴细胞上清液对黑素细胞增殖活性的影响,从而揭示CLEC2B基因参与白癜风发病的免疫学机制.方法 选用Jurkat淋巴瘤细胞及B16黑素瘤细胞为研究对象,应用RNA干扰技术沉默淋巴细胞CLEC2BmRNA的表达,MTT法检测CLEC2B基因沉默后淋巴细胞增殖情况及其上清液作用后对黑素细胞增殖情况.结果 CLEC2B基因沉默后淋巴细胞增殖无明显影响,CLEC2B基因沉默后淋巴细胞上清液对黑素细胞的抑制作用降低,黑素细胞存活率增加(P<0.05).结论 CLEC2B基因表达下调后减弱对黑素细胞的抑制作用,提示CLEC2B基因影响黑素细胞增殖活性,从而参与白癜风发病.

  14. Comparison of sequences of hypervariable region (HVR subunit S-1 gene of field isolate I-37 infectious bronchitis virus with Connecticut serotype

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    N.L.P Indi Dharmayanti

    2003-06-01

    Full Text Available Infectious Bronchitis is a contagious and acute respiratory disease in chickens caused by infectious bronchitis virus (IBV.Antigenic differences in IBV are associated with changes in the sequence of the spike glycoprotein (S. The subunit S1 which demonstrates more sequence variability than S-2 have been identified as hypervariable region (HVR-1 and 2. There were several IB virus field isolates included I-37 have been identified in Indonesia by serum neutralization method. However, gene sequence variation in HVR subunit S-1 had not yet been identified. Isolate I-37 was close to the serotype Connecticut 46 (Conn 46. The aim of this study is to identify sequence variation of HVR subunit S-1 gene of isolate I-37 produced by Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR and sequencing. Several procedures were carried out in the study including virus titration, propagation and was concentrated from the allantoic fluid infected with IBV. Then, RNA was extracted for RTPCR. urther the product was sequnced and its homology with IBV references from GenBank was compared by GenMac version 8.0. Result showed that isolate I-37 produced 515 bp of amplification product. Isolate I-37 and Conn 46 are same serotype, yet their HVR subunit S-1 nucleotides and amino acids (protein differ by 6.9% and 15.6% respectively. It might be concluded that isolate I-37 was variant of Conn 46.

  15. Interferon Alfa-2b Injection

    Science.gov (United States)

    Interferon alfa-2b injection is used to treat a number of conditions.Interferon alfa-2b injection is used alone or in ... Hodgkin's lymphoma (NHL; a slow-growing blood cancer). Interferon alfa-2b is in a class of medications ...

  16. The subunit gene Ldα1 of nicotinic acetylcholine receptors plays important roles in the toxicity of imidacloprid and thiamethoxam against Leptinotarsa decemlineata.

    Science.gov (United States)

    Qu, Yang; Chen, Jinhua; Li, Chenge; Wang, Qiang; Guo, Wenchao; Han, Zhaojun; Jiang, Weihua

    2016-02-01

    Nicotinic acetylcholine receptors (nAChRs) are pentameric ACh-gated ion channels. It is believed that nAChRs composed of different subunits may vary in their function and toxicological characteristics. Neonicotinoids are activators of nAChRs and important insecticides that are extensively used for crop protection and resistance has been developed by some pests. They are also major insecticides for the control of Leptinotarsa decemlineata, which is a destructive defoliator pest that invaded the Xinjiang region of China in the 1990s. However, little is known about the constitution or subunits of the target in this pest. In this study, the full-length cDNAs encoding four new nAChR subunits (named Ldα3, Ldα6, Ldα10, and Ldβ1) were cloned from L. decemlineata. These genes encode 822-, 753-, 672-, and 759-amino acid proteins, respectively, which share typical features of insect nAChRs subunits and closely resemble the corresponding subunits of the nAChRs from Tribolium castaneum. Temporal and spatial expression analyses showed that these genes, as well as the previously identified Ldα1, Ldα2, and Ldα8 genes, are widely expressed in all developmental stages, including eggs, larvae of various instars, pupae, and adults. All genes monitored were expressed at higher levels in the head than in the thorax and abdomen, except for Ldα10. Dietary ingestion of double-stranded RNA bacterially expressed for Ldα1 (dsLdα1) significantly reduced the mRNA level of Ldα1 in treated larvae and adults by 48.0% and 78.6%, respectively. Among the non-target genes, Ldα3, Ldα9, and Ldβ1 were significantly up-regulated in larvae. A toxicity bioassay showed that dsLdα1 treatment greatly decreased the sensitivity to imidacloprid and thiamethoxam in adults. The larval susceptibility to thiamethoxam but not to imidacloprid was also reduced because of the lower down-regulation of Ldα1. Thus, our results suggest that Ldα1 encodes a subunit of a functional nAChR that mediates the

  17. Coat protein mutations in an attenuated Cucumber mosaic virus encoding mutant 2b protein that lacks RNA silencing suppressor activity induces chlorosis with photosynthesis gene repression and chloroplast abnormalities in infected tobacco plants.

    Science.gov (United States)

    Mochizuki, Tomofumi; Yamazaki, Ryota; Wada, Tomoya; Ohki, Satoshi T

    2014-05-01

    In tobacco plants, the Cucumber mosaic virus (CMV) pepo strain induces mosaic symptoms, including pale green chlorosis and malformed tissues. Here, we characterized the involvement of 2b protein and coat protein (CP) in the development of mosaic symptoms. A 2b mutant (R46C) that lacks viral suppressor of RNA silencing (VSR) activity showed an asymptomatic phenotype with low levels of virus accumulation. Tomato spotted wilt virus NSs protein did not complement the virulence of the R46C, although it did restore high-level virus accumulation. However, R46C mutants expressing mutated CP in which the amino acid P129 was mutated to A, E, C, Q, or S induced chlorosis that was associated with reduced expression of chloroplast and photosynthesis related genes (CPRGs) and abnormal chloroplasts with fewer thylakoid membranes. These results suggest that the CP of the CMV pepo strain acquires virulence by amino acid mutations, which causes CPRG repression and chloroplast abnormalities.

  18. EFFECTS OF OUABAIN AND DIGOXIN ON THE GENE EXPRESSION OF SODIUM PUMP α-SUBUNIT ISOFORM IN AORTIC SMOOTH MUSCLE OF RATS

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Objective To compare the effects of ouabain and digoxin on both the systolic blood pressure and sodium pump α-subunit isoforms gene expression in the aortic smooth muscle of rats. Methods Normal SpragueDawley rats were injected with ouabain (20μg·kg-1 ·d-1 ,i. p),digoxin (32 μg·kg-1 ·d-1,i. p)and normal saline once a day, respectively, and indirect systolic blood pressure was recorded once a week. Six weeks later,all the rats were killed and sodium pump α1-,α2-,and α3-subunit mRNA levels were detected in the aortic smooth muscle with reverse transcription polymerase chain reaction(RT-PCR) method. Results The systolic blood pressure of rats infused with ouabain increased significantly at the end of week 6 [132. 6± 9. 0 mmHg (1 mmHg = 0. 133 kPa)vs 115. 7±8.2mmHg, P <0. 01] ,while no difference of blood pressure was found between digoxin group and NS group (P>0.05).The expression of sodium pump α-subunit isoforms in aortic smooth muscle was regulated by either ouabain or digoxin:both ouabain and digoxin increased α1- and α3-subunit expression, α2-subunit decreased in digoxin group but unchanged in ouabain group. Conclusion These results suggest that both ouabain and digoxin could regulate sodium pump α-subunit isoform expression, which might be related to the physiological roles of endogenous ouabain and might be responsible for the difference between the pharmacological and toxicological effects of ouabain and digoxin,including their effects on blood pressure.

  19. Absence of the A4 peptide in the G4 glycinin subunit of soybean cultivar Enrei is caused by a point mutation in the Gy4 gene

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    Yu Kangfu

    2005-01-01

    Full Text Available Functional properties of soy proteins for food are closely related to the composition of their storage protein subunits. Using base excision sequence scanning (BESS, we show that the absence of the A4 peptide in the G4 glycinin subunit of the soybean (Glycine max L. cultivar Enrei was caused by the same point mutation in the Gy4 gene as previously reported in the soybean cultivar Raiden. Although the genetic relationship between Raiden and Enrei is not known, the same point mutation in their Gy4 genes may indicate that they probably share a related origin. The application of BESS to identify single nucleotide polymorphisms (SNPs as co-dominant markers for marker-assisted selection (MAS of a recessive null allele is also discussed.

  20. Characterization of low-molecular-weight-glutenin subunit genes from the D-genome of Triticum aestivum, Aegilops crassa, Ae. cylindrica and Ae. tauschii

    NARCIS (Netherlands)

    Naghavi, M.R.; Ahmadi, S.; Shanejat-Boushehri, A.A.; Komaei, G.; Struik, P.C.

    2013-01-01

    Twenty low-molecular-weight-glutenin subunit (LMW-GS) gene sequences from the D-genome from Aegilops crassa (2n ¼ 4x ¼ 28), Ae. cylindrica (2n ¼ 4x ¼ 28), Ae. tauschii (2n ¼ 2x ¼ 14) and Triticum aestivum (2n ¼ 6x ¼ 42) were obtained using five sets of specific allele primer pairs. Only the sequence

  1. Cloning and sequence analysis of the gene encoding 19-kD subunit of Complex I from Dunaliella salina.

    Science.gov (United States)

    Liu, Yi; Qiao, Dai Rong; Zheng, Hong Bo; Dai, Xu Lan; Bai, Lin Han; Zeng, Jing; Cao, Yi

    2008-09-01

    NADH:ubiquinone oxidoreductase (complex I ) of the mitochondrial respiratory chain catalyzes the transfer of electrons from NADH to ubiquinone coupled to proton translocation across the membrane. The cDNA sequence of Dunaliella salina mitochondrial NADH: ubiquinone oxidoreductase 19-kD subunit contains a 682-bp ORF encoding a protein with an apparent molecular mass of 19 kD. The sequence has been submitted to the GenBank database under Accession No. EF566890 (cDNA sequences) and EF566891 (genomic sequence). The deduced amino-acid sequence is 74% identical to Chlamydomonas reinhardtii mitochondrial NADH:ubiquinone oxidoreductase 18-kD subunit. The 19-kD subunit mRNA expression was observed in oxygen deficiency, salt treatment, and rotenone treatment with lower levels. It demonstrate that the 19-kD subunit of Complex I from Dunaliella salina is regulated by these stresses.

  2. Genetic enhancement of memory and long-term potentiation but not CA1 long-term depression in NR2B transgenic rats.

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    Deheng Wang

    Full Text Available One major theory in learning and memory posits that the NR2B gene is a universal genetic factor that acts as rate-limiting molecule in controlling the optimal NMDA receptor's coincidence-detection property and subsequent learning and memory function across multiple animal species. If so, can memory function be enhanced via transgenic overexpression of NR2B in another species other than the previously reported mouse species? To examine these crucial issues, we generated transgenic rats in which NR2B is overexpressed in the cortex and hippocampus and investigated the role of NR2B gene in NMDA receptor-mediated synaptic plasticity and memory functions by combining electrophysiological technique with behavioral measurements. We found that overexpression of the NR2B subunit had no effect on CA1-LTD, but rather resulted in enhanced CA1-LTP and improved memory performances in novel object recognition test, spatial water maze, and delayed-to-nonmatch working memory test. Our slices recordings using NR2A- and NR2B-selective antagonists further demonstrate that the larger LTP in transgenic hippocampal slices was due to contribution from the increased NR2B-containing NMDARs. Therefore, our genetic experiments suggest that NR2B at CA1 synapses is not designated as a rate-limiting factor for the induction of long-term synaptic depression, but rather plays a crucial role in initiating the synaptic potentiation. Moreover, our studies provide strong evidence that the NR2B subunit represents a universal rate-limiting molecule for gating NMDA receptor's optimal coincidence-detection property and for enhancing memory function in adulthood across multiple mammalian species.

  3. Chromosomal location of genes for novel glutenin subunits and gliadins in wild emmer wheat (Triticum turgidum L. var. dicoccoides).

    Science.gov (United States)

    Xu, S S; Khan, K; Klindworth, D L; Faris, J D; Nygard, G

    2004-05-01

    The glutenin and gliadin proteins of wild emmer wheat, Triticum turgidum L. var. dicoccoides, have potential for improvement of durum wheat ( T. turgidum L. var. durum) quality. The objective of this study was to determine the chromosomes controlling the high molecular weight (HMW) glutenin subunits and gliadin proteins present in three T. turgidum var. dicoccoides accessions (Israel-A, PI-481521, and PI-478742), which were used as chromosome donors in Langdon durum- T. turgidum var. dicoccoides (LDN-DIC) chromosome substitution lines. The three T. turgidum var. dicoccoides accessions, their respective LDN-DIC substitution lines, and a number of controls with known HMW glutenin subunits were analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), urea/SDS-PAGE, and acid polyacrylamide gel electrophoresis (A-PAGE). The results revealed that all three T. turgidum var. dicoccoides accessions possess Glu-A1 alleles that are the same as or similar to those reported previously. However, each T. turgidum var. dicoccoides accession had a unique Glu-B1 allele. PI-478742 had an unusual 1Bx subunit, which had mobility slightly slower than the 1Ax subunit in 12% SDS-PAGE gels. The subunits controlled by chromosome 1B of PI-481521 were slightly faster in mobility than the subunits of the Glu-B1n allele, and the 1By subunit was identified as band 8. The 1B subunits of Israel-A had similar mobility to subunits 14 and 16. The new Glu-B1 alleles were designated as Glu-B1be in Israel-A, Glu-B1bf in PI-481521, and Glu-B1bg in PI-478742. Results from A-PAGE revealed that PI-481521, PI-478742, and Israel-A had eight, 12, and nine unique gliadin bands, respectively, that were assigned to specific chromosomes. The identified glutenin subunits and gliadin proteins in the LDN-DIC substitution lines provide the basis for evaluating their effects on end-use quality, and they are also useful biochemical markers for identifying specific chromosomes or chromosome

  4. Phylogenetic positions of two marine ciliates, Metanophrys similis and Pseudocohnilembus hargisi (Protozoa, Ciliophora, Scuticociliatia), inferred from complete small subunit rRNA gene sequences

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    The small subunit rRNA (SSrRNA) gene was sequenced for two marine scuticociliates Metanophrys similis and Pseudocohnilembus hargisi. The results show that this gene comprises 1763 and 1753 nucleotides in the two marine ciliates respectively.Metanophrys similis is phylogenetically closely related to the clade containing Mesanophrys carcini and Anophyroides haemophila, which branches basally to other species within the order Philasterida. Pseudocohnilembus hargisi groups with its congener, P. marinus, with strong bootstrap support. Paranophrys magna groups with the clade including Cohnilembus and Uronema, representing a sister clade to that containing the two Pseudocohnilembus species.

  5. Vanishing white matter disease: an Italian case with A638G mutation in exon 5 of EIF2B2 gene, an unusual early onset and a long course.

    Science.gov (United States)

    Sambati, Luisa; Agati, Raffaele; Bacci, Antonella; Bianchi, Silvia; Capellari, Sabina

    2013-07-01

    We report the clinical description of an Italian patient with c.638A>G mutation in exon 5 of EIF2B2 gene and a very slow progressive Vanishing White Matter disease phenotype. Infact, in relation to her causative mutation, our patient had an unusual early onset and long course. Furthermore, other than standard MRI examination and spectroscopy study, we report DWI and ADC maps and FA maps reconstruction from DTI in order to describe brain tissue degeneration in vanishing white matter disease.

  6. GRIN2B Mutations in West Syndrome and Intellectual Disability with Focal Epilepsy

    Science.gov (United States)

    Lemke, Johannes R; Hendrickx, Rik; Geider, Kirsten; Laube, Bodo; Schwake, Michael; Harvey, Robert J; James, Victoria M; Pepler, Alex; Steiner, Isabelle; Hörtnagel, Konstanze; Neidhardt, John; Ruf, Susanne; Wolff, Markus; Bartholdi, Deborah; Caraballo, Roberto; Platzer, Konrad; Suls, Arvid; De Jonghe, Peter; Biskup, Saskia; Weckhuysen, Sarah

    2014-01-01

    Objective To identify novel epilepsy genes using a panel approach and describe the functional consequences of mutations. Methods Using a panel approach, we screened 357 patients comprising a vast spectrum of epileptic disorders for defects in genes known to contribute to epilepsy and/or intellectual disability (ID). After detection of mutations in a novel epilepsy gene, we investigated functional effects in Xenopus laevis oocytes and screened a follow-up cohort. Results We revealed de novo mutations in GRIN2B encoding the NR2B subunit of the N-methyl-D-aspartate (NMDA) receptor in 2 individuals with West syndrome and severe developmental delay as well as 1 individual with ID and focal epilepsy. The patient with ID and focal epilepsy had a missense mutation in the extracellular glutamate-binding domain (p.Arg540His), whereas both West syndrome patients carried missense mutations within the NR2B ion channel-forming re-entrant loop (p.Asn615Ile, p.Val618Gly). Subsequent screening of 47 patients with unexplained infantile spasms did not reveal additional de novo mutations, but detected a carrier of a novel inherited GRIN2B splice site variant in close proximity (c.2011-5_2011-4delTC). Mutations p.Asn615Ile and p.Val618Gly cause a significantly reduced Mg2+ block and higher Ca2+ permeability, leading to a dramatically increased Ca2+ influx, whereas p.Arg540His caused less severe disturbance of channel function, corresponding to the milder patient phenotype. Interpretation We identified GRIN2B gain-of-function mutations as a cause of West syndrome with severe developmental delay as well as of ID with childhood onset focal epilepsy. Severely disturbed channel function corresponded to severe clinical phenotypes, underlining the important role of facilitated NMDA receptor signaling in epileptogenesis. PMID:24272827

  7. The acquired radioresistance in HeLa cells under conditions mimicking hypoxia was attenuated by a decreased expression of HIF subunit genes induced by RNA interference

    Energy Technology Data Exchange (ETDEWEB)

    Doi, Nobutaka [Department of Radiological Sciences, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194 (Japan); New Products Research & Development, Gene Engineering Division, NIPPON GENE Co., Ltd. (Japan); Ogawa, Ryohei, E-mail: ogawa@med.u-toyama.ac.jp [Department of Radiological Sciences, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194 (Japan); Cui, Zheng-Guo [Department of Public Health, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama (Japan); Morii, Akihiro; Watanabe, Akihiko [Department of Urology, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama (Japan); Kanayama, Shinji; Yoneda, Yuko [New Products Research & Development, Gene Engineering Division, NIPPON GENE Co., Ltd. (Japan); Kondo, Takashi [Department of Radiological Sciences, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194 (Japan)

    2015-05-01

    The cancer cells residing in the hypoxic layer are resistant to radiation and these are ones responsible for cancer recurrence after radiation therapy. One of the reasons why hypoxic cancer cells acquire radioresistance may be attributable to changes in the gene expression profile by the activation of hypoxia inducible factors (HIFs). However, the details underlying this process remain unknown. In this study, we investigated the effects of knockdown of HIF subunit genes to elucidate how HIF subunit genes may be involved in the radioresistance acquired by HeLa cells following exposure to a hypoxia mimic. Interestingly, HIF-1α and HIF-2α seemed mutually complementary for each other when either of them was suppressed. We thus suppressed the expression of both genes simultaneously. To do this, we developed a short hairpin RNA (shRNA) targeting a high homology region between HIF-1α and HIF-2α. It was shown that the expression of the shRNA effectively suppressed the acquisition of radioresistance following the hypoxia mimic. Moreover, it was confirmed that suppression of both subunits resulted in the downregulation of stem cell markers and the suppression of spheroid formation during the hypoxia mimicking-conditions. This shRNA-mediated knockdown method targeting a common region shared by a family of genes may offer a new candidate cancer treatment. - Highlights: • Incubation with CoCl{sub 2} confers radioresistance to HeLa cells. • Both HIF-1α and HIF-2α are involved in the acquisition of radioresistance. • An shRNA to a homology region of HIF-1α and HIF-2α suppressed the radioresistance. • The shRNA decreased cells with stem cell markers and a stem cell phenotype.

  8. Neuron-specific specificity protein 4 bigenomically regulates the transcription of all mitochondria- and nucleus-encoded cytochrome c oxidase subunit genes in neurons.

    Science.gov (United States)

    Johar, Kaid; Priya, Anusha; Dhar, Shilpa; Liu, Qiuli; Wong-Riley, Margaret T T

    2013-11-01

    Neurons are highly dependent on oxidative metabolism for their energy supply, and cytochrome c oxidase (COX) is a key energy-generating enzyme in the mitochondria. A unique feature of COX is that it is one of only four proteins in mammalian cells that are bigenomically regulated. Of its thirteen subunits, three are encoded in the mitochondrial genome and ten are nuclear-encoded on nine different chromosomes. The mechanism of regulating this multisubunit, bigenomic enzyme poses a distinct challenge. In recent years, we found that nuclear respiratory factors 1 and 2 (NRF-1 and NRF-2) mediate such bigenomic coordination. The latest candidate is the specificity factor (Sp) family of proteins. In N2a cells, we found that Sp1 regulates all 13 COX subunits. However, we discovered recently that in primary neurons, it is Sp4 and not Sp1 that regulates some of the key glutamatergic receptor subunit genes. The question naturally arises as to the role of Sp4 in regulating COX in primary neurons. The present study utilized multiple approaches, including chromatin immunoprecipitation, promoter mutational analysis, knockdown and over-expression of Sp4, as well as functional assays to document that Sp4 indeed functionally regulate all 13 subunits of COX as well as mitochondrial transcription factors A and B. The present study discovered that among the specificity family of transcription factors, it is the less known neuron-specific Sp4 that regulates the expression of all 13 subunits of mitochondrial cytochrome c oxidase (COX) enzyme in primary neurons. Sp4 also regulates the three mitochondrial transcription factors (TFAM, TFB1M, and TFB2M) and a COX assembly protein SURF-1 in primary neurons.

  9. Novel insights into the composition, variation, organization, and expression of the low-molecular-weight glutenin subunit gene family in common wheat.

    Science.gov (United States)

    Zhang, Xiaofei; Liu, Dongcheng; Zhang, Jianghua; Jiang, Wei; Luo, Guangbin; Yang, Wenlong; Sun, Jiazhu; Tong, Yiping; Cui, Dangqun; Zhang, Aimin

    2013-04-01

    Low-molecular-weight glutenin subunits (LMW-GS), encoded by a complex multigene family, play an important role in the processing quality of wheat flour. Although members of this gene family have been identified in several wheat varieties, the allelic variation and composition of LMW-GS genes in common wheat are not well understood. In the present study, using the LMW-GS gene molecular marker system and the full-length gene cloning method, a comprehensive molecular analysis of LMW-GS genes was conducted in a representative population, the micro-core collections (MCC) of Chinese wheat germplasm. Generally, >15 LMW-GS genes were identified from individual MCC accessions, of which 4-6 were located at the Glu-A3 locus, 3-5 at the Glu-B3 locus, and eight at the Glu-D3 locus. LMW-GS genes at the Glu-A3 locus showed the highest allelic diversity, followed by the Glu-B3 genes, while the Glu-D3 genes were extremely conserved among MCC accessions. Expression and sequence analysis showed that 9-13 active LMW-GS genes were present in each accession. Sequence identity analysis showed that all i-type genes present at the Glu-A3 locus formed a single group, the s-type genes located at Glu-B3 and Glu-D3 loci comprised a unique group, while high-diversity m-type genes were classified into four groups and detected in all Glu-3 loci. These results contribute to the functional analysis of LMW-GS genes and facilitate improvement of bread-making quality by wheat molecular breeding programmes.

  10. Determination of the relative expression levels of rubisco small subunit genes in Arabidopsis by rapid amplification of cDNA ends.

    Science.gov (United States)

    Yoon, M; Putterill, J J; Ross, G S; Laing, W A

    2001-04-15

    Multigene families are common in higher organisms. However, due to the close similarities between members, it is often difficult to assess the individual contribution of each gene to the overall expression of the family. In Arabidopsis thaliana, there are four genes encoding the small subunits (SSU) of ribulose-1.5-bisphosphate carboxylase oxygenase (rubisco) whose nucleotide sequences are up to 98.4% identical. In order to overcome the technical limitations associated with gene-specific probes (or primers) commonly used in existing methods, we developed a new gene expression assay based on the RACE (rapid amplification of cDNA ends) technique with a single pair of primers. With this RACE gene expression assay, we were able to determine the relative transcript levels between four Arabidopsis SSU genes. We found that the relative SSU gene expression differed significantly between plants grown at different temperatures. Our observation raises the possibility that an adaptation of rubisco to the environment may be achieved through the specific synthesis of the SSU proteins, which is determined by the relative expression levels between the SSU genes.

  11. Identification of Botrytis cinerea genes up-regulated during infection and controlled by the Galpha subunit BCG1 using suppression subtractive hybridization (SSH).

    Science.gov (United States)

    Schulze Gronover, Christian; Schorn, Corinna; Tudzynski, Bettina

    2004-05-01

    The Galpha subunit BCG1 plays an important role during the infection of host plants by Botrytis cinerea. Delta bcg1 mutants are able to conidiate, penetrate host leaves, and produce small primary lesions. However, in contrast to the wild type, the mutants completely stop invasion of plant tissue at this stage; secondary lesions have never been observed. Suppression subtractive hybridization (SSH) was used to identify fungal genes whose expression on the host plant is specifically affected in bcg1 mutants. Among the 22 differentially expressed genes, we found those which were predicted to encode proteases, enzymes involved in secondary metabolism, and others encoding cell wall-degrading enzymes. All these genes are highly expressed during infection in the wild type but not in the mutant. However, the genes are expressed in both the wild type and the mutant under certain conditions in vitro. Most of the BCG1-controlled genes are still expressed in adenylate cyclase (bac) mutants in planta, suggesting that BCG1 is involved in at least one additional signaling cascade in addition to the cAMP-depending pathway. In a second SSH approach, 1,500 clones were screened for those that are specifically induced by the wild type during the infection of bean leaves. Of the 22 BCG1-controlled genes, 11 also were found in the in planta SSH library. Therefore, SSH technology can be successfully applied to identify target genes of signaling pathways and differentially expressed genes in planta.

  12. The tryptophan photoproduct 6-formylindolo[3,2-b]carbazole (FICZ) binds multiple AHRs and induces multiple CYP1 genes via AHR2 in zebrafish

    DEFF Research Database (Denmark)

    Jönsson, Maria E.; Franks, Diana G.; Woodin, Bruce R.;

    2009-01-01

    to FICZ is evolutionarily conserved in vertebrates by measuring FICZ binding to two zebrafish AHRs (AHR1B and AHR2) and its ability to induce zebrafish CYP1 genes (CYP1A, CYP1B1, CYP1C1, CYP1C2, and CYP1D1) in vivo. Exposure of zebrafish embryos (48 h-post-fertilization; hpf) to 10 nM FICZ for 6 h caused...... strong induction of CYP1A mRNA and a statistically significant but modest induction of CYP1B1 and CYP1C1. Neither CYP1C2 nor CYP1D1 expression was induced by FICZ under the conditions of dose, time or developmental stage examined here. CYP1A induction was significantly greater after 6 h than after 12 h...... of exposure to FICZ, suggesting a rapid degradation of inducer. The 6-h EC50 values for induction of CYP1A and CYP1B1 by FICZ were 0.6 and 0.5 nM compared to 72-h EC50 values of 2.3 and 2.7 nM for PCB126, indicating that in zebrafish embryos FICZ is a more potent inducer than PCB126. FICZ at 10 nM was able...

  13. Elevated breast cancer risk in irradiated BALB/c mice associates with unique functional polymorphism of the Prkdc (DNA-dependent protein kinase catalytic subunit) gene

    Science.gov (United States)

    Yu, Y.; Okayasu, R.; Weil, M. M.; Silver, A.; McCarthy, M.; Zabriskie, R.; Long, S.; Cox, R.; Ullrich, R. L.

    2001-01-01

    Female BALB/c mice are unusually radiosensitive and more susceptible than C57BL/6 and other tested inbred mice to ionizing radiation (IR)-induced mammary tumors. This breast cancer susceptibility is correlated with elevated susceptibility for mammary cell transformation and genomic instability following irradiation. In this study, we report the identification of two BALB/c strain-specific polymorphisms in the coding region of Prkdc, the gene encoding the DNA-dependent protein kinase catalytic subunit, which is known to be involved in DNA double-stranded break repair and post-IR signal transduction. First, we identified an A --> G transition at base 11530 resulting in a Met --> Val conversion at codon 3844 (M3844V) in the phosphatidylinositol 3-kinase domain upstream of the scid mutation (Y4046X). Second, we identified a C --> T transition at base 6418 resulting in an Arg --> Cys conversion at codon 2140 (R2140C) downstream of the putative leucine zipper domain. This unique PrkdcBALB variant gene is shown to be associated with decreased DNA-dependent protein kinase catalytic subunit activity and with increased susceptibility to IR-induced genomic instability in primary mammary epithelial cells. The data provide the first evidence that naturally arising allelic variation in a mouse DNA damage response gene may associate with IR response and breast cancer risk.

  14. Nuclear-cytoplasmic conflict in pea (Pisum sativum L.) is associated with nuclear and plastidic candidate genes encoding acetyl-CoA carboxylase subunits.

    Science.gov (United States)

    Bogdanova, Vera S; Zaytseva, Olga O; Mglinets, Anatoliy V; Shatskaya, Natalia V; Kosterin, Oleg E; Vasiliev, Gennadiy V

    2015-01-01

    In crosses of wild and cultivated peas (Pisum sativum L.), nuclear-cytoplasmic incompatibility frequently occurs manifested as decreased pollen fertility, male gametophyte lethality, sporophyte lethality. High-throughput sequencing of plastid genomes of one cultivated and four wild pea accessions differing in cross-compatibility was performed. Candidate genes for involvement in the nuclear-plastid conflict were searched in the reconstructed plastid genomes. In the annotated Medicago truncatula genome, nuclear candidate genes were searched in the portion syntenic to the pea chromosome region known to harbor a locus involved in the conflict. In the plastid genomes, a substantial variability of the accD locus represented by nucleotide substitutions and indels was found to correspond to the pattern of cross-compatibility among the accessions analyzed. Amino acid substitutions in the polypeptides encoded by the alleles of a nuclear locus, designated as Bccp3, with a complementary function to accD, fitted the compatibility pattern. The accD locus in the plastid genome encoding beta subunit of the carboxyltransferase of acetyl-coA carboxylase and the nuclear locus Bccp3 encoding biotin carboxyl carrier protein of the same multi-subunit enzyme were nominated as candidate genes for main contribution to nuclear-cytoplasmic incompatibility in peas. Existence of another nuclear locus involved in the accD-mediated conflict is hypothesized.

  15. Switch in glutamate receptor subunit gene expression in CA1 subfield of hippocampus following global ischemia in rats.

    OpenAIRE

    Pellegrini-Giampietro, D E; Zukin, R.S.; Bennett, M V; Cho, S; Pulsinelli, W. A.

    1992-01-01

    Severe, transient global ischemia of the brain induces delayed damage to specific neuronal populations. Sustained Ca2+ influx through glutamate receptor channels is thought to play a critical role in postischemic cell death. Although most kainate-type glutamate receptors are Ca(2+)-impermeable, Ca(2+)-permeable kainate receptors have been reported in specific kinds of neurons and glia. Recombinant receptors assembled from GluR1 and/or GluR3 subunits in exogenous expression systems are permeab...

  16. Association study of common polymorphisms in MSRA, TFAP2B, MC4R, NRXN3, PPARGC1A, TMEM18, SEC16B, HOXB5 and OLFM4 genes with obesity-related traits among Portuguese children.

    Science.gov (United States)

    Albuquerque, David; Nóbrega, Clévio; Rodríguez-López, Raquel; Manco, Licínio

    2014-06-01

    At least 52 genetic loci were associated with obesity-related traits. However, little is known about the genetic basis of obesity among children. This study aims to test whether 10 polymorphisms in obesity-related genes methionine sulfoxide reductase A (MSRA), transcription factor AP-2 beta (TFAP2B), melanocortin 4 receptor (MC4R), neurexin 3 (NRXN3), peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PPARGC1A), transmembrane protein 18 (TMEM18), homolog of S. cerevisiae Sec16 (SEC16B), homeobox B5 (HOXB5) and olfactomedin 4 (OLFM4) are associated with the risk of obesity in Portuguese children. A total of 730 children aging from 6 to 12 years old, recruited randomly from public schools in Portugal, were analysed. Anthropometric measurements were obtained and children were classified into three phenotypic groups, normal weight (n=256), overweight (n=320) and obese (n=154), according to the International Obesity Task Force cutoffs. Polymorphisms were genotyped by allelic discrimination TaqMan assays. The MC4R rs12970134 polymorphism was nominally associated with body mass index (BMI) (P=0.035), BMI Z-score (P=0.043) and waist circumference (P=0.020), and borderline associated with weight (P=0.053). Near nominal associations were also found for the PPARGC1A rs8192678 polymorphism with weight (P=0.061), and for the MSRA rs545854 polymorphism with BMI (P=0.055) and BMI Z-score (P=0.056). Furthermore, logistic regression showed that MC4R rs12970134 and TFAP2B rs987237 were nominally, respectively, associated (P=0.029) and borderline associated (P=0.056) with the obese phenotype. This study highlighted the possible association of MC4R, PPARGC1A, MSRA and TFAP2B polymorphisms with several obesity-related traits in a sample of Portuguese children. The two significant associated TFAP2B rs987237 and MC4R rs12970134 polymorphisms showed an opposite direction of effect to that in the original reports.

  17. SWR1 Chromatin-Remodeling Complex Subunits and H2A.Z Have Non-overlapping Functions in Immunity and Gene Regulation in Arabidopsis.

    Science.gov (United States)

    Berriri, Souha; Gangappa, Sreeramaiah N; Kumar, S Vinod

    2016-07-06

    Incorporation of the histone variant H2A.Z into nucleosomes by the SWR1 chromatin remodeling complex is a critical step in eukaryotic gene regulation. In Arabidopsis, SWR1c and H2A.Z have been shown to control gene expression underlying development and environmental responses. Although they have been implicated in defense, the specific roles of the complex subunits and H2A.Z in immunity are not well understood. In this study, we analyzed the roles of the SWR1c subunits, PHOTOPERIOD-INDEPENDENT EARLY FLOWERING1 (PIE1), ACTIN-RELATED PROTEIN6 (ARP6), and SWR1 COMPLEX 6 (SWC6), as well as H2A.Z, in defense and gene regulation. We found that SWR1c components play different roles in resistance to different pathogens. Loss of PIE1 and SWC6 function as well as depletion of H2A.Z led to reduced basal resistance, while loss of ARP6 fucntion resulted in enhanced resistance. We found that mutations in PIE1 and SWC6 resulted in impaired effector-triggered immunity. Mutation in SWR1c components and H2A.Z also resulted in compromised jasmonic acid/ethylene-mediated immunity. Genome-wide expression analyses similarly reveal distinct roles for H2A.Z and SWR1c components in gene regulation, and suggest a potential role for PIE1 in the regulation of the cross talk between defense signaling pathways. Our data show that although they are part of the same complex, Arabidopsis SWR1c components could have non-redundant functions in plant immunity and gene regulation.

  18. Relationship between reduced nicotinamide adenine dinucleotide phosphate oxidase subunit p22phox gene polymorphism and obstructive sleep apnea-hypopnea syndrome in the Chinese Han population

    Institute of Scientific and Technical Information of China (English)

    LIU Hui-guo; LIU Kui; ZHOU Yan-ning; XU Yong-jian

    2009-01-01

    Background Increased production of reactive oxygen species (ROS) is thought to play a major role in the pathogenesis of obstructive sleep apnea-hypopnea syndrome (OSAHS). The reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase complex is an important source of ROS. The p22phox subunit is polymorphic with a C242T variant that changes histidine-72 for a tyrosine in the potential heme binding site. This study aimed to investigate the relationship between NADPH oxidase subunit p22phox gene polymorphism and OSAHS. Methods The genotypes of p22phox polymorphism were determined by polymerase chain reaction-restriction fragment length polymorphisms (PCR-RFLP) assay in 176 unrelated subjects of the Han population in southern region of China (including 107 OSAHS subjects and 69 non-OSAHS subjects), while the plasma concentration of superoxide dismutase (SOD) was detected in the two groups, and p22phox mRNA expression in peripheral blood mononuclear cell (PBMC) was determined with reverse transcription polymerase chain reaction (RT-PCR).Results The phagocyte NADPH oxidase subunit p22phox mRNA expression was significantly increased in the OSAHS group than that in the non-OSAHS group (P<0.01). Compared with the non-OSAHS control group ((85.31±9.23) U/ml), the levels of SOD were lower in patients with OSAHS ((59.65±11.61) U/ml (P<0.01). There were significant differences in genotypes distribution in p22phox polymorphism between the two groups (P=0.02). Compared with the non-OSAHS control group, the OSAHS group had a significantly higher T allele frequency in p22phox polymorphism (P=0.03). There were independent effects of p22phox polymorphism on body mass index (BMI), neck circumference (NC), waist-to-hip ratio (WHR) in the OSAHS group, and the carriers of the T allele of p22phox polymorphism had greater NC, WHR, systolic blood pressure (SBP), diastolic blood pressure (DBP) and apnea-hypopnea index (AHI) (P <0.05), but the carriers of the T allele had lower SOD

  19. Isolation and characterization of BetaM protein encoded by ATP1B4 - a unique member of the Na,K-ATPase {beta}-subunit gene family

    Energy Technology Data Exchange (ETDEWEB)

    Pestov, Nikolay B. [Department of Physiology and Pharmacology, University of Toledo College of Medicine, 3000 Arlington Ave., Toledo, OH 43614 (United States); Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Moscow 117997 (Russian Federation); Zhao, Hao [Department of Physiology and Pharmacology, University of Toledo College of Medicine, 3000 Arlington Ave., Toledo, OH 43614 (United States); Basrur, Venkatesha [Department of Pathology, University of Michigan Medical School, Ann Arbor, MI 48109 (United States); Modyanov, Nikolai N., E-mail: nikolai.modyanov@utoledo.edu [Department of Physiology and Pharmacology, University of Toledo College of Medicine, 3000 Arlington Ave., Toledo, OH 43614 (United States)

    2011-09-09

    Highlights: {yields} Structural properties of BetaM and Na,K-ATPase {beta}-subunits are sharply different. {yields} BetaM protein is concentrated in nuclear membrane of skeletal myocytes. {yields} BetaM does not associate with a Na,K-ATPase {alpha}-subunit in skeletal muscle. {yields} Polypeptide chain of the native BetaM is highly sensitive to endogenous proteases. {yields} BetaM in neonatal muscle is a product of alternative splice mRNA variant B. -- Abstract: ATP1B4 genes represent a rare instance of the orthologous gene co-option that radically changed functions of encoded BetaM proteins during vertebrate evolution. In lower vertebrates, this protein is a {beta}-subunit of Na,K-ATPase located in the cell membrane. In placental mammals, BetaM completely lost its ancestral role and through acquisition of two extended Glu-rich clusters into the N-terminal domain gained entirely new properties as a muscle-specific protein of the inner nuclear membrane possessing the ability to regulate gene expression. Strict temporal regulation of BetaM expression, which is the highest in late fetal and early postnatal myocytes, indicates that it plays an essential role in perinatal development. Here we report the first structural characterization of the native eutherian BetaM protein. It should be noted that, in contrast to structurally related Na,K-ATPase {beta}-subunits, the polypeptide chain of BetaM is highly sensitive to endogenous proteases that greatly complicated its isolation. Nevertheless, using a complex of protease inhibitors, a sample of authentic BetaM was isolated from pig neonatal skeletal muscle by a combination of ion-exchange and lectin-affinity chromatography followed by SDS-PAGE. Results of the analysis of the BetaM tryptic digest using MALDI-TOF and ESI-MS/MS mass spectrometry have demonstrated that native BetaM in neonatal skeletal muscle is a product of alternative splice mRNA variant B and comprised of 351 amino acid residues. Isolated BetaM protein was

  20. A genetic suppressor of two dominant temperature-sensitive lethal proteasome mutants of Drosophila melanogaster is itself a mutated proteasome subunit gene.

    Science.gov (United States)

    Neuburger, Peter J; Saville, Kenneth J; Zeng, Jue; Smyth, Kerrie-Ann; Belote, John M

    2006-07-01

    Two dominant temperature-sensitive (DTS) lethal mutants of Drosophila melanogaster are Pros26(1) and Prosbeta2(1), previously known as DTS5 and DTS7. Heterozygotes for either mutant die as pupae when raised at 29 degrees , but are normally viable and fertile at 25 degrees . Previous studies have identified these as missense mutations in the genes encoding the beta6 and beta2 subunits of the 20S proteasome, respectively. In an effort to isolate additional proteasome-related mutants a screen for dominant suppressors of Pros26(1) was carried out, resulting in the identification of Pros25(SuDTS) [originally called Su(DTS)], a missense mutation in the gene encoding the 20S proteasome alpha2 subunit. Pros25(SuDTS) acts in a dominant manner to rescue both Pros26(1) and Prosbeta2(1) from their DTS lethal phenotypes. Using an in vivo protein degradation assay it was shown that this suppression occurs by counteracting the dominant-negative effect of the DTS mutant on proteasome activity. Pros25(SuDTS) is a recessive polyphasic lethal at ambient temperatures. The effects of these mutants on larval neuroblast mitosis were also examined. While Prosbeta2(1) shows a modest increase in the number of defective mitotic figures, there were no defects seen with the other two mutants, other than slightly reduced mitotic indexes.

  1. SRC Inhibition Reduces NR2B Surface Expression and Synaptic Plasticity in the Amygdala

    Science.gov (United States)

    Sinai, Laleh; Duffy, Steven; Roder, John C.

    2010-01-01

    The Src protein tyrosine kinase plays a central role in the regulation of N-methyl-d-aspartate receptor (NMDAR) activity by regulating NMDAR subunit 2B (NR2B) surface expression. In the amygdala, NMDA-dependent synaptic plasticity resulting from convergent somatosensory and auditory inputs contributes to emotional memory; however, the role of Src…

  2. Disease-associated mutations in the HSPD1 gene encoding the large subunit of the mitochondrial HSP60/HSP10 chaperonin complex

    Directory of Open Access Journals (Sweden)

    Peter Bross

    2016-08-01

    Full Text Available Heat shock protein 60 (HSP60 forms together with heat shock protein 10 (HSP10 double-barrel chaperonin complexes that are essential for folding to the native state of proteins in the mitochondrial matrix space. Two extremely rare monogenic disorders have been described that are caused by missense mutations in the HSPD1 gene that encodes the HSP60 subunit of the HSP60/HSP10 chaperonin complex. Investigations of the molecular mechanisms underlying these disorders have revealed that different degrees of reduced HSP60 function produce distinct neurological phenotypes. While mutations with deleterious or strong dominant negative effects are not compatible with life, HSPD1 gene variations found in the human population impair HSP60 function and depending on the mechanism and degree of HSP60 dys- and malfunction cause different phenotypes. We here summarize the knowledge on the effects of disturbances of the function of the HSP60/HSP10 chaperonin complex by disease-associated mutations.

  3. Characterization and variation of a human inwardly-rectifying-K-channel gene (KCNJ6): a putative ATP-sensitive K-channel subunit.

    Science.gov (United States)

    Sakura, H; Bond, C; Warren-Perry, M; Horsley, S; Kearney, L; Tucker, S; Adelman, J; Turner, R; Ashcroft, F M

    1995-06-26

    The ATP-sensitive K-channel plays a central role in insulin release from pancreatic beta-cells. We report here the cloning of the gene (KCNJ6) encoding a putative subunit of a human ATP-sensitive K-channel expressed in brain and beta-cells, and characterisation of its exon-intron structure. Screening of a somatic cell mapping panel and fluorescent in situ hybridization place the gene on chromosome 21 (21q22.1-22.2). Analysis of single-stranded conformational polymorphisms revealed the presence of two silent polymorphisms (Pro-149: CCG-CCA and Asp-328: GAC-GAT) with similar frequencies in normal and non-insulin-dependent diabetic patients.

  4. Loss-of-function mutations of retromer large subunit genes suppress the phenotype of an Arabidopsis zig mutant that lacks Qb-SNARE VTI11.

    Science.gov (United States)

    Hashiguchi, Yasuko; Niihama, Mitsuru; Takahashi, Tetsuya; Saito, Chieko; Nakano, Akihiko; Tasaka, Masao; Morita, Miyo Terao

    2010-01-01

    Arabidopsis thaliana zigzag (zig) is a loss-of-function mutant of Qb-SNARE VTI11, which is involved in membrane trafficking between the trans-Golgi network and the vacuole. zig-1 exhibits abnormalities in shoot gravitropism and morphology. Here, we report that loss-of-function mutants of the retromer large subunit partially suppress the zig-1 phenotype. Moreover, we demonstrate that three paralogous VPS35 genes of Arabidopsis have partially overlapping but distinct genetic functions with respect to zig-1 suppression. Tissue-specific complementation experiments using an endodermis-specific SCR promoter show that expression of VPS35B or VPS35C cannot complement the function of VPS35A. The data suggest the existence of functionally specialized paralogous VPS35 genes that nevertheless share common functions.

  5. Loss-of-Function Mutations of Retromer Large Subunit Genes Suppress the Phenotype of an Arabidopsis zig Mutant That Lacks Qb-SNARE VTI11[C][W

    Science.gov (United States)

    Hashiguchi, Yasuko; Niihama, Mitsuru; Takahashi, Tetsuya; Saito, Chieko; Nakano, Akihiko; Tasaka, Masao; Morita, Miyo Terao

    2010-01-01

    Arabidopsis thaliana zigzag (zig) is a loss-of-function mutant of Qb-SNARE VTI11, which is involved in membrane trafficking between the trans-Golgi network and the vacuole. zig-1 exhibits abnormalities in shoot gravitropism and morphology. Here, we report that loss-of-function mutants of the retromer large subunit partially suppress the zig-1 phenotype. Moreover, we demonstrate that three paralogous VPS35 genes of Arabidopsis have partially overlapping but distinct genetic functions with respect to zig-1 suppression. Tissue-specific complementation experiments using an endodermis-specific SCR promoter show that expression of VPS35B or VPS35C cannot complement the function of VPS35A. The data suggest the existence of functionally specialized paralogous VPS35 genes that nevertheless share common functions. PMID:20086190

  6. Association of polymorphisms in calpain 1, (mu/I) large subunit, calpastatin, and cathepsin D genes with meat quality traits in double-muscled Piemontese cattle.

    Science.gov (United States)

    Ribeca, Cinzia; Bonfatti, Valentina; Cecchinato, Alessio; Albera, Andrea; Maretto, Fabio; Gallo, Luigi; Carnier, Paolo

    2013-04-01

    Five single-nucleotide polymorphisms (SNPs) located in the calpain 1, (mu/I) large subunit (CAPN1), calpastatin (CAST), and cathepsin D (CTSD) genes were analyzed in a large sample of Piemontese cattle. The aim of this study was to evaluate allele and genotype frequencies of these SNPs and to investigate associations of CAPN1, CAST, and CTSD gene variants with meat quality traits. Minor allele frequencies ranged from 30 to 48%. The presence of the A allele at CAPN530 increased yellowness and drip loss. The CAST282 G allele was associated with an increased drip loss compared to the C allele, and the CAST2959 A allele decreased redness compared to the G allele.

  7. Neurotransmitter release: vacuolar ATPase V0 sector c-subunits in possible gene or cell therapies for Parkinson's, Alzheimer's, and psychiatric diseases.

    Science.gov (United States)

    Higashida, Haruhiro; Yokoyama, Shigeru; Tsuji, Chiharu; Muramatsu, Shin-Ichi

    2017-01-01

    We overview the 16-kDa proteolipid mediatophore, the transmembrane c-subunit of the V0 sector of the vacuolar proton ATPase (ATP6V0C) that was shown to mediate the secretion of acetylcholine. Acetylcholine, serotonin, and dopamine (DA) are released from cell soma and/or dendrites if ATP6V0C is expressed in cultured cells. Adeno-associated viral vector-mediated gene transfer of ATP6V0C into the caudate putamen enhanced the depolarization-induced overflow of endogenous DA in Parkinson-model mice. Motor impairment was ameliorated in hemiparkinsonian model mice when ATP6V0C was expressed with DA-synthesizing enzymes. The review discusses application in the future as a potential tool for gene therapy, cell transplantation therapy, and inducible pluripotent stem cell therapy in neurological diseases, from the view point of recent findings regarding vacuolar ATPase.

  8. Cloning and sequencing of the genes coding for the A and B subunits of vacuolar-type Na(+)-ATPase from Enterococcus hirae. Coexistence of vacuolar- and F0F1-type ATPases in one bacterial cell.

    Science.gov (United States)

    Takase, K; Yamato, I; Kakinuma, Y

    1993-06-05

    The eubacterium Enterococcus hirae ATCC 9790 possesses a H(+)-translocating ATPase, and the deduced amino acid sequences of the genes coding for this enzyme have indicated that it is a typical F0F1-type ATPase (Shibata, C., Ehara, T., Tomura, K., Igarashi, K., and Kobayashi, H. (1992) J. Bacteriol. 174, 6117-6124). We cloned the ntpA and ntpB genes coding for the A and B subunits, respectively, of Na(+)-translocating ATPase from the same bacterium, and the full amino acid sequences of the two subunits were deduced from the nucleotide sequence. The A (593 amino acid residues) and B (458 amino acid residues) subunits were highly homologous (48-60% identical) to the A (large or alpha) and the B (small or beta) subunits, respectively, of vacuolar-type H(+)-ATPases which have been found in eukaryotic endomembrane systems (Neurospora crassa, Saccharomyces cerevisiae, Arabidopsis thaliana, and carrot) and archaebacterial cell membranes (Sulfolobus acidocaldarius and Methanosarcina barkeri). The A and B subunits of Na(+)-ATPase showed about 23-28% identities with the beta and alpha subunits of E. hirae F1-ATPase and of Escherichia coli F1-ATPase, respectively. These results indicate that E. hirae Na(+)-ATPase belongs to the vacuolar-type ATPase. This is the first demonstration that both genes for V- and F-type ATPases are functionally expressed in one bacterial cell.

  9. INCURVATA2 Encodes the Catalytic Subunit of DNA Polymerase α and Interacts with Genes Involved in Chromatin-Mediated Cellular Memory in Arabidopsis thaliana

    Science.gov (United States)

    Barrero, José María; González-Bayón, Rebeca; del Pozo, Juan Carlos; Ponce, María Rosa; Micol, José Luis

    2007-01-01

    Cell type–specific gene expression patterns are maintained by the stable inheritance of transcriptional states through mitosis, requiring the action of multiprotein complexes that remodel chromatin structure. Genetic and molecular interactions between chromatin remodeling factors and components of the DNA replication machinery have been identified in Schizosaccharomyces pombe, indicating that some epigenetic marks are replicated simultaneously to DNA with the participation of the DNA replication complexes. This model of epigenetic inheritance might be extended to the plant kingdom, as we report here with the positional cloning and characterization of INCURVATA2 (ICU2), which encodes the putative catalytic subunit of the DNA polymerase α of Arabidopsis thaliana. The strong icu2-2 and icu2-3 insertional alleles caused fully penetrant zygotic lethality when homozygous and incompletely penetrant gametophytic lethality, probably because of loss of DNA polymerase activity. The weak icu2-1 allele carried a point mutation and caused early flowering, leaf incurvature, and homeotic transformations of sepals into carpels and of petals into stamens. Further genetic analyses indicated that ICU2 interacts with TERMINAL FLOWER2, the ortholog of HETEROCHROMATIN PROTEIN1 of animals and yeasts, and with the Polycomb group (PcG) gene CURLY LEAF. Another PcG gene, EMBRYONIC FLOWER2, was found to be epistatic to ICU2. Quantitative RT-PCR analyses indicated that a number of regulatory genes were derepressed in the icu2-1 mutant, including genes associated with flowering time, floral meristem, and floral organ identity. PMID:17873092

  10. INCURVATA2 encodes the catalytic subunit of DNA Polymerase alpha and interacts with genes involved in chromatin-mediated cellular memory in Arabidopsis thaliana.

    Science.gov (United States)

    Barrero, José María; González-Bayón, Rebeca; del Pozo, Juan Carlos; Ponce, María Rosa; Micol, José Luis

    2007-09-01

    Cell type-specific gene expression patterns are maintained by the stable inheritance of transcriptional states through mitosis, requiring the action of multiprotein complexes that remodel chromatin structure. Genetic and molecular interactions between chromatin remodeling factors and components of the DNA replication machinery have been identified in Schizosaccharomyces pombe, indicating that some epigenetic marks are replicated simultaneously to DNA with the participation of the DNA replication complexes. This model of epigenetic inheritance might be extended to the plant kingdom, as we report here with the positional cloning and characterization of INCURVATA2 (ICU2), which encodes the putative catalytic subunit of the DNA polymerase alpha of Arabidopsis thaliana. The strong icu2-2 and icu2-3 insertional alleles caused fully penetrant zygotic lethality when homozygous and incompletely penetrant gametophytic lethality, probably because of loss of DNA polymerase activity. The weak icu2-1 allele carried a point mutation and caused early flowering, leaf incurvature, and homeotic transformations of sepals into carpels and of petals into stamens. Further genetic analyses indicated that ICU2 interacts with TERMINAL FLOWER2, the ortholog of HETEROCHROMATIN PROTEIN1 of animals and yeasts, and with the Polycomb group (PcG) gene CURLY LEAF. Another PcG gene, EMBRYONIC FLOWER2, was found to be epistatic to ICU2. Quantitative RT-PCR analyses indicated that a number of regulatory genes were derepressed in the icu2-1 mutant, including genes associated with flowering time, floral meristem, and floral organ identity.

  11. Analysis and validation of genome-specific DNA variations in 5' flanking conserved sequences of wheat low-molecular-weight glutenin subunit genes

    Institute of Scientific and Technical Information of China (English)

    LONG; Hai; WEI; Yuming

    2006-01-01

    The thirty-three 5' flanking conserved sequences of the known low-molecular-weight subunit (LMW-GS) genes have been divided into eight clusters, which was in agreement with the classification based on the deduced N-terminal protein sequences. The DNA polymorphism between the eight clusters was obtained by sequence alignment, and a total of 34 polymorphic positions were observed in the approximately 200 bp regions, among which 18 polymorphic positions were candidate SNPs. Seven cluster-specific primer sets were designed for seven out of eight clusters containing cluster-specific bases, with which the genomic DNA of the ditelosomic lines of group 1 chromosomes of a wheat variety 'Chinese Spring' was employed to carry out chromosome assignment. The subsequent cloning and DNA sequencing of PCR fragments validated the sequences specificity of the 5' flanking conserved sequences between LMW-GS gene groups in different genomes. These results suggested that the coding and 5' flanking regions of LMW-GS genes are likely to have evolved in a concerted fashion. The seven primer sets developed in this study could be used to isolate the complete ORFs of seven groups of LMW-GS genes, respectively, and therefore possess great value for further research in the contributions of a single LMW-GS gene to wheat quality in the complex genetic background and the efficient selections of quality-related components in breeding programs.

  12. Phylogenetic diversity of ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit genes of bacterioplankton in the East China Sea

    Institute of Scientific and Technical Information of China (English)

    ZENG Yonghui; JIAO Nianzhi; CAI Haiyuan; CHEN Xihan; WEI Chaoling

    2004-01-01

    Phylogenetic diversity of Form I and Form Ⅱ ribulose-1, 5-bisphosphate carboxylase/oxygenase (RubisCO) large subunit (rbcL) genes in the inshore and offshore areas of the East China Sea were investigated. Two new primer sets were designed for amplifying partial sequences of rbcL genes from Proteobacteria. Four rbcL gene clone libraries were constructed by amplification and cloning of approximately 640~800 bp sequences of bacterioplankton populations.The method of screening library by denaturing gradient gel electrophoresis (DGGE) was introduced. The results show that the diversity of Form I is higher in offshore waters with higher salinity and lower productivity, while that of Form Ⅱ is higher at the inshore station where salinity is lower and productivity is higher. Several clusters of sequences obtained are deeply rooted and show low similarity (60%~78%) to the known rbcL in existing databases.The degree of diversity of rbcL genes is directly related to environmental variables, including temperature, salinity,pH, dissolved oxygen, etc. These results indicate that rbcL gene can be used as an effective indicator for genetic diversity and population variability of bacterioplankton with the ability of carbon dioxide fixation in the sea.

  13. Association of polymorphisms in nicotinic acetylcholine receptor alpha 4 subunit gene (CHRNA4), mu-opioid receptor gene (OPRM1), and ethanol-metabolizing enzyme genes with alcoholism in Korean patients.

    Science.gov (United States)

    Kim, Soon Ae; Kim, Jong-Woo; Song, Ji-Young; Park, Sunny; Lee, Hee Jae; Chung, Joo-Ho

    2004-01-01

    Findings obtained from several studies indicate that ethanol enhances the activity of alpha4beta2 neuronal nicotinic acetylcholine receptor and support the possibility that a polymorphism of the nicotinic acetylcholine receptor alpha4 subunit gene (CHRNA4) modulates enhancement of nicotinic receptor function by ethanol. To identify the association between the CfoI polymorphism of the CHRNA4 and alcoholism, we examined distribution of genotypes and allele frequencies in Korean patients diagnosed with alcoholism (n = 127) and Korean control subjects without alcoholism (n = 185) with polymerase chain reaction-restriction fragment length polymorphism methods. We were able to detect the association between the CfoI polymorphism of the CHRNA4 and alcoholism in Korean patients (genotype P = .023; allele frequency P = .047). The genotypes and allele frequencies of known polymorphisms in other alcoholism candidate genes, such as alcohol metabolism-related genes [alcohol dehydrogenase 2 (ADH2), aldehyde dehydrogenase 2 (ALDH2), alcohol dehydrogenase 3 (ADH3), and cytochrome P450 2E1 (CYP2E1)] and mu-opioid receptor gene (OPRM1), were studied. The polymorphisms of ADH2, ALDH2, and CYP2E1 were significantly different in Korean patients with alcoholism and Korean control subjects without alcoholism, but ADH3 and OPRM1 did not differ between the two groups.

  14. Analysis of the type IV fimbrial-subunit gene fimA of Xanthomonas hyacinthi: application in PCR-mediated detection of yellow disease in Hyacinths.

    Science.gov (United States)

    van Doorn, J; Hollinger, T C; Oudega, B

    2001-02-01

    A sensitive and specific detection method was developed for Xanthomonas hyacinthi; this method was based on amplification of a subsequence of the type IV fimbrial-subunit gene fimA from strain S148. The fimA gene was amplified by PCR with degenerate DNA primers designed by using the N-terminal and C-terminal amino acid sequences of trypsin fragments of FimA. The nucleotide sequence of fimA was determined and compared with the nucleotide sequences coding for the fimbrial subunits in other type IV fimbria-producing bacteria, such as Xanthomonas campestris pv. vesicatoria, Neisseria gonorrhoeae, and Moraxella bovis. In a PCR internal primers JAAN and JARA, designed by using the nucleotide sequences of the variable central and C-terminal region of fimA, amplified a 226-bp DNA fragment in all X. hyacinthi isolates. This PCR was shown to be pathovar specific, as assessed by testing 71 Xanthomonas pathovars and bacterial isolates belonging to other genera, such as Erwinia and Pseudomonas. Southern hybridization experiments performed with the labelled 226-bp DNA amplicon as a probe suggested that there is only one structural type IV fimbrial-gene cluster in X. hyacinthi. Only two Xanthomonas translucens pathovars cross-reacted weakly in PCR. Primers amplifying a subsequence of the fimA gene of X. campestris pv. vesicatoria (T. Ojanen-Reuhs, N. Kalkkinen, B. Westerlund-Wikström, J. van Doorn, K. Haahtela, E.-L. Nurmiaho-Lassila, K. Wengelink, U. Bonas, and T. K. Korhonen, J. Bacteriol. 179: 1280-1290, 1997) were shown to be pathovar specific, indicating that the fimbrial-subunit sequences are more generally applicable in xanthomonads for detection purposes. Under laboratory conditions, approximately 1,000 CFU of X. hyacinthi per ml could be detected. In inoculated leaves of hyacinths the threshold was 5,000 CFU/ml. The results indicated that infected hyacinths with early symptoms could be successfully screened for X. hyacinthi with PCR.

  15. Cloning, characterization, and expression of a novel member of proteasomal subunits gene in turbot,Scophthalmus maximus

    Institute of Scientific and Technical Information of China (English)

    ZHANG Bo; WANG Xianli; SONG Wenping; ZHENG Debin; MA Chao; XIAO Guangxia

    2015-01-01

    The proteasome is a large, polymeric protease complex responsible for the degradation of intracellular pro-teins and generation of peptides that bind to class I major histocompatibility complex (MHC) molecules. This study identified a new member of proteasomal subunits in turbots (Scophthalmus maximus). The full-length cDNA sequence of turbot proteasomal subunit was obtained. Sequence analysis indicated that its primary structure is highly similar to that ofLMP7 from other vertebrates. The relationship between the turbotLMP7 expression and immune responses to pathogen infection was reported. Quantitative reverse transcriptase polymerase chain reaction showed thatLMP7 was expressed differently in various tissues, with higher expression in the spleen, liver, muscle, and skin. TheLMP7 expression was the highest at 96 h after challenge with lymphocyctis disease virus (LCDV) and at 12 h after challenge withVibrio anguillarum in the turbot liver, kidney, and spleen. Furthermore, theLMP7 expression distinctly increased in turbot kidney cells at 24 h after challenge withV. anguillarumand at 96 h after challenge with LCDV. These results indicate that the turbot LMP7 protein participates in immune responses and may play a significant role in the immune process.

  16. Novel gene sequence variants of pbp2b in penicillin-nonsusceptible Streptococcus pneumonia isolates%青霉素不敏感的肺炎链球菌pbp2b基因新的变异

    Institute of Scientific and Technical Information of China (English)

    田素飞; 褚云卓; 陈佰义; 万建华

    2008-01-01

    Objective To investigate the alternations in gene/amino acid sequence of penicillin-binding protein (PBP) 2b from clinical isolates of penicillin-nonsusceptible Streptococcus pneumonia(PNSP) in this region.Methods 24 strains of Streptococcus pneumonia were collected from January to December 2006.The antibiotics susceptibility of these strains was detected.PCR amplification and direct sequencing of pbp2b genes were performed.The sequence variations of PBP genes of the PNSP in this region were studied with sequence BLAST analysis.Results Three prominent substitutions were common tO 13 PNSP isolates with minimal inhibitory concentration(MIC) at least 0.1 mg/L.These included the replacement of Thr445→Ala following the conservative motif SSN,Glu475→Gly and Thr488→Ala/Ser.The exchange of Glu332→Gly was identified in 12 PNSP isolates of which the MIC was at least 0.25 mg/L.Seven penicillin resistant Streptococcus pneumonia (PRSP) isolates (MIC≥3 mg/L)shared the amino acid substitution Ala618→Gly adiacent to third conserved (KTG) motif and the PBP2b sequences of seven PRSP isolates were classified within Back's group Ⅱ and were very similar to those of the Korean J77 isolate.Novel gene and amino acid sequence variants in isolate 14,15,8,11 and 24 was identified in this study and these gene sequences have been deposited in the GenBank database and assigned accession no.EU035970,EU056919,EU056920,EU056921 and EU106886.Conclusion Analysis of pbp2b genes revealed highly similar patterns of nucleotide and amino acid sequence variation among most resistant isolates.while penicillin intermediate Streptococcus pneumonia might be associated with novel gene sequence variants.%目的 调查本地区青霉素不敏感肺炎链球菌(PNSP)的pbp2b基因和氨基酸序列的变异特点.方法 2006年1-12月收集肺炎链球菌临床分离株24株,检测其对青霉素的敏感性,对青霉素不敏感肺炎链球菌的青霉素结合蛋白pbp2b基因进行PCR扩增和

  17. Cloning and expression of the human N-methyl-D-aspartate receptor subunit NR3A

    DEFF Research Database (Denmark)

    Eriksson, Maria; Nilsson, Anna; Froelich-Fabre, Susanne

    2002-01-01

    Native N-methyl-D-aspartate (NMDA) receptors are heteromeric assemblies of four or five subunits. The NMDA receptor subunits, NR1, NR2A, NR2B, NR2C, and NR2D have been cloned in several species, including man. The NR3A subunit, which in rodents is predominantly expressed during early development......, seems to function by reducing the NMDA receptor response. The human homologue to the rat NR3A, however, had not been cloned. In order to study the functions of the human NR3A (hNR3A), we have cloned and sequenced the hNR3A. It was found to share 88% of the DNA sequence with the rat gene, corresponding...

  18. Genetic Differences of Mitten Crabs Based on RFLP Analysis on Mitochondrial Cytochrome Oxidase Subunit I (COI) Gene

    Institute of Scientific and Technical Information of China (English)

    HU Pengfei; WANG Qian; DAI Wei; WANG Xiaomei

    2008-01-01

    The genetic differences of 15 mitten crab populations from 6 river systems in mainland China and 1 population from Russia were studied based on RFLP analysis of mitochondrial cytochrome oxidase subunit I (COI).The results showed that Tas I-RFLP pattern could be used as a genetic marker to distinguish Eriocheir hepuensis from Eriocheir sinensis, Eriocheirjaponica and Eriocheir leptognathus;genetic distances among 13 populations ofEriocheir sinensis range from 0 to 0.015, indicating that they were different geographic strains;the subspecies status ofEriocheir sinensis and Eriocheir hepuensis (population from Nanliujiang) were considered owning to their genetic distances of 0.02-0.044,indicating that genetic divergence between them was low; Eriocheir leptognathus (population from Nanpaihe, Tianjin) was the most distant taxon with genetic distances value of 0.147-0.195,which could be defined as genetic distances between species in genus Eriocheir.

  19. Efficient cell culture system for hepatitis C virus genotype 2B

    DEFF Research Database (Denmark)

    2014-01-01

    The present inventors developed hepatitis C virus 2b/2a intergenotypic recombinants in which the JFH1 structural genes (Core, E1 and E2), p7 and the complete NS2 were replaced by the corresponding genes of the genotype 2b reference strain J8. Sequence analysis of recovered 2b/2a recombinants from...... transfection experiments revealed that 2b/2a was genetically stable. Conclusion: The developed 2b/2a viruses provide a robust in vitro tool for research in HCV genotype 2b, including vaccine studies and functional analysis.......The present inventors developed hepatitis C virus 2b/2a intergenotypic recombinants in which the JFH1 structural genes (Core, E1 and E2), p7 and the complete NS2 were replaced by the corresponding genes of the genotype 2b reference strain J8. Sequence analysis of recovered 2b/2a recombinants from 2...

  20. 动脉导管未闭患儿TFAP-2B基因突变的研究%Mutation of TFAP-2B gene in patients with patent ductus arteriosus

    Institute of Scientific and Technical Information of China (English)

    陈轶维; 赵武; 李奋; 吉炜; 傅启华; 张志芳; 王剑

    2010-01-01

    目的 发现我国动脉导管未闭(patent ductus arteriosus,PDA)患儿分子遗传方面缺陷,为PDA早期预防及遗传咨询提供支持.方法 收集100例单纯性PDA患儿的临床资料和外周静脉血样本,以100名健康儿童为对照.应用聚合酶链式反应(polymerase chain reaction,PCR)扩增TFAP-2B基因的全部外显子和外显子两侧部分内含子,并对扩增片段进行双向测序.应用BLAST程序将所测TFAP-2B基因序列与GeneBank中的已知序列进行对比以检测基因突变.采用逆转录聚合酶链式反应(reverse transcription polymerase chain reaction,RT-PCR)对1例家族史阳性患儿及其家属共16人TFAP-2B部分cDNA片段进行扩增,扩增产物直接进行双向序列测定.结果 基因分析显示,在1例家族史阳性患儿及其患病亲属中,TFAP-2B第3内含子剪接位点+5位发生突变[intron3(+5)G>A],患儿TFAP-2B基因部分cDNA巢式PCR扩增结果 提示3号外显子完全缺失.此外,还发现了一个新的单核苷酸多态性,即转录起始点上游第34位的鸟嘌呤变为腺嘌呤,这个多态在PDA患者和健康对照组的频率分布差异有统计学意义(Z=-2.513,P=0.012).结论 TFAP-2B基因突变能够导致家族型PDA.%Objective To identify novel genetic mutations in Chinese patients with congenital patent ductus arteriosus (PDA). Method Clinical data and peripheral blood specimens from a kindred spanning 3 generations in which 5 of 16 individuals had PDA and a cohort of 95 unrelated subjects with PDA were collected, and 100 unrelated healthy individuals were included as controls. The coding exons and flanking introns of TFAP-2B gene were amplified by polymerase chain reaction ( PCR ) with specific primers. We aligned the acquired sequences with which publicized in GenBank by the aid of program BLAST. Reverse transcription-polymerase chain reaction (RT-PCR) was used to amplify the parts of TFAP-2B and sequencing was performed on PCR products forward and reversely

  1. The carB gene encoding the large subunit of carbamoylphosphate synthetase from Lactococcus lactis is transcribed monocistronically

    DEFF Research Database (Denmark)

    Martinussen, Jan; Hammer, Karin

    1998-01-01

    the requirement for carbamoylphosphate in pyrimidine biosynthesis through degradation by the arginine deiminase pathway. The expression of the carB gene is subject to regulation at the level of transcription by pyrimidines most probably by an attenuator mechanism. Upstream of the carB gene, an open reading frame...... to be an isolated transcriptional unit. Carbamoylphosphate is a precursor in the biosynthesis of both pyrimidine nucleotides and arginine. By mutant analysis L. lactis is shown to possess only one carB gene; the same gene product is thus required for both biosynthetic pathways. Furthermore, arginine may satisfy...

  2. The Characteristics of Cytochrome C Oxidase Gene Subunit I in Wild Silkmoth Cricula trifenestrata Helfer and Its Evaluation for Species Marker

    Directory of Open Access Journals (Sweden)

    Suriana

    2012-08-01

    Full Text Available The study was conducted to assess the characteristics of partial gene of cytochrome C oxidase subunit I (COI of wild silkmoth Cricula trifenestrata, and to detect the diagnostic sites from these gene for evaluation as species marker. A total of fifteen larvae of C. tifenestrata were collected from Bogor, Purwakarta, and Bantul Regencies. Genomic DNA was extracted from silk gland of individual larvae, then amplified by PCR method and sequenced. DNA sequencing was done to characterize their nucleotide and amino acid contents. The results showed that 595 nucleotides at the 5 ‘end of COI gene of C. tifenestrata was conserved at the species level, but varies at the family level. Nucleotide dominated by thymine and adenine bases (± 70%. There were 25 diagnostic sites for C. tifenestrata, and four diagnostic sites for genus level. One hundred eigthty nine (189 amino acids were alignment, and only one percent of the genes was varied among species. The 107th amino acid (valine and 138th (threonine were diagnostics amino acid for C. tifenestrata. Based on nucleotides and amino acids sequences, the phylogeny showed that C. tifenestrata lied on the same nodes with Antheraea, so the Saturniidae family is monophyletic.

  3. An intact F1ATPase alpha-subunit gene and a pseudogene with differing genomic organization are detected in both male-fertile and CMS petunia mitochondria.

    Science.gov (United States)

    Yesodi, V; Hauschner, H; Tabib, Y; Firon, N

    1997-11-01

    The gene copies for the alpha-subunit of the mitochondrial F1ATPase (atpA) were isolated and characterized in both male-fertile and cytoplasmic male sterile (CMS) petunia. Two copies, an intact gene and a truncated gene, were detected in both cytoplasms. The accumulated data, based upon a comparison of the sequences (the open reading frames as well as the 5' and 3' flanking regions) of the two atpA copies, both in male-fertile and CMS Petunia, indicate that: (1) they differ in their genomic organization and (2) a common progenitor cytoplasm, containing two copies of an intact atpA sequence, served as the origin for the atpA copies of the fertility and CMS-inducing cytoplasms. Homologous recombination through the progenitor intact atpA sequences is assumed to have caused the rearrangement in the 3' portion of the atpA open reading frame and the generation of the truncated atpA gene. It is thus suggested that the atpA pseudogenes, in both male-fertile and CMS cytoplasms, originated from a common progenitor atpA pseudogene sequence.

  4. Increase of AMPA receptor glutamate receptor 1 subunit and B-cell receptor-associated protein 31 gene expression in hippocampus of fatigued mice.

    Science.gov (United States)

    Kamakura, Masaki; Tamaki, Keisuke; Sakaki, Toshiyuki; Yoneda, Yukio

    2005-10-14

    Central fatigue is an indispensable biosignal for maintaining life, but the neuronal and molecular mechanisms involved remain unclear. In this study, we searched for genes differentially expressed in the hippocampus of fatigued mice to elucidate the mechanisms underlying fatigue. Mice were forced to swim in an adjustable-current water pool, and the maximum swimming time (endurance) until fatigue was measured thrice. Fatigued and nonfatigued mice with equal swimming capacity and body weight were compared. We found that the genes of GluR1 and B-cell receptor-associated protein 31 (Bap31), which acts as a transport molecule in the secretory pathway or as a mediator of apoptosis, were upregulated in the hippocampus of fatigued mice, and increases of GluR1 and Bap31 were confirmed by Northern blotting and real-time PCR. No change of gene expression of AMPA receptor subunits other than GluR1 was observed. These results suggest that a compositional change of AMPA receptor (increase of GluR1) and upregulation of the Bap31 gene may be implicated in fatigue in mice.

  5. SNP detection in Na/K ATP-ase gene α1 subunit of bisexual and parthenogenetic Artemia strains by RFLP screening.

    Science.gov (United States)

    Manaffar, R; Zare, S; Agh, N; Abdolahzadeh, N; Soltanian, S; Sorgeloos, P; Bossier, P; Van Stappen, G

    2011-01-01

    In order to find a marker for differentiating between a bisexual and a parthenogenetic Artemia strain, Exon-7 of the Na/K ATPase α(1) subunit gene was screened by RFLP technique. The results revealed a constant synonymous SNP (single nucleotide polymorphism) in digestion by the Tru1I enzyme that was consistent with these two types of Artemia. This SNP was identified as an accurate molecular marker for discrimination between bisexual and parthenogenetic Artemia. According to the Nei's genetic distance (1973), the lowest genetic distance was found between individuals from Artemia urmiana Günther 1890 and parthenogenetic populations, making the described marker the first marker to easily distinguish between these two cooccurring species.

  6. Population structure of the Monocelis lineata (Proseriata, Monocelididae species complex assessed by phylogenetic analysis of the mitochondrial Cytochrome c Oxidase subunit I (COI gene

    Directory of Open Access Journals (Sweden)

    Daria Sanna

    2009-01-01

    Full Text Available Monocelis lineata consists of a complex of sibling species, widespread in the Mediterranean and Atlantic Ocean. Previous genetic analysis placed in evidence at least four sibling species. Nevertheless, this research was not conclusive enough to fully resolve the complex or to infer the phylogeny/phylogeography of the group. We designed specific primers aiming at obtaining partial sequences of the mtDNA gene Cytochrome c Oxidase subunit I (COI of M. lineata, and have identified 25 different haplotypes in 32 analyzed individuals. The dendrogram generated by Neighbor-Joining analysis confirmed the differentiation between Atlantic and Mediterranean siblings, as well as the occurrence of at least two Mediterranean sibling species. Thus validated, the method here presented appears as a valuable tool in population genetics and biodiversity surveys on the Monocelis lineata complex.

  7. Differentiation of expression proifles of two calcineurin subunit genes in chicken skeletal muscles during early postnatal growth depending on anatomical location of muscles and breed

    Institute of Scientific and Technical Information of China (English)

    SHAN Yan-ju; XU Wen-juan; SHU Jing-ting; ZHANG Ming; SONG Wei-tao; TAO Zhi-yun; ZHU Chun-hong; LI Hui-fang

    2016-01-01

    Calcineurin (Cn or CaN) is implicated in the control of skeletal muscle ifber phenotype and hypertrophy. However, little information is available concerning the expression of Cn in chickens. In the present study, the expression of two Cn subunit genes (CnAα andCnB1) was quantiifed by qPCR in the lateral gastrocnemius (LG, mainly composing of red fast-twitch myoifbers), the soleus (mainly composing of red slow-twitch myoifbers) and the extensor digitorum longus (EDL, mainly composing of white fast-twitch myoifbers) from Qingyuan partridge chickens (QY, slow-growing chicken breed) and Recessive White chickens (RW, fast-growing chicken breed) on different days (1, 8, 22, 36, 50 and 64 days post-hatching). Although CnAα andCnB1 gene expressions were variable with different trends in different skeletal muscles in the two chicken breeds during postnatal growth, it is highly muscle phenotype and breed speciifc. In general, the levels ofCnAαandCnB1gene expressions of the soleus were lower than those of EDL and LG in both chicken breeds at the same stages. Compared be-tween the two chicken breeds, the levels ofCnAα gene expression of the three skeletal muscles in QY chickens were higher than those in RW chickens on days 1 and 22. However, on day 64, the levels of bothCnAα andCnB1 gene expressions of the three skeletal muscles were lower in QY chickens than those in RW chickens. Correlation analysis of the levels of CnAα andCnB1 gene expressions of the same skeletal muscle showed that there were positive correlations for al three skeletal muscle tissues in two chicken breeds. These results provide some valuable clues to understand the role of Cn in the development of chicken skeletal muscles, with a function that may be related to meat quality.

  8. Expression of NR2B in cerebellar granule cells specifically facilitates effect of motor training on motor learning.

    Science.gov (United States)

    Jiao, Jianwei; Nakajima, Akira; Janssen, William G M; Bindokas, Vytautas P; Xiong, Xiaoli; Morrison, John H; Brorson, James R; Tang, Ya-Ping

    2008-02-27

    It is believed that gene/environment interaction (GEI) plays a pivotal role in the development of motor skills, which are acquired via practicing or motor training. However, the underlying molecular/neuronal mechanisms are still unclear. Here, we reported that the expression of NR2B, a subunit of NMDA receptors, in cerebellar granule cells specifically enhanced the effect of voluntary motor training on motor learning in the mouse. Moreover, this effect was characterized as motor learning-specific and developmental stage-dependent, because neither emotional/spatial memory was affected nor was the enhanced motor learning observed when the motor training was conducted starting at the age of 3 months old in these transgenic mice. These results indicate that changes in the expression of gene(s) that are involved in regulating synaptic plasticity in cerebellar granule cells may constitute a molecular basis for the cerebellum to be involved in the GEI by facilitating motor skill learning.

  9. Expression of NR2B in cerebellar granule cells specifically facilitates effect of motor training on motor learning.

    Directory of Open Access Journals (Sweden)

    Jianwei Jiao

    Full Text Available It is believed that gene/environment interaction (GEI plays a pivotal role in the development of motor skills, which are acquired via practicing or motor training. However, the underlying molecular/neuronal mechanisms are still unclear. Here, we reported that the expression of NR2B, a subunit of NMDA receptors, in cerebellar granule cells specifically enhanced the effect of voluntary motor training on motor learning in the mouse. Moreover, this effect was characterized as motor learning-specific and developmental stage-dependent, because neither emotional/spatial memory was affected nor was the enhanced motor learning observed when the motor training was conducted starting at the age of 3 months old in these transgenic mice. These results indicate that changes in the expression of gene(s that are involved in regulating synaptic plasticity in cerebellar granule cells may constitute a molecular basis for the cerebellum to be involved in the GEI by facilitating motor skill learning.

  10. An upstream initiator caspase 10 of snakehead murrel Channa striatus, containing DED, p20 and p10 subunits: molecular cloning, gene expression and proteolytic activity.

    Science.gov (United States)

    Arockiaraj, Jesu; Gnanam, Annie J; Muthukrishnan, Dhanaraj; Pasupuleti, Mukesh; Milton, James; Singh, Arun

    2013-02-01

    Caspase 10 (CsCasp10) was identified from a constructed cDNA library of freshwater murrel (otherwise called snakehead) Channa striatus. The CsCasp10 is 1838 base pairs (bp) in length and it is encoding 549 amino acid (aa) residues. CsCasp10 amino acid contains two death effector domains (DED) in the N-terminal at 2-77 and 87-154 and it contains caspase family p20 domain (large subunit) and caspase family p10 domain (small subunit) in the C-terminal at 299-425 and 449-536 respectively. Pairwise analysis of CsCasp10 showed the highest sequence similarity (79%) with caspase 10 of Paralichthys olivaceus. Moreover, the phylogenetic analysis showed that CsCasp10 is clustered together with other fish caspase 10, formed a sister group with caspase 10 from other lower vertebrates including amphibian, reptile and birds and finally clustered together with higher vertebrates such as mammals. Significantly (P < 0.05) highest CsCasp10 gene expression was noticed in gills and lowest in intestine. Furthermore, the CsCasp10 gene expression in C. striatus was up-regulated in gills by fungus Aphanomyces invadans and bacteria Aeromonas hydrophila induction. The proteolytic activity was analyzed using the purified recombinant CsCasp10 protein. The results showed the proteolytic activity of CsCasp10 for caspase 10 substrate was 2.5 units per μg protein. Moreover, the proteolytic activities of CsCasp10 in kidney and spleen induced by A. invadans and A. hydrophila stimulation were analyzed by caspase 10 activity assay kit. All these results showed that CsCasp10 are participated in immunity of C. striatus against A. invadans and A. hydrophila infection.

  11. Association of Common Polymorphisms in the Nicotinic Acetylcholine Receptor Alpha4 Subunit Gene with an Electrophysiological Endophenotype in a Large Population-Based Sample.

    Directory of Open Access Journals (Sweden)

    A Mobascher

    Full Text Available Variation in genes coding for nicotinic acetylcholine receptor (nAChR subunits affect cognitive processes and may contribute to the genetic architecture of neuropsychiatric disorders. Single nucleotide polymorphisms (SNPs in the CHRNA4 gene that codes for the alpha4 subunit of alpha4/beta2-containing receptors have previously been implicated in aspects of (mostly visual attention and smoking-related behavioral measures. Here we investigated the effects of six synonymous but functional CHRNA4 exon 5 SNPs on the N100 event-related potential (ERP, an electrophysiological endophenotype elicited by a standard auditory oddball. A total of N = 1,705 subjects randomly selected from the general population were studied with electroencephalography (EEG as part of the German Multicenter Study on nicotine addiction. Two of the six variants, rs1044396 and neighboring rs1044397, were significantly associated with N100 amplitude. This effect was pronounced in females where we also observed an effect on reaction time. Sequencing of the complete exon 5 region in the population sample excluded the existence of additional/functional variants that may be responsible for the observed effects. This is the first large-scale population-based study investigation the effects of CHRNA4 SNPs on brain activity measures related to stimulus processing and attention. Our results provide further evidence that common synonymous CHRNA4 exon 5 SNPs affect cognitive processes and suggest that they also play a role in the auditory system. As N100 amplitude reduction is considered a schizophrenia-related endophenotype the SNPs studied here may also be associated with schizophrenia outcome measures.

  12. The archipelago ubiquitin ligase subunit acts in target tissue to restrict tracheal terminal cell branching and hypoxic-induced gene expression.

    Directory of Open Access Journals (Sweden)

    Nathan T Mortimer

    Full Text Available The Drosophila melanogaster gene archipelago (ago encodes the F-box/WD-repeat protein substrate specificity factor for an SCF (Skp/Cullin/F-box-type polyubiquitin ligase that inhibits tumor-like growth by targeting proteins for degradation by the proteasome. The Ago protein is expressed widely in the fly embryo and larva and promotes degradation of pro-proliferative proteins in mitotically active cells. However the requirement for Ago in post-mitotic developmental processes remains largely unexplored. Here we show that Ago is an antagonist of the physiologic response to low oxygen (hypoxia. Reducing Ago activity in larval muscle cells elicits enhanced branching of nearby tracheal terminal cells in normoxia. This tracheogenic phenotype shows a genetic dependence on sima, which encodes the HIF-1α subunit of the hypoxia-inducible transcription factor dHIF and its target the FGF ligand branchless (bnl, and is enhanced by depletion of the Drosophila Von Hippel Lindau (dVHL factor, which is a subunit of an oxygen-dependent ubiquitin ligase that degrades Sima/HIF-1α protein in metazoan cells. Genetic reduction of ago results in constitutive expression of some hypoxia-inducible genes in normoxia, increases the sensitivity of others to mild hypoxic stimulus, and enhances the ability of adult flies to recover from hypoxic stupor. As a molecular correlate to these genetic data, we find that Ago physically associates with Sima and restricts Sima levels in vivo. Collectively, these findings identify Ago as a required element of a circuit that suppresses the tracheogenic activity of larval muscle cells by antagonizing the Sima-mediated transcriptional response to hypoxia.

  13. Regulation of the nuclear gene that encodes the alpha-subunit of the mitochondrial F0F1-ATP synthase complex. Activation by upstream stimulatory factor 2.

    Science.gov (United States)

    Breen, G A; Jordan, E M

    1997-04-18

    We have previously identified several positive cis-acting regulatory regions in the promoters of the bovine and human nuclear-encoded mitochondrial F0F1-ATP synthase alpha-subunit genes (ATPA). One of these cis-acting regions contains the sequence 5'-CACGTG-3' (an E-box), to which a number of transcription factors containing a basic helix-loop-helix motif can bind. This E-box element is required for maximum activity of the ATPA promoter in HeLa cells. The present study identifies the human transcription factor, upstream stimulatory factor 2 (USF2), as a nuclear factor that binds to the ATPA E-box and demonstrates that USF2 plays a critical role in the activation of the ATPA gene in vivo. Evidence includes the following. Antiserum directed against USF2 recognized factors present in HeLa nuclear extracts that interact with the ATPA promoter in mobility shift assays. Wild-type USF2 proteins synthesized from expression vectors trans-activated the ATPA promoter through the E-box, whereas truncated USF2 proteins devoid of the amino-terminal activation domains did not. Importantly, expression of a dominant-negative mutant of USF2 lacking the basic DNA binding domain but able to dimerize with endogenous USF proteins significantly reduced the level of activation of the ATPA promoter caused by ectopically coexpressed USF2, demonstrating the importance of endogenous USF2 in activation of the ATPA gene.

  14. The largest subunit of RNA polymerase II as a new marker gene to study assemblages of arbuscular mycorrhizal fungi in the field.

    Directory of Open Access Journals (Sweden)

    Herbert Stockinger

    Full Text Available Due to the potential of arbuscular mycorrhizal fungi (AMF, Glomeromycota to improve plant growth and soil quality, the influence of agricultural practice on their diversity continues to be an important research question. Up to now studies of community diversity in AMF have exclusively been based on nuclear ribosomal gene regions, which in AMF show high intra-organism polymorphism, seriously complicating interpretation of these data. We designed specific PCR primers for 454 sequencing of a region of the largest subunit of RNA polymerase II gene, and established a new reference dataset comprising all major AMF lineages. This gene is known to be monomorphic within fungal isolates but shows an excellent barcode gap between species. We designed a primer set to amplify all known lineages of AMF and demonstrated its applicability in combination with high-throughput sequencing in a long-term tillage experiment. The PCR primers showed a specificity of 99.94% for glomeromycotan sequences. We found evidence of significant shifts of the AMF communities caused by soil management and showed that tillage effects on different AMF taxa are clearly more complex than previously thought. The high resolving power of high-throughput sequencing highlights the need for quantitative measurements to efficiently detect these effects.

  15. Phylogenetic relationships within Taenia taeniaeformis variants and other taeniid cestodes inferred from the nucleotide sequence of the cytochrome c oxidase subunit I gene.

    Science.gov (United States)

    Okamoto, M; Bessho, Y; Kamiya, M; Kurosawa, T; Horii, T

    1995-01-01

    Nucleotide sequence variations in a region of the mitochondrial cytochrome c oxidase subunit I (COI) gene (391 bp) were examined within seven species of the genus Taenia and two species of the genus Echinococcus, including ten isolates of T. taeniaeformis and six isolates of E. multilocularis. More than a 12% rate of nucleotide differences between taeniid species was found, allowing the species to be distinguished. In E. multilocularis, no sequence variation was observed among isolates, regardless of the host (gray red-backed vole, tundra vole, pig, Norway rat) or area (Japan, Alaska) from which each metacestode had been isolated. In contrast, six distinct sequences were detected among the ten T. taeniaeformis isolates examined. The level of nucleotide variation in the COI gene within T. taeniaeformis isolates except for one isolate from the gray red-backed vole (TtACR), which has been proposed as a distinct strain or a different species, was about 0.3%-4.1%, whereas the COI gene sequence for TtACR differed from those of the other isolates, with levels being 9.0%-9.5%. Phylogenetic trees were then inferred from these sequence data using two different algorithms.

  16. Characterization of a gene from chromosome 1B encoding the large subunit of ADPglucose pyrophosphorylase from wheat: evolutionary divergence and differential expression of Agp2 genes between leaves and developing endosperm.

    Science.gov (United States)

    Thorneycroft, David; Hosein, Felicia; Thangavelu, Madan; Clark, Joanna; Vizir, Igor; Burrell, Michael M; Ainsworth, Charles

    2003-07-01

    A full-length genomic clone containing the gene encoding the large subunit of the ADPglucose pyrophosphorylase (Agp2), was isolated from a genomic library prepared from etiolated shoots of hexaploid wheat (Triticum aestivum L., cv, Chinese Spring). The coding region of this gene is identical to one of the cDNA clones previously isolated from a developing wheat grain cDNA library and is therefore an actively transcribed gene. The sequence represented by the cDNA spans 4.8 kb of the genomic clone and contains 15 introns. 2852 bp of DNA flanking the transcription start site of the gene was cloned upstream of the GUS (beta-glucuronidase) reporter gene. This Agp2::GUS construct and promoter deletions were used to study the pattern of reporter gene expression in both transgenic tobacco and wheat plants. Histochemical analysis of GUS expression in transgenic tobacco demonstrated that the reporter gene was expressed in guard cells of leaves and throughout the seed. In transgenic wheat, reporter gene expression was confined to the endosperm and aleurone with no expression in leaves. The cloned Agp2 gene was located to chromosome 1B by gene-specific PCR with nullisomic-tetrasomic lines. Northern analysis demonstrated that the Agp2 genes are differentially expressed in leaves and developing endosperm; while all three classes of Agp2 genes are transcribed in developing wheat grain endosperm, only one is transcribed in leaves. The differences between the Agp2 genes are discussed in relation to the evolution of hexaploid wheat.

  17. A Cyclin Dependent Kinase Regulatory Subunit (CKS) Gene of Pigeonpea Imparts Abiotic Stress Tolerance and Regulates Plant Growth and Development in Arabidopsis.

    Science.gov (United States)

    Tamirisa, Srinath; Vudem, Dashavantha R; Khareedu, Venkateswara R

    2017-01-01

    Frequent climatic changes in conjunction with other extreme environmental factors are known to affect growth, development and productivity of diverse crop plants. Pigeonpea, a major grain legume of the semiarid tropics, endowed with an excellent deep-root system, is known as one of the important drought tolerant crop plants. Cyclin dependent kinases (CDKs) are core cell cycle regulators and play important role in different aspects of plant growth and development. The cyclin-dependent kinase regulatory subunit gene (CKS) was isolated from the cDNA library of pigeonpea plants subjected to drought stress. Pigeonpea CKS (CcCKS) gene expression was detected in both the root and leaf tissues of pigeonpea and was upregulated by polyethylene glycol (PEG), mannitol, NaCl and abscisic acid (ABA) treatments. The overexpression of CcCKS gene in Arabidopsis significantly enhanced tolerance of transgenics to drought and salt stresses as evidenced by different physiological parameters. Under stress conditions, transgenics showed higher biomass, decreased rate of water loss, decreased MDA levels, higher free proline contents, and glutathione levels. Moreover, under stress conditions transgenics exhibited lower stomatal conductance, lower transpiration, and higher photosynthetic rates. However, under normal conditions, CcCKS-transgenics displayed decreased plant growth rate, increased cell size and decreased stomatal number compared to those of wild-type plants. Real-time polymerase chain reaction revealed that CcCKS could regulate the expression of both ABA-dependent and ABA-independent genes associated with abiotic stress tolerance as well as plant growth and development. As such, the CcCKS seems promising and might serve as a potential candidate gene for enhancing the abiotic stress tolerance of crop plants.

  18. GluN2B-containing NMDA receptors promote glutamate synapse development in hippocampal interneurons.

    Science.gov (United States)

    Kelsch, Wolfgang; Li, Zhijun; Wieland, Sebastian; Senkov, Oleg; Herb, Anne; Göngrich, Christina; Monyer, Hannah

    2014-11-26

    In postnatal development, GluN2B-containing NMDARs are critical for the functional maturation of glutamatergic synapses. GluN2B-containing NMDARs prevail until the second postnatal week when GluN2A subunits are progressively added, conferring mature properties to NMDARs. In cortical principal neurons, deletion of GluN2B results in an increase in functional AMPAR synapses, suggesting that GluN2B-containing NMDARs set a brake on glutamate synapse maturation. The function of GluN2B in the maturation of glutamatergic inputs to cortical interneurons is not known. To examine the function of GluN2B in interneurons, we generated mutant mice with conditional deletion of GluN2B in interneurons (GluN2B(ΔGAD67)). In GluN2B(ΔGAD67) mice interneurons distributed normally in cortical brain regions. After the second postnatal week, GluN2B(ΔGAD67) mice developed hippocampal seizures and died shortly thereafter. Before the onset of seizures, GluN2B-deficient hippocampal interneurons received fewer glutamatergic synaptic inputs than littermate controls, indicating that GluN2B-containing NMDARs positively regulate the maturation of glutamatergic input synapses in interneurons. These findings suggest that GluN2B-containing NMDARs keep the circuit activity under control by promoting the maturation of excitatory synapses in interneurons.

  19. Knockdown of the Rhipicephalus microplus cytochrome c oxidase subunit III gene is associated with a failure of Anaplasma marginale transmission.

    Directory of Open Access Journals (Sweden)

    Thais D Bifano

    Full Text Available Rhipicephalus microplus is an obligate hematophagous ectoparasite of cattle and an important biological vector of Anaplasma marginale in tropical and subtropical regions. The primary determinants for A. marginale transmission are infection of the tick gut, followed by infection of salivary glands. Transmission of A. marginale to cattle occurs via infected saliva delivered during tick feeding. Interference in colonization of either the tick gut or salivary glands can affect transmission of A. marginale to naïve animals. In this study, we used the tick embryonic cell line BME26 to identify genes that are modulated in response to A. marginale infection. Suppression-subtractive hybridization libraries (SSH were constructed, and five up-regulated genes {glutathione S-transferase (GST, cytochrome c oxidase sub III (COXIII, dynein (DYN, synaptobrevin (SYN and phosphatidylinositol-3,4,5-triphosphate 3-phosphatase (PHOS} were selected as targets for functional in vivo genomic analysis. RNA interference (RNAi was used to determine the effect of tick gene knockdown on A. marginale acquisition and transmission. Although RNAi consistently knocked down all individually examined tick genes in infected tick guts and salivary glands, only the group of ticks injected with dsCOXIII failed to transmit A. marginale to naïve calves. To our knowledge, this is the first report demonstrating that RNAi of a tick gene is associated with a failure of A. marginale transmission.

  20. Analysis of the cytochrome c oxidase subunit 1 (COX1) gene reveals the unique evolution of the giant panda.

    Science.gov (United States)

    Hu, Yao-Dong; Pang, Hui-Zhong; Li, De-Sheng; Ling, Shan-Shan; Lan, Dan; Wang, Ye; Zhu, Yun; Li, Di-Yan; Wei, Rong-Ping; Zhang, He-Min; Wang, Cheng-Dong

    2016-11-05

    As the rate-limiting enzyme of the mitochondrial respiratory chain, cytochrome c oxidase (COX) plays a crucial role in biological metabolism. "Living fossil" giant panda (Ailuropoda melanoleuca) is well-known for its special bamboo diet. In an effort to explore functional variation of COX1 in the energy metabolism behind giant panda's low-energy bamboo diet, we looked at genetic variation of COX1 gene in giant panda, and tested for its selection effect. In 1545 base pairs of the gene from 15 samples, 9 positions were variable and 1 mutation leaded to an amino acid sequence change. COX1 gene produces six haplotypes, nucleotide (pi), haplotype diversity (Hd). In addition, the average number of nucleotide differences (k) is 0.001629±0.001036, 0.8083±0.0694 and 2.517, respectively. Also, dN/dS ratio is significantly below 1. These results indicated that giant panda had a low population genetic diversity, and an obvious purifying selection of the COX1 gene which reduces synthesis of ATP determines giant panda's low-energy bamboo diet. Phylogenetic trees based on the COX1 gene were constructed to demonstrate that giant panda is the sister group of other Ursidae.

  1. The effects of clobazam treatment in rats on the expression of genes and proteins encoding glucronosyltransferase 1A/2B (UGT1A/2B) and multidrug resistance-associated protein-2 (MRP2), and development of thyroid follicular cell hypertrophy.

    Science.gov (United States)

    Miyawaki, Izuru; Tamura, Akitoshi; Matsumoto, Izumi; Inada, Hiroshi; Kunimatsu, Takeshi; Kimura, Juki; Funabashi, Hitoshi

    2012-12-15

    Clobazam (CLB) is known to increase hepatobiliary thyroxine (T4) clearance in Sprague-Dawley (SD) rats, which results in hypothyroidism followed by thyroid follicular cell hypertrophy. However, the mechanism of the acceleration of T4-clearance has not been fully investigated. In the present study, we tried to clarify the roles of hepatic UDP-glucronosyltransferase (UGT) isoenzymes (UGT1A and UGT2B) and efflux transporter (multidrug resistance-associated protein-2; MRP2) in the CLB-induced acceleration of T4-clearance using two mutant rat strains, UGT1A-deficient mutant (Gunn) and MRP2-deficient mutant (EHBR) rats, especially focusing on thyroid morphology, levels of circulating hormones (T4 and triiodothyronine (T3)) and thyroid-stimulating hormone (TSH), and mRNA or protein expressions of UGTs (Ugt1a1, Ugt1a6, and Ugt2b1/2) and MRP2 (Mrp). CLB induced thyroid morphological changes with increases in TSH in SD and Gunn rats, but not in EHBR rats. T4 was slightly decreased in SD and Gunn rats, and T3 was decreased in Gunn rats, whereas these hormones were maintained in EHBR rats. Hepatic Ugt1a1, Ugt1a6, Ugt2b1/2, and Mrp2 mRNAs were upregulated in SD rats. In Gunn rats, UGT1A mRNAs (Ugt1a1/6) and protein levels were quite low, but UGT2B mRNAs (Ugt2b1/2) and protein were prominently upregulated. In SD and Gunn rats, MRP2 mRNA and protein were upregulated to the same degree. These results suggest that MRP2 is an important contributor in development of the thyroid cellular hypertrophy in CLB-treated rats, and that UGT1A and UGT2B work in concert with MRP2 in the presence of MRP2 function to enable the effective elimination of thyroid hormones.

  2. Forebrain NR2B overexpression facilitating the prefrontal cortex long-term potentiation and enhancing working memory function in mice.

    Directory of Open Access Journals (Sweden)

    Yihui Cui

    Full Text Available Prefrontal cortex plays an important role in working memory, attention regulation and behavioral inhibition. Its functions are associated with NMDA receptors. However, there is little information regarding the roles of NMDA receptor NR2B subunit in prefrontal cortical synaptic plasticity and prefrontal cortex-related working memory. Whether the up-regulation of NR2B subunit influences prefrontal cortical synaptic plasticity and working memory is not yet clear. In the present study, we measured prefrontal cortical synaptic plasticity and working memory function in NR2B overexpressing transgenic mice. In vitro electrophysiological data showed that overexpression of NR2B specifically in the forebrain region resulted in enhancement of prefrontal cortical long-term potentiation (LTP but did not alter long-term depression (LTD. The enhanced LTP was completely abolished by a NR2B subunit selective antagonist, Ro25-6981, indicating that overexpression of NR2B subunit is responsible for enhanced LTP. In addition, NR2B transgenic mice exhibited better performance in a set of working memory paradigms including delay no-match-to-place T-maze, working memory version of water maze and odor span task. Our study provides evidence that NR2B subunit of NMDA receptor in prefrontal cortex is critical for prefrontal cortex LTP and prefrontal cortex-related working memory.

  3. Association of the deleted DAZ gene copy related to gr/gr and b2/b3 deletions with spermatogenic impairment%gr/gr和b2/b3缺失中不同DAZ拷贝缺失与精子发生障碍的相关性研究

    Institute of Scientific and Technical Information of China (English)

    王亚民; 李权; 宋乐彬; 张嘉宜; 杨杰; 宋宁宏

    2016-01-01

    目的:研究AZFc区gr/gr和b2/b3缺失中DAZ基因拷贝缺失对中国男性精子发生障碍的影响.方法:试验组纳入121例不同程度生精障碍的不育男性,对照组选择了95例健康男性且均符合《WHO人类精液检查与处理实验室手册》第5版标准,通过常规PCR和PCR-RFLP以及Y染色体特异性序列标签位点(STS),分析AZFc区不同类型gr/gr和b2/b3缺失中DAZ基因拷贝缺失与男性精子发生障碍的关系. 结果:对照组中共观察到13例(13.68%) gr/gr缺失和1例(1.05%) b2/b3缺失,试验组中发现15例(12.40%) gr/gr缺失及6例(4.96%) b2/b3缺失.DAZ特异性单核苷酸变异位点(SNV)分析显示对照组中有3例(3.16%) gr/gr-DAZ1/DAZ2缺失和10例(10.53%) gr/gr-DAZ3/DAZ4缺失以及1例(1.05%) b2/b3-DAZ3/DAZ4缺失,试验组中有11例(9.09%) gr/gr-DAZ1/DAZ2缺失和4例(3.31%) gr/gr-DAZ3/DAZ4缺失以及6例(4.96%) b2/b3-DAZ1/DAZ2缺失. 结论:中国男性Y染色体AZFc部分缺失(gr/gr、b2/b3缺失)在正常生精者和不同程度的生精障碍患者中均存在,且差异无统计学意义,不能作为精子发生障碍的危险因素.但gr/铲和b2/b3缺失中不同类型的DAZ拷贝子缺失对男性精子发生的影响不同,DAZ1/DAZ2缺失与男性生精功能障碍相关,可能是导致男性不育的危险因素,而DAZ3/DAZ4缺失似乎对生精的影响很小或不起作用.

  4. Role of positron emission tomography and bone scintigraphy in the evaluation of bone involvement in metastatic pheochromocytoma and paraganglioma: specific implications for succinate dehydrogenase enzyme subunit B gene mutations.

    NARCIS (Netherlands)

    Zelinka, T.; Timmers, H.J.L.M.; Kozupa, A.; Chen, C.C.; Carrasquillo, J.A.; Reynolds, J.C.; Ling, A.; Eisenhofer, G.; Lazurova, I.; Adams, K.T.; Whatley, M.A.; Widimsky, J.Jr.; Pacak, K.

    2008-01-01

    We performed a retrospective analysis of 71 subjects with metastatic pheochromocytoma and paraganglioma (30 subjects with mutation of succinate dehydrogenase enzyme subunit B (SDHB) gene and 41 subjects without SDHB mutation). Sixty-nine percent presented with bone metastases (SDHB +/-: 77% vs 63%),

  5. Molecular cloning and functional expression of the Equine K+ channel KV11.1 (Ether à Go-Go-related/KCNH2 gene) and the regulatory subunit KCNE2 from equine myocardium

    DEFF Research Database (Denmark)

    Pedersen, Philip Juul; Thomsen, Kirsten Brolin; Olander, Emma Rie;

    2015-01-01

    The KCNH2 and KCNE2 genes encode the cardiac voltage-gated K+ channel KV11.1 and its auxiliary β subunit KCNE2. KV11.1 is critical for repolarization of the cardiac action potential. In humans, mutations or drug therapy affecting the KV11.1 channel are associated with prolongation of the QT inter...

  6. Gene expression profile of sodium channel subunits in the anterior cingulate cortex during experimental paclitaxel-induced neuropathic pain in mice

    Directory of Open Access Journals (Sweden)

    Willias Masocha

    2016-11-01

    Full Text Available Paclitaxel, a chemotherapeutic agent, causes neuropathic pain whose supraspinal pathophysiology is not fully understood. Dysregulation of sodium channel expression, studied mainly in the periphery and spinal cord level, contributes to the pathogenesis of neuropathic pain. We examined gene expression of sodium channel (Nav subunits by real time polymerase chain reaction (PCR in the anterior cingulate cortex (ACC at day 7 post first administration of paclitaxel, when mice had developed paclitaxel-induced thermal hyperalgesia. The ACC was chosen because increased activity in the ACC has been observed during neuropathic pain. In the ACC of vehicle-treated animals the threshold cycle (Ct values for Nav1.4, Nav1.5, Nav1.7, Nav1.8 and Nav1.9 were above 30 and/or not detectable in some samples. Thus, comparison in mRNA expression between untreated control, vehicle-treated and paclitaxel treated animals was done for Nav1.1, Nav1.2, Nav1.3, Nav1.6, Nax as well as Navβ1–Navβ4. There were no differences in the transcript levels of Nav1.1–Nav1.3, Nav1.6, Nax, Navβ1–Navβ3 between untreated and vehicle-treated mice, however, vehicle treatment increased Navβ4 expression. Paclitaxel treatment significantly increased the mRNA expression of Nav1.1, Nav1.2, Nav1.6 and Nax, but not Nav1.3, sodium channel alpha subunits compared to vehicle-treated animals. Treatment with paclitaxel significantly increased the expression of Navβ1 and Navβ3, but not Navβ2 and Navβ4, sodium channel beta subunits compared to vehicle-treated animals. These findings suggest that during paclitaxel-induced neuropathic pain (PINP there is differential upregulation of sodium channels in the ACC, which might contribute to the increased neuronal activity observed in the area during neuropathic pain.

  7. Gene expression profile of sodium channel subunits in the anterior cingulate cortex during experimental paclitaxel-induced neuropathic pain in mice

    Science.gov (United States)

    2016-01-01

    Paclitaxel, a chemotherapeutic agent, causes neuropathic pain whose supraspinal pathophysiology is not fully understood. Dysregulation of sodium channel expression, studied mainly in the periphery and spinal cord level, contributes to the pathogenesis of neuropathic pain. We examined gene expression of sodium channel (Nav) subunits by real time polymerase chain reaction (PCR) in the anterior cingulate cortex (ACC) at day 7 post first administration of paclitaxel, when mice had developed paclitaxel-induced thermal hyperalgesia. The ACC was chosen because increased activity in the ACC has been observed during neuropathic pain. In the ACC of vehicle-treated animals the threshold cycle (Ct) values for Nav1.4, Nav1.5, Nav1.7, Nav1.8 and Nav1.9 were above 30 and/or not detectable in some samples. Thus, comparison in mRNA expression between untreated control, vehicle-treated and paclitaxel treated animals was done for Nav1.1, Nav1.2, Nav1.3, Nav1.6, Nax as well as Navβ1–Navβ4. There were no differences in the transcript levels of Nav1.1–Nav1.3, Nav1.6, Nax, Navβ1–Navβ3 between untreated and vehicle-treated mice, however, vehicle treatment increased Navβ4 expression. Paclitaxel treatment significantly increased the mRNA expression of Nav1.1, Nav1.2, Nav1.6 and Nax, but not Nav1.3, sodium channel alpha subunits compared to vehicle-treated animals. Treatment with paclitaxel significantly increased the expression of Navβ1 and Navβ3, but not Navβ2 and Navβ4, sodium channel beta subunits compared to vehicle-treated animals. These findings suggest that during paclitaxel-induced neuropathic pain (PINP) there is differential upregulation of sodium channels in the ACC, which might contribute to the increased neuronal activity observed in the area during neuropathic pain. PMID:27896032

  8. Extracellular signal-regulated kinase mediates gonadotropin subunit gene expression and LH release responses to endogenous gonadotropin-releasing hormones in goldfish.

    Science.gov (United States)

    Klausen, Christian; Booth, Morgan; Habibi, Hamid R; Chang, John P

    2008-08-01

    The possible involvement of extracellular signal-regulated kinase (ERK) in mediating the stimulatory actions of two endogenous goldfish gonadotropin-releasing hormones (salmon (s)GnRH and chicken (c)GnRH-II) on gonadotropin synthesis and secretion was examined. Western blot analysis revealed the presence of ERK and phosphorylated (p)ERK in goldfish brain, pituitary, liver, ovary, testis and muscle tissue extracts, as well as extracts of dispersed goldfish pituitary cells and HeLa cells. Interestingly, a third ERK-like immunoreactive band of higher molecular mass was detected in goldfish tissue and pituitary cell extracts in addition to the ERK1-p44- and ERK2-p42-like immunoreactive bands. Incubation of primary cultures of goldfish pituitary cells with either a PKC-activating 4beta-phorbol ester (TPA) or a synthetic diacylglycerol, but not a 4alpha-phorbol ester, elevated the ratio of pERK/total (t)ERK for all three ERK isoforms. The stimulatory effects of TPA were attenuated by the PKC inhibitor GF109203X and the MEK inhibitor PD98059. sGnRH and cGnRH-II also elevated the ratio of pERK/tERK for all three ERK isoforms, in a time-, dose- and PD98059-dependent manner. In addition, treatment with PD98059 reduced the sGnRH-, cGnRH-II- and TPA-induced increases in gonadotropin subunit mRNA levels in Northern blot studies and sGnRH- and cGnRH-II-elicited LH release in cell column perifusion studies with goldfish pituitary cells. These results indicate that GnRH and PKC can activate ERK through MEK in goldfish pituitary cells. More importantly, the present study suggests that GnRH-induced gonadotropin subunit gene expression and LH release involve MEK/ERK signaling in goldfish.

  9. Alternative-splicing in the exon-10 region of GABA(A receptor beta(2 subunit gene: relationships between novel isoforms and psychotic disorders.

    Directory of Open Access Journals (Sweden)

    Cunyou Zhao

    Full Text Available BACKGROUND: Non-coding single nucleotide polymorphisms (SNPs in GABRB2, the gene for beta(2-subunit of gamma-aminobutyric acid type A (GABA(A receptor, have been associated with schizophrenia (SCZ and quantitatively correlated to mRNA expression and alternative splicing. METHODS AND FINDINGS: Expression of the Exon 10 region of GABRB2 from minigene constructs revealed this region to be an "alternative splicing hotspot" that readily gave rise to differently spliced isoforms depending on intron sequences. This led to a search in human brain cDNA libraries, and the discovery of two novel isoforms, beta(2S1 and beta(2S2, bearing variations in the neighborhood of Exon-10. Quantitative real-time PCR analysis of postmortem brain samples showed increased beta(2S1 expression and decreased beta(2S2 expression in both SCZ and bipolar disorder (BPD compared to controls. Disease-control differences were significantly correlated with SNP rs187269 in BPD males for both beta(2S1 and beta(2S2 expressions, and significantly correlated with SNPs rs2546620 and rs187269 in SCZ males for beta(2S2 expression. Moreover, site-directed mutagenesis indicated that Thr(365, a potential phosphorylation site in Exon-10, played a key role in determining the time profile of the ATP-dependent electrophysiological current run-down. CONCLUSION: This study therefore provided experimental evidence for the importance of non-coding sequences in the Exon-10 region in GABRB2 with respect to beta(2-subunit splicing diversity and the etiologies of SCZ and BPD.

  10. Marketing Optimization for B2B Market

    OpenAIRE

    Kaynova Tatyana V.

    2012-01-01

    The article presents market definition B2B, the necessity to optimize marketing B2B market, provides a system for B2B-marketing and developed stages of its formation. On this basis it was identified key factors of customer loyalty and are the stages of development of loyalty programs for customers market B2B.

  11. New genes encoding subunits of a cytochrome bc1-analogous complex in the respiratory chain of the hyperthermoacidophilic crenarchaeon Sulfolobus acidocaldarius.

    Science.gov (United States)

    Hiller, A; Henninger, T; Schäfer, G; Schmidt, C L

    2003-04-01

    The soxL gene from Sulfolobus acidocaldarius (DSM 639) encodes a Rieske iron-sulfur protein. In this study we report the identification of two open reading frames in its downstream region. The first one, named soxN, codes for a membrane protein bearing a resemblance to the b-type cytochromes of the cytochrome bc1 and b6f complexes. The protein is predicted to contain at least 10 transmembrane helices and features the two conserved histidine pairs coordinating the heme groups of these cytochromes. The second open reading frame, named odsN, encodes a soluble protein of unknown function. The genomic region displays a complex transcription pattern. Northern blot and RT-PCR analyses revealed the presence of mono- and bi-cistronic transcripts as well as a tri-cistronic transcript of soxL and cbsAB, encoding the mono-heme cytochrome b558/566. Phylogenetic analyses of the genes of the soxLN pair and of other archaeal gene pairs encoding Rieske iron-sulfur proteins and b-type cytochromes revealed an identical branching patterns for both protein families, suggesting an evolutionary link of these genes provided by the functional interaction of the proteins. On the basis of the findings of this study and the previously studied properties of the soxL and cbsA proteins, we propose the occurrence of a novel cytochrome bc1-analogous complex in the membranes of Sulfolobus, consisting of the cytochrome b homolog soxN, the Rieske protein soxL, the high potential cytochrome cbsA, as well as the non-redox-active subunits cbsB and odsN.

  12. Gene splicing of an invertebrate beta subunit (LCavβ in the N-terminal and HOOK domains and its regulation of LCav1 and LCav2 calcium channels.

    Directory of Open Access Journals (Sweden)

    Taylor F Dawson

    Full Text Available The accessory beta subunit (Ca(vβ of calcium channels first appear in the same genome as Ca(v1 L-type calcium channels in single-celled coanoflagellates. The complexity of this relationship expanded in vertebrates to include four different possible Ca(vβ subunits (β1, β2, β3, β4 which associate with four Ca(v1 channel isoforms (Ca(v1.1 to Ca(v1.4 and three Ca(v2 channel isoforms (Ca(v2.1 to Ca(v2.3. Here we assess the fundamentally-shared features of the Ca(vβ subunit in an invertebrate model (pond snail Lymnaea stagnalis that bears only three homologous genes: (LCa(v1, LCa(v2, and LCa(vβ. Invertebrate Ca(vβ subunits (in flatworms, snails, squid and honeybees slow the inactivation kinetics of Ca(v2 channels, and they do so with variable N-termini and lacking the canonical palmitoylation residues of the vertebrate β2a subunit. Alternative splicing of exon 7 of the HOOK domain is a primary determinant of a slow inactivation kinetics imparted by the invertebrate LCa(vβ subunit. LCa(vβ will also slow the inactivation kinetics of LCa(v3 T-type channels, but this is likely not physiologically relevant in vivo. Variable N-termini have little influence on the voltage-dependent inactivation kinetics of differing invertebrate Ca(vβ subunits, but the expression pattern of N-terminal splice isoforms appears to be highly tissue specific. Molluscan LCa(vβ subunits have an N-terminal "A" isoform (coded by exons: 1a and 1b that structurally resembles the muscle specific variant of vertebrate β1a subunit, and has a broad mRNA expression profile in brain, heart, muscle and glands. A more variable "B" N-terminus (exon 2 in the exon position of mammalian β3 and has a more brain-centric mRNA expression pattern. Lastly, we suggest that the facilitation of closed-state inactivation (e.g. observed in Ca(v2.2 and Ca(vβ3 subunit combinations is a specialization in vertebrates, because neither snail subunit (LCa(v2 nor LCa(vβ appears to be compatible

  13. Purification of FLAG-tagged eukaryotic initiation factor 2B complexes, subcomplexes, and fragments from Saccharomyces cerevisiae.

    Science.gov (United States)

    Mohammad-Qureshi, Sarah S; Haddad, Raphaël; Palmer, Karren S; Richardson, Jonathan P; Gomez, Edith; Pavitt, Graham D

    2007-01-01

    The eukaryotic initiation factor 2B (eIF2B) is a five-subunit guanine nucleotide exchange factor, that functions during translation initiation to catalyze the otherwise slow exchange of GDP for GTP on its substrate eIF2. Assays to measure substrate interaction and guanine nucleotide release ability of eIF2B require the complex to be purified free of interacting proteins. We have also found that a subcomplex of two subunits, gamma and epsilon or the largest one, epsilon alone, promotes this activity. Within eIF2Bepsilon, the catalytic center requires the C-terminal 200 residues only. Here, we describe our protocols for purifying the Saccharomyces cerevisiae eIF2B complexes and the catalytic subunit using FLAG-tagged proteins overexpressed in yeast cells. Using commercially available FLAG-affinity resin and high salt buffer, we are able to purify active eIF2B virtually free of contaminants.

  14. Role of the Rubisco Small Subunit

    Energy Technology Data Exchange (ETDEWEB)

    Spreitzer, Robert Joseph [Univ. of Nebraska, Lincoln, NE (United States)

    2016-11-05

    Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) catalyzes the rate-limiting step of CO2 fixation in photosynthesis. However, it is a slow enzyme, and O2 competes with CO2 at the active site. Oxygenation initiates the photorespiratory pathway, which also results in the loss of CO2. If carboxylation could be increased or oxygenation decreased, an increase in net CO2 fixation would be realized. Because Rubisco provides the primary means by which carbon enters all life on earth, there is much interest in engineering Rubisco to increase the production of food and renewable energy. Rubisco is located in the chloroplasts of plants, and it is comprised of two subunits. Much is known about the chloroplast-gene-encoded large subunit (rbcL gene), which contains the active site, but much less is known about the role of the nuclear-gene-encoded small subunit in Rubisco function (rbcS gene). Both subunits are coded by multiple genes in plants, which makes genetic engineering difficult. In the eukaryotic, green alga Chlamydomonas reinhardtii, it has been possible to eliminate all the Rubisco genes. These Rubisco-less mutants can be maintained by providing acetate as an alternative carbon source. In this project, focus has been placed on determining whether the small subunit might be a better genetic-engineering target for improving Rubisco. Analysis of a variable-loop structure (βA-βB loop) of the small subunit by genetic selection, directed mutagenesis, and construction of chimeras has shown that the small subunit can influence CO2/O2 specificity. X-ray crystal structures of engineered chimeric-loop enzymes have indicated that additional residues and regions of the small subunit may also contribute to Rubisco function. Structural dynamics of the small-subunit carboxyl terminus was also investigated. Alanine-scanning mutagenesis of the most-conserved small-subunit residues has identified a

  15. Impairment of the tRNA-splicing endonuclease subunit 54 (tsen54) gene causes neurological abnormalities and larval death in zebrafish models of pontocerebellar hypoplasia.

    Science.gov (United States)

    Kasher, Paul R; Namavar, Yasmin; van Tijn, Paula; Fluiter, Kees; Sizarov, Aleksander; Kamermans, Maarten; Grierson, Andrew J; Zivkovic, Danica; Baas, Frank

    2011-04-15

    Pontocerebellar hypoplasia (PCH) represents a group (PCH1-6) of neurodegenerative autosomal recessive disorders characterized by hypoplasia and/or atrophy of the cerebellum, hypoplasia of the ventral pons, progressive microcephaly and variable neocortical atrophy. The majority of PCH2 and PCH4 cases are caused by mutations in the TSEN54 gene; one of the four subunits comprising the tRNA-splicing endonuclease (TSEN) complex. We hypothesized that TSEN54 mutations act through a loss of function mechanism. At 8 weeks of gestation, human TSEN54 is expressed ubiquitously in the brain, yet strong expression is seen within the telencephalon and metencephalon. Comparable expression patterns for tsen54 are observed in zebrafish embryos. Morpholino (MO) knockdown of tsen54 in zebrafish embryos results in loss of structural definition in the brain. This phenotype was partially rescued by co-injecting the MO with human TSEN54 mRNA. A developmental patterning defect was not associated with tsen54 knockdown; however, an increase in cell death within the brain was observed, thus bearing resemblance to PCH pathophysiology. Additionally, N-methyl-N-nitrosourea mutant zebrafish homozygous for a tsen54 premature stop-codon mutation die within 9 days post-fertilization. To determine whether a common disease pathway exists between TSEN54 and other PCH-related genes, we also monitored the effects of mitochondrial arginyl-tRNA synthetase (rars2; PCH1 and PCH6) knockdown in zebrafish. Comparable brain phenotypes were observed following the inhibition of both genes. These data strongly support the hypothesis that TSEN54 mutations cause PCH through a loss of function mechanism. Also we suggest that a common disease pathway may exist between TSEN54- and RARS2-related PCH, which may involve a tRNA processing-related mechanism.

  16. Fifteen novel mutations in the mitochondrial NADH dehydrogenase subunit 1, 2, 3, 4, 4L, 5 and 6 genes from Iranian patients with Leber's hereditary optic neuropathy (LHON).

    Science.gov (United States)

    Rezvani, Zahra; Didari, Elmira; Arastehkani, Ahoura; Ghodsinejad, Vadieh; Aryani, Omid; Kamalidehghan, Behnam; Houshmand, Massoud

    2013-12-01

    Leber's hereditary optic neuropathy (LHON) is an optic nerve dysfunction resulting from mutations in mitochondrial DNA (mtDNA), which is transmitted in a maternal pattern of inheritance. It is caused by three primary point mutations: G11778A, G3460A and T14484C; in the mitochondrial genome. These mutations are sufficient to induce the disease, accounting for the majority of LHON cases, and affect genes that encode for the different subunits of mitochondrial complexes I and III of the mitochondrial respiratory chain. Other mutations are secondary mutations associated with the primary mutations. The purpose of this study was to determine MT-ND variations in Iranian patients with LHON. In order to determine the prevalence and distribution of mitochondrial mutations in the LHON patients, their DNA was studied using PCR and DNA sequencing analysis. Sequencing of MT-ND genes from 35 LHON patients revealed a total of 44 nucleotide variations, in which fifteen novel variations-A14020G, A13663G, C10399T, C4932A, C3893G, C10557A, C12012A, C13934T, G4596A, T12851A, T4539A, T4941A, T13255A, T14353C and del A 4513-were observed in 27 LHON patients. However, eight patients showed no variation in the ND genes. These mutations contribute to the current database of mtDNA polymorphisms in LHON patients and may facilitate the definition of disease-related mutations in human mtDNA. This research may help to understand the disease mechanism and open up new diagnostic opportunities for LHON.

  17. A new sodium channel alpha-subunit gene (Scn9a) from Schwann cells maps to the Scn1a, Scn2a, Scn3a cluster of mouse chromosome 2.

    Science.gov (United States)

    Beckers, M C; Ernst, E; Belcher, S; Howe, J; Levenson, R; Gros, P

    1996-08-15

    We have used a total of 27 AXB/BXA recombinant inbred mouse strains to determine the chromosomal location of a newly identified gene encoding an alpha-subunit isoform of the sodium channel from Schwann cells, Scn9a. Linkage analysis established that Scn9a mapped to the proximal segment of mouse chromosome 2. The segregation of restriction fragment length polymorphisms in 145 progeny from a Mus spretus x C57BL/6J backcross indicates that Scn9a is very tightly linked to Scn1a (gene encoding the type I sodium channel alpha-subunit of the brain) and forms part of a cluster of four Scna genes located on mouse chromosome 2.

  18. A new sodium channel {alpha}-subunit gene (Scn9a) from Schwann cells maps to the Scn1a, Scn2a, Scn3a cluster of mouse chromosome 2

    Energy Technology Data Exchange (ETDEWEB)

    Beckers, M.C.; Ernst, E.; Gros, P. [McGill Univ., Montreal (Canada)

    1996-08-15

    We have used a total of 27 AXB/BXA recombinant inbred mouse strains to determine the chromosomal location of a newly identified gene encoding an {alpha}-subunit isoform of the sodium channel from Schwann cells, Scn9a. Linkage analysis established that Scn9a mapped to the proximal segment of mouse chromosome 2. The segregation of restriction fragment length polymorphisms in 145 progeny from a Mus spretus x C57BL/6J backcross indicates that Scn9a is very tightly linked to Scn1a (gene encoding the type I sodium channel {alpha}-subunit of the brain) and forms part of a cluster of four Scna genes located on mouse chromosome 2. 17 refs., 1 fig., 3 tabs.

  19. A multilevel prediction of physiological response to challenge: Interactions among child maltreatment, neighborhood crime, endothelial nitric oxide synthase gene (eNOS), and GABA(A) receptor subunit alpha-6 gene (GABRA6).

    Science.gov (United States)

    Lynch, Michael; Manly, Jody Todd; Cicchetti, Dante

    2015-11-01

    Physiological response to stress has been linked to a variety of healthy and pathological conditions. The current study conducted a multilevel examination of interactions among environmental toxins (i.e., neighborhood crime and child maltreatment) and specific genetic polymorphisms of the endothelial nitric oxide synthase gene (eNOS) and GABA(A) receptor subunit alpha-6 gene (GABRA6). One hundred eighty-six children were recruited at age 4. The presence or absence of child maltreatment as well as the amount of crime that occurred in their neighborhood during the previous year were determined at that time. At age 9, the children were brought to the lab, where their physiological response to a cognitive challenge (i.e., change in the amplitude of the respiratory sinus arrhythmia) was assessed and DNA samples were collected for subsequent genotyping. The results confirmed that complex Gene × Gene, Environment × Environment, and Gene × Environment interactions were associated with different patterns of respiratory sinus arrhythmia reactivity. The implications for future research and evidence-based intervention are discussed.

  20. Lithium decreased NR2B tyrosine phosphorylation and interactions of NR2B and PSD-95 with Src in rat hippocampus following cerebral ischemia

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    Objective: To study the effects of chronic lithium on N-methyl-d-aspartate receptor subunit 2B (NR2B) tyrosine phosphorylation and the interactions of NR2B and PSD-95 with Src induced by cerebral ischemia/reperfusion (I/R). Methods: Transient (15 min) cerebral ischemia was induced by four-vessel occlusion procedure in SD rats. Immunoprecipitation (IP) and immunoblotting (IB)were performed to investigate the phosphorylation and interactions of proteins. The effects of lithium on tyrosine phosphorylation of NR2B and its interactions with PSD-95 and Src were examined. Results: Transient cerebral ischemia 15 min followed by reperfusion 6 h (I/R 6h) caused a significant increase in tyrosine phosphorylation of NR2B. Administration of LiCl for 7days before ischemia caused a profound decrease in tyrosine phosphorylation of NR2B. Similiarly, the interactions of NR2B and PSD-95 with Src were also enhanced by I/R 6 h.moreover, these interactions were also inhibited by chronic lithium. Conclusion: Pretreatment with lithium decrease tyrosine phosphorylation of NR2B and interactions of NR2B and PSD-95 with Src during cerebral I/R.

  1. Distribution of genotypes C825T polymorphism G-protein β3-subunit gene in patients with hypertension depending on body mass index

    Directory of Open Access Journals (Sweden)

    Prystupa L.N.

    2015-09-01

    Full Text Available The aim of the study was to investigate the frequency of genotypes of C825T polymorphism G-protein β3-subunit gene (GNB3 in patients with arterial hypertension (AH, depending on body mass index (BMI. The study involved 155 patients with verified diagnosis of AH (study group and 50 healthy individuals (control group. The patients of the main group were divided into 3 groups according to BMI: I - 35 patients with normal body weight, II - 38 patients with overweight, III - 82 patients with obesity. We used general clinical, anthropometric, instrumental, molecular-genetic and statistical methods. Probability of differences in the frequency of alleles and genotypes was determined using χ² criteria. Pairwise comparison of groups was made using nonparametric Mann-Whitney test. The difference was considered statistically significant at p <0,05. Investigation of the distribution of genotypes C825T polymorphism GNB3 in patients with AH according to BMI showed statistically significant increase in the frequency of genotypes C / T and T / T and T allele in patients with overweight and obesity as compared with patients with normal body weight (χ² = 26 8; p <0.001. The risk of weight increase in AH patients with T allele carriers is 2,2 times higher than in C allele carriers. Association of C825T polymorphism of GNB3 with a tendency to obesity and overweight in patients with AH was proved.

  2. Genetic structure of the snakehead murrel, Channa striata (channidae) based on the cytochrome c oxidase subunit I gene: Influence of historical and geomorphological factors.

    Science.gov (United States)

    Jamsari, Amirul Firdaus Jamaluddin; Jamaluddin, Jamsari Amirul Firdaus; Pau, Tan Min; Siti-Azizah, Mohd Nor

    2011-01-01

    Nucleotide sequences of a partial cytochrome c oxidase subunit I gene were used to assess the manner in which historical processes and geomorphological effects may have influenced genetic structuring and phylogeographic patterns in Channa striata. Assaying was based on individuals from twelve populations in four river systems, which were separated into two regions, the eastern and western, of the biodiversely rich state of Perak in central Peninsular Malaysia. In 238 specimens, a total of 368-bp sequences with ten polymorphic sites and eleven unique haplotypes were detected. Data on all the twelve populations revealed incomplete divergence due to past historical coalescence and the short period of separation. Nevertheless, SAMOVA and F(ST) revealed geographical structuring existed to a certain extent in both regions. For the eastern region, the data also showed that the upstream populations were genetically significantly different compared to the mid- and downstream ones. It is inferred that physical barriers and historical processes played a dominant role in structuring the genetic dispersal of the species. A further inference is that the Grik, Tanjung Rambutan and Sungkai are potential candidates for conservation and aquaculture programmes since they contained most of the total diversity in this area.

  3. Genetic structure of the snakehead murrel, Channa striata (channidae based on the cytochrome c oxidase subunit I gene: influence of historical and geomorphological factors

    Directory of Open Access Journals (Sweden)

    Jamsari Amirul Firdaus Jamaluddin

    2011-01-01

    Full Text Available Nucleotide sequences of a partial cytochrome c oxidase subunit I gene were used to assess the manner in which historical processes and geomorphological effects may have influenced genetic structuring and phylogeographic patterns in Channa striata. Assaying was based on individuals from twelve populations in four river systems, which were separated into two regions, the eastern and western, of the biodiversely rich state of Perak in central Peninsular Malaysia. In 238 specimens, a total of 368-bp sequences with ten polymorphic sites and eleven unique haplotypes were detected. Data on all the twelve populations revealed incomplete divergence due to past historical coalescence and the short period of separation. Nevertheless, SAMOVA and F ST revealed geographical structuring existed to a certain extent in both regions. For the eastern region, the data also showed that the upstream populations were genetically significantly different compared to the mid- and downstream ones. It is inferred that physical barriers and historical processes played a dominant role in structuring the genetic dispersal of the species. A further inference is that the Grik, Tanjung Rambutan and Sungkai are potential candidates for conservation and aquaculture programmes since they contained most of the total diversity in this area.

  4. Correlation of oxygen consumption, cytochrome c oxidase, and cytochrome c oxidase subunit I gene expression in the termination of larval diapause in the bamboo borer, Omphisa fuscidentalis.

    Science.gov (United States)

    Singtripop, Tippawan; Saeangsakda, Manasawan; Tatun, Nujira; Kaneko, Yu; Sakurai, Sho

    2007-09-01

    The moth Omphisa fuscidentalis (Lepidoptera, Pyralidae) is a univoltine insect with a larval diapause period lasting up to 9 months. We studied changes in O(2) consumption in conjunction with cytochrome c oxidase activity and cytochrome c oxidase subunit I (cox1) gene expression. O(2) consumption changed within a day, showing a supradian rhythm with a ca.12-h cycle at 25 degrees C. During the first two-thirds of the diapause period, from October to March, O(2) consumption was constant until January and then increased by March. Topical application of methoprene, a juvenile hormone analog (JHA), to diapausing larvae terminated the diapause and was associated with an increase in O(2) consumption rate at diapause termination. In JHA-treated larvae, cytochrome c oxidase activity in fat bodies was high at the beginning of the prepupal period and highest at pupation. cox1 expression in fat bodies displayed a transient peak 8 days after JHA application and peaked in the prepupal period. Taken together, our results show that the break of diapause by JHA is associated with the activation of cox1, bringing about an increase in cytochrome c oxidase activity, followed by an increase in O(2) consumption rate.

  5. The effect of high glucose levels on the hypermethylation of protein phosphatase 1 regulatory subunit 3C (PPP1R3C) gene in colorectal cancer

    Indian Academy of Sciences (India)

    Soo Kyung Lee; Ji Wook Moon; Yong Woo Lee; Jung Ok Lee; Su Jin Kim; Nami Kim; Jin Kim; Hyeon Soo Kim; Sun-Hwa Park

    2015-03-01

    DNA methylation is an epigenetic event that occurs frequently in colorectal cancer (CRC). Increased glucose level is a strong risk factor for CRC. Protein phosphatase 1 regulatory subunit 3C (PPP1R3C) modulates glycogen metabolism, particularly glycogen synthesis. The aim of this study was to investigate the effect of high glucose levels on DNA methylation of PPP1R3C in CRC. PPP1R3C was significantly hypermethylated in CRC tissues (76/105, 72.38%, < 0.05) and colon cancer cell lines ( < 0.05). CRC tissues obtained from patients with high glucose levels showed that the methylation of PPP1R3C was lower than in patients who had normal levels of glucose. When DLD-1 cells were cultured under conditions of high glucose, the methylation of PPP1R3C was repressed. The expression of PPP1R3C was inversely related to methylation status. In addition, a promoter luciferase assay showed that the transcriptional activity of PPP1R3C was increased in high glucose culture conditions. The number of cells decreased when PPP1R3C was silenced in DLD-1 cells. These results suggest that PPP1R3C, a novel hypermethylated gene in CRC, may play a critical role in cancer cell growth in association with glucose levels.

  6. Expression of the gene for large subunit of m-calpain is elevated in skeletal muscle from Duchenne muscular dystrophy patients

    Indian Academy of Sciences (India)

    Tajamul Hussain; Harleen Mangath; C. Sundaram; M. P. J. S. Anandaraj

    2000-08-01

    Calpain is an intracellular nonlysosomal protease involved in essential regulatory or processing functions of the cell, mediated by physiological concentrations of Ca2+. However, in an environment of abnormal intracellular calcium, such as that seen in Duchenne muscular dystrophy (DMD), calpain is suggested to cause degeneration of muscle owing to enhanced activity. To test whether the reported increase in calpain activity in DMD results from de novo synthesis of the protease, we have assessed the quantitative changes in mRNA specific for m-calpain. mRNA isolated from DMD and control muscle was analysed by dot blot hybridization using a cDNA probe for the large subunit of m-calpain. Compared to control a four-fold increase in specific mRNAwas observed in dystrophic muscle. This enhanced expression of the m-calpain gene in dystrophic condition suggests that the reported increase in m-calpain activity results from de novo synthesis of protease and underlines the important role of m-calpain in DMD.

  7. Reconsideration of the phylogenetic positions of five peritrich genera, Vorticella, Pseudovorticella, Zoothamnopsis, Zoothamnium, and Epicarchesium (Ciliophora, Peritrichia, Sessilida), based on small subunit rRNA gene sequences.

    Science.gov (United States)

    Li, Lifang; Song, Weibo; Warren, Alan; Shin, Mann Kyoon; Chen, Zigui; Ji, Daode; Sun, Ping

    2008-01-01

    In order to re-evaluate the systematics of sessilid peritrich ciliates, small subunit (SSU) rRNA gene sequences were determined for 12 species belonging to five genera: Vorticella, Pseudovorticella, Epicarchesium, Zoothamnium, and Zoothamnopsis. Phylogenetic trees were deduced using Bayesian inference, maximum parsimony, and maximum likelihood methods. The phylogenetic analyses suggest that (1) sessilids which have stalks with continuous myonemes that contract in a zig-zag fashion form a separate clade from those which have stalks that contract independently and in a spiral fashion, supporting the separation of the family Zoothamniidae from the family Vorticellidae and (2) Epicarchesium and Pseudovorticella, both of which have reticulate silverline systems, are more closely related to each other than to other vorticellids, suggesting that differences in the silverline system (i.e. transverse vs. reticulate) may be the result of genuine evolutionary divergence among sessilid peritrichs. However, the newly sequenced Zoothamnopsis sinica, which has a reticulate silverline pattern, nests within the unresolved Zoothamnium species that have transverse silverline patterns. Thus, there were at least two evolutions of the reticulate silverline pattern character state from a plesiomorphic transverse state in the peritrichid ciliates. The molecular work demonstrates the genus Zoothamnium to be paraphyletic in relation to morphological studies, and suggests that Astylozoon, Opisthonecta, and Vorticella microstoma possibly share a SSU rRNA secondary structure in the helix E10-1 region.

  8. [DISTRIBUTION OF GENOTYPES OF C825T POLYMORPHISM β3-SUBUNIT G-PROTEIN GENE IN PATIENTS WITH ARTERIAL HYPERTENSION ACCORDING THE DEGREE OF OBESITY].

    Science.gov (United States)

    Moiseyenko, I; Prystupa, L; Garbuzova, V; Pogorielova, O; Opolonskaya, N

    2015-01-01

    Arterial hypertension (AH) and obesity - risk factors for cardiovascular diseases and their complications, leading to high morbidity and mortality. These nosologies notedly linked, because have common etiological factors, pathophysiological mechanisms and genetic determination. The aim this research was to analyze the distribution of genotypes of the C825T polymorphism of β3-subunit G-protein gene (GNB3) according the degree of obesity and to assess the risk of obesity in patients with AH. Patients were divided into three groups according the degree of obesity. We used clinical, anthropometric, instrumental, molecular-genetic and statistical methods. The significance of differences of alleles and genotypes frequency was determined by test χ². For comparing the groups used nonparametric Mann-Whitney and Kruskal-Wallis tests. A value of phypertension and obesity (χ² = 27,976, p obesity. The risk of obesity in T allele carriers was in 2.2 times higher than in C allele carriers in patients with AH. In summary, our study showed association of C825T polymorphism of the GNB3 with obesity, but did not prove the association this with the degree of obesity i patients with AH.

  9. Proliferation of transformed somatotroph cells related to low or absent expression of protein kinase a regulatory subunit 1A protein.

    Science.gov (United States)

    Lania, Andrea G; Mantovani, Giovanna; Ferrero, Stefano; Pellegrini, Caterina; Bondioni, Sara; Peverelli, Erika; Braidotti, Paola; Locatelli, Marco; Zavanone, Mario L; Ferrante, Emanuela; Bosari, Silvano; Beck-Peccoz, Paolo; Spada, Anna

    2004-12-15

    The two regulatory subunits (R1 and R2) of protein kinase A (PKA) are differentially expressed in cancer cell lines and exert diverse roles in growth control. Recently, mutations of the PKA regulatory subunit 1A gene (PRKAR1A) have been identified in patients with Carney complex. The aim of this study was to evaluate the expression of the PKA regulatory subunits R1A, R2A, and R2B in a series of 30 pituitary adenomas and the effects of subunit activation on cell proliferation. In these tumors, neither mutation of PRKAR1A nor loss of heterozygosity was identified. By real-time PCR, mRNA of the three subunits was detected in all of the tumors, R1A being the most represented in the majority of samples. By contrast, immunohistochemistry documented low or absent R1A levels in all tumors, whereas R2A and R2B were highly expressed, thus resulting in an unbalanced R1/R2 ratio. The low levels of R1A were, at least in part, due to proteasome-mediated degradation. The effect of the R1/R2 ratio on proliferation was assessed in GH3 cells, which showed a similar unbalanced pattern of R subunits expression, and in growth hormone-secreting adenomas. The R2-selective cAMP analog 8-Cl cAMP and R1A RNA silencing, stimulated cell proliferation and increased Cyclin D1 expression, respectively, in human and rat adenomatous somatotrophs. These data show that a low R1/R2 ratio promoted proliferation of transformed somatotrophs and are consistent with the Carney complex model in which R1A inactivating mutations further unbalance this ratio in favor of R2 subunits. These results suggest that low expression of R1A protein may favor cAMP-dependent proliferation of transformed somatotrophs.

  10. Infantile onset Vanishing White Matter disease associated with a novel EIF2B5 variant, remarkably long life span, severe epilepsy, and hypopituitarism.

    Science.gov (United States)

    Woody, April L; Hsieh, David T; McIver, Harkirtin K; Thomas, Linda P; Rohena, Luis

    2015-04-01

    Vanishing White Matter disease (VWM) is an inherited progressive leukoencephalopathy caused by mutations in the genes EIF2B1-5, which encode for the 5 subunits of the eukaryotic initiation factor 2B (eIF2B), a regulator of protein synthesis. VWM typically presents with acute neurological decline following febrile infections or minor head trauma, and subsequent progressive neurological and cognitive regression. There is a varied clinical spectrum of VWM, with earlier onset associated with more severe phenotypes. Brain magnetic resonance imaging is usually diagnostic with diffusely abnormal white matter, progressing over time to cystic degeneration. We are reporting on a patient with infantile onset VWM associated with three heterozygous missense variants in EIF2B5, including a novel missense variant on exon 6 of EIF2B5 (D262N), as well as an interstitial duplication at 7q21.12. In addition, our case is unusual because of a severe epilepsy course, a novel clinical finding of hypopituitarism manifested by hypothyroidism and adrenal insufficiency, and a prolonged life span with current age of survival of 4 years and 11 months.

  11. The roles of Na⁺/K⁺-ATPase α-subunit gene from the ridgetail white prawn Exopalaemon carinicauda in response to salinity stresses.

    Science.gov (United States)

    Li, Jitao; Ma, Peng; Liu, Ping; Chen, Ping; Li, Jian

    2015-02-01

    Na(+)/K(+)-ATPase (NAK) is one important transporter protein and plays a key role in maintaining osmotic homeostasis in low and high salinity acclimation in variety of crustacean species. The ridgetail white prawn Exopalaemon carinicauda is an euryhaline and economic shrimp species in China, but it remains unclear about its mechanism of salinity adaption. In this study, a full-length of Na(+)/K(+)-ATPase α-subunit (α-NAK) cDNA was cloned from E. carinicauda by using rapid amplification of cDNA ends (RACE) approaches. The full-length cDNA of α-NAK was of 3680 bp, containing an open reading frame (ORF) of 3030 bp encoding a polypeptide of 1009 amino acids with the predicted molecular weight of 112.27 kDa. Eight transmembrane domains and two sites of phosphorylation and ATP binding were identified in E. carinicauda α-NAK. BLAST analysis revealed that the sequence of α-NAK amino acids of E. carinicauda shared more than 75% homologies with those of other crustacean. Real time quantitative RT-PCR analysis indicated that E. carinicauda α-NAK gene could be detected in all the tested tissues with highest expression level in gill. The expression profiles of E. carinicauda α-NAK transcripts were analyzed in gill and hepatopancreas tissues after salinity stresses. The results showed that the expression level of E. carinicauda α-NAK gene in both gill and hepatopancreas reached peak at different time after low and high salinity stresses, and showed different expression profiles. The expression profiles of proPO transcripts in gills after salinity stresses also indicated α-NAK and proPO played synergistic actions for salinity responses in E. carinicauda. These results indicated that E. carinicauda α-NAK involved in stress responses against salinity.

  12. GATAD2B loss-of-function mutations cause a recognisable syndrome with intellectual disability and are associated with learning deficits and synaptic undergrowth in Drosophila

    NARCIS (Netherlands)

    Willemsen, M.H.; Nijhof, B.; Fenckova, M.; Nillesen, W.M.; Bongers, E.M.H.F.; Castells-Nobau, A.; Asztalos, L.; Viragh, E.; Bon, B.W.M. van; Tezel, E.; Veltman, J.A.; Brunner, H.G.; Vries, B.B. de; Ligt, J. de; Yntema, H.G.; Bokhoven, H. van; Isidor, B.; Caignec, C. Le; Lorino, E.; Asztalos, Z.; Koolen, D.A.; Vissers, L.E.L.M.; Schenck, A.; Kleefstra, T.

    2013-01-01

    BACKGROUND: GATA zinc finger domain containing 2B (GATAD2B) encodes a subunit of the MeCP1-Mi-2/nucleosome remodelling and deacetylase complex involved in chromatin modification and regulation of transcription. We recently identified two de novo loss-of-function mutations in GATAD2B by whole exome s

  13. [Molecular cloning of the DNA sequence of activin beta A subunit gene mature peptides from panda and related species and its application in the research of phylogeny and taxonomy].

    Science.gov (United States)

    Wang, Xiao-Jing; Wang, Xiao-Xing; Wang, Ya-Jun; Wang, Xi-Zhong; He, Guang-Xin; Chen, Hong-Wei; Fei, Li-Song

    2002-09-01

    Activin, which is included in the transforming growth factor-beta (TGF beta) superfamily of proteins and receptors, is known to have broad-ranging effects in the creatures. The mature peptide of beta A subunit of this gene, one of the most highly conserved sequence, can elevate the basal secretion of follicle-stimulating hormone (FSH) in the pituitary and FSH is pivotal to organism's reproduction. Reproduction block is one of the main reasons which cause giant panda to extinct. The sequence of Activin beta A subunit gene mature peptides has been successfully amplified from giant panda, red panda and malayan sun bear's genomic DNA by using polymerase chain reaction (PCR) with a pair of degenerate primers. The PCR products were cloned into the vector pBlueScript+ of Esherichia coli. Sequence analysis of Activin beta A subunit gene mature peptides shows that the length of this gene segment is the same (359 bp) and there is no intron in all three species. The sequence encodes a peptide of 119 amino acid residues. The homology comparison demonstrates 93.9% DNA homology and 99% homology in amino acid among these three species. Both GenBank blast search result and restriction enzyme map reveal that the sequences of Activin beta A subunit gene mature peptides of different species are highly conserved during the evolution process. Phylogeny analysis is performed with PHYLIP software package. A consistent phylogeny tree has been drawn with three different methods. The software analysis outcome accords with the academic view that giant panda has a closer relationship to the malayan sun bear than the red panda. Giant panda should be grouped into the bear family (Uersidae) with the malayan sun bear. As to the red panda, it would be better that this animal be grouped into the unique family (red panda family) because of great difference between the red panda and the bears (Uersidae).

  14. Somatic mutations in ATP1A1 and ATP2B3 lead to aldosterone-producing adenomas and secondary hypertension

    DEFF Research Database (Denmark)

    Beuschlein, Felix; Boulkroun, Sheerazed; Osswald, Andrea

    2013-01-01

    Primary aldosteronism is the most prevalent form of secondary hypertension. To explore molecular mechanisms of autonomous aldosterone secretion, we performed exome sequencing of aldosterone-producing adenomas (APAs). We identified somatic hotspot mutations in the ATP1A1 (encoding an Na+/K+ ATPase α...... subunit) and ATP2B3 (encoding a Ca2+ ATPase) genes in three and two of the nine APAs, respectively. These ATPases are expressed in adrenal cells and control sodium, potassium and calcium ion homeostasis. Functional in vitro studies of ATP1A1 mutants showed loss of pump activity and strongly reduced...... affinity for potassium. Electrophysiological ex vivo studies on primary adrenal adenoma cells provided further evidence for inappropriate depolarization of cells with ATPase alterations. In a collection of 308 APAs, we found 16 (5.2%) somatic mutations in ATP1A1 and 5 (1.6%) in ATP2B3. Mutation...

  15. GRIN2B predicts attention problems among disadvantaged children.

    Science.gov (United States)

    Riva, Valentina; Battaglia, Marco; Nobile, Maria; Cattaneo, Francesca; Lazazzera, Claudio; Mascheretti, Sara; Giorda, Roberto; Mérette, Chantal; Émond, Claudia; Maziade, Michel; Marino, Cecilia

    2015-07-01

    It is well established that adversities and GRIN2B (coding an N-methyl-D-aspartate receptor subunit) are independently associated with behavioral and cognitive impairments in childhood. However, a high proportion of children exposed to adversities have good, long-term outcomes. We hypothesized that among children exposed to adversities, GRIN2B variants would predict the worst cognitive and behavioral outcomes. 6 single nucleotide polymorphisms of GRIN2B were genotyped in 625 children aged 6-11 years from an Italian community-based sample. The interacting effect of GRIN2B variants with 4 measures of adversities [low socioeconomic status (SES), preterm delivery, maternal smoking during pregnancy, and absence of breastfeeding] was investigated upon blindly assessed cognitive abilities (vocabulary, block design, digit spans of Wechsler's Intelligence Scale, and Rey complex figure) and parents-rated behavioral problems (Child Behavior Checklist/6-18). Rs2268119 × SES interaction (Hotelling's Trace = 0.07; F(12,1154) = 3.53; p = 0.00004) influenced behavior, with more attention problems among children in the 'either A/T or T/T genotype and low SES' group, compared to all other groups. This interaction effect was not significant in an independent, replication sample of 475 subjects from an Italian community-based sample. GRIN2B variants predict children with the worst outcome in attention functioning among children exposed to low SES. Our findings, if replicated, could help in the identification of children with the highest risk and may prompt cost-effective preventive/treatment strategies.

  16. Expression of zebrafish nos2b surrounds oral cavity.

    Science.gov (United States)

    Poon, Kar-Lai; Richardson, Michael; Korzh, Vladimir

    2008-06-01

    Inducible nitric oxide synthase (NOS2) catalyzes the production of nitric oxide (NO), and is one of the factors establishing innate immunity. In zebrafish, Nos2 is represented by nos2a and nos2b. Here, we report the cloning and expression pattern of the zebrafish nos2b gene, which does not seem to participate in induced immune response. nos2b was mapped to zebrafish linkage group 15. The spatial and temporal expression pattern of nos2b in embryonic zebrafish was analyzed by whole-mount in situ hybridization. nos2b is expressed constitutively in two primordia located along the ventral midline. The first group of cells contributes to the neurohypophysis. Initially at the level of the ventral hindbrain, the second group of cells migrates closely with the thyroid primordium to its final position at the basihyal by 3 dpf. Thus, the analysis of expression pattern of nos2b reveals complex morphogenetic movements resulting in its expression surrounding the oral cavity.

  17. Implication of human UGT2B7, 2B15 and 2B17 in 19-norandrosterone metabolism

    OpenAIRE

    Emmanuel eStrahm; Ulf eSjöberg; Mats eGarle; Anders eRane; Lena eEkström

    2013-01-01

    Nandrolone (19-nortestosterone) is an anabolic androgenic steroid commonly abused for doping purposes. Nandrolone is mainly metabolized in the liver into 19-norandrosterone prior to glucuronidation and excretion through urine over an extended period of time. Several UGTs (i.e. UGT2B7, UGT2B15 and UGT2B17) are thought to be the major enzymes responsible for conjugation of androgens in human. An in-vitro study using recombinant enzymes expressed in insect cells showed that UGT1A4 and UGT2B7 are...

  18. Phylogenetic position of Linguatula arctica and Linguatula serrata (Pentastomida) as inferred from the nuclear 18S rRNA gene and the mitochondrial cytochrome c oxidase subunit I gene.

    Science.gov (United States)

    Gjerde, Bjørn

    2013-10-01

    Genomic DNA was isolated from a Linguatula serrata female expelled from a dog imported to Norway from Romania and from four Linguatula arctica females collected from semi-domesticated reindeer from northern Norway and subjected to PCR amplification of the complete nuclear 18S rRNA gene and a 1,045-bp portion of the mitochondrial cytochrome c oxidase subunit I gene (cox1). The two species differed at two of 1,830 nucleotide positions (99.9% identity) of the complete 18S rRNA gene sequences and at 102 of 1,045 nucleotide positions (90.2% identity) of the partial cox1 sequences. The four isolates of L. arctica showed no genetic variation in either gene. The new cox1 primers may facilitate the diagnosis of various developmental stages of L. arctica and L. serrata in their hosts. In separate phylogenetic analyses using the maximum likelihood method on sequence data from either gene, L. arctica and L. serrata clustered with members of the order Cephalobaenida rather than with members of the order Porocephalida, in which the genus Linguatula is currently placed based on morphological characters. The phylogenetic relationship of L. arctica, L. serrata and other pentastomids to other metazoan groups could not be clearly resolved, but the pentastomids did not seem to have a sister relationship to crustaceans of the subclass Branchiura as found in other studies. A more extensive taxon sampling, including molecular characterisation of more pentastomid taxa across different genera, seems to be necessary in order to estimate the true relationship of the Pentastomida to other metazoan groups.

  19. G-protein beta3 subunit gene variant is unlikely to have a significant influence on serum uric acid level in Japanese workers.

    Science.gov (United States)

    Suwazono, Yasushi; Kobayashi, Etsuko; Uetani, Mirei; Miura, Katsuyuki; Morikawa, Yuko; Ishizaki, Masao; Kido, Teruhiko; Nakagawa, Hideaki; Nogawa, Koji

    2006-06-01

    The C825T variant of the G-protein beta3 subunit (GNB3) gene has attracted renewed attention as a candidate gene for obesity, hypertension and hyperuricemia. The main role of G-protein is to translate signals from the cell surface into a cellular response. The 825T allele is associated with a splice variant of GNB3 protein and enhanced G-protein activation. We examined the relationship between this variant and the risk of hyperuricemia in Japanese workers. The study subjects were 1,452 men and 1,169 women selected from 3,834 men and 2,591 women in 1997. On the basis of common clinical criteria, hyperuricemia I was defined as serum uric acid >or= 7.0 mg/dl in men and 6.0 mg/dl in women or taking antihyperuricemic medication. The hyperuricemia I group consisted of 186 men and 20 women and its control of 1,266 men and 1,149 women. Hyperuricemia II was defined as serum uric acid > 5.7 mg/dl (median) in men and 3.9 mg/dl (median) in women or taking antihyperuricemic medication. The hyperuricemic II group consisted of 684 men and 570 women and its control of 768 men and 599 women. To replicate previous significant results in young Caucasian men, we selected these criteria because the authors of the study in young Caucasian men adopted the median in their subjects as a cut-off. The statistical power was estimated as 99% based on the significant results in Caucasians. Genotype and allele distributions in men and women with hyperuricemia I and II were not significantly different from those in the corresponding control groups. Logistic regression analysis on hyperuricemia I and II, and multiple regression on serum uric acid level demonstrated no significant effect of the C825T genotype. Despite the sufficient statistical power, this study could not demonstrate the significant influence of C825T on hyperuricemia or serum uric acid. The targeting of this polymorphism is unlikely to be beneficial in the prevention of hyperuricemia in the general Japanese population.

  20. GATAD2B loss-of-function mutations cause a recognisable syndrome with intellectual disability and are associated with learning deficits and synaptic undergrowth in Drosophila

    OpenAIRE

    Willemsen, M. H.; Nijhof, B.; Fenckova, M.; Nillesen, W.M.; Bongers, E M H F; Castells-Nobau, A.; Asztalos, L.; Viragh, E.; Bon, B.W.M. van; Tezel, E.; Veltman, J.A.; Brunner, H G; de Vries, B B; de Ligt, J.; Yntema, H.G.

    2013-01-01

    BACKGROUND: GATA zinc finger domain containing 2B (GATAD2B) encodes a subunit of the MeCP1-Mi-2/nucleosome remodelling and deacetylase complex involved in chromatin modification and regulation of transcription. We recently identified two de novo loss-of-function mutations in GATAD2B by whole exome sequencing in two unrelated individuals with severe intellectual disability. METHODS: To identify additional individuals with GATAD2B aberrations, we searched for microdeletions overlapping with GAT...

  1. Cloning and activity analysis of in vitro expression of plant NAD-IDH genes

    Institute of Scientific and Technical Information of China (English)

    CHEN Defu; CHEN Xiwen

    2004-01-01

    The plant NAD+-dependent isocitrate dehydrogenase (NAD-IDH) is an important multifunctional enzyme. The cDNAs encoding three different subunits of the NAD-IDH were cloned successfully from leaves of Arabidopsis thaliana ecotype Columbia gl1 and three lines (Shaan 2A, Shaan 2B and Ken C1) of Brassica napus by RT-PCR method. By searching the sequences in GenBank Database, it is shown that these sequences from B. napus were novel. Their encoding regions of a functional protein were then inserted into the multiple cloning sites of pMID1 vector for the expression of recombinant protein. All the recombinants were successfully expressed in the in vitro expression system. When the transcripts of subunits 0, 1 and 2 were added together to the in vitro system with 36 μg/mL of protein disulfide isomerase, the expressed products had NAD-IDH enzymatic activity. Comparison of different kinds of gene and molecular ratio of the transcripts showed that the NAD-IDH enzyme is composed of three subunits designated subunit 0, subunit 1 and subunit 2. All the three subunits are essential to catalytic activity. Missing subunit 0 could abolish activity. Missing either subunit 1 or subunit 2 would cause severe impact on activity. Deletions, which would cause frameshift mutation or nonsense mutation, were also found in some transcripts of subunit 1 gene from Shaan 2A and Shaan 2B of B. napus. The mutated subunit 1 lost its proper function. This may explain why there is difference of the NAD-IDH activity among three lines of B. napus.

  2. Pharmacogenetics of Cytochrome P450 2B6 (CYP2B6: Advances on Polymorphisms, Mechanisms, and Clinical Relevance

    Directory of Open Access Journals (Sweden)

    Ulrich M Zanger

    2013-03-01

    Full Text Available Cytochrome P450 2B6 (CYP2B6 belongs to the minor drug metabolizing P450s in human liver. Expression is highly variable both between individuals and within individuals, owing to nongenetic factors, genetic polymorphisms, inducibility and irreversible inhibition by many compounds. Drugs metabolized mainly by CYP2B6 include artemisinin, bupropion, cyclophosphamide, efavirenz, ketamine, and methadone. CYP2B6 is one of the most polymorphic CYP genes in humans and variants have been shown to affect transcriptional regulation, splicing, mRNA and protein expression, and catalytic activity. Some variants appear to affect several functional levels simultaneously, thus, combined in haplotypes, leading to complex interactions between substrate-dependent and -independent mechanisms. The most common functionally deficient allele is CYP2B6*6 [Q172H, K262R], which occurs at frequencies of 15 to over 60% in different populations. The allele leads to lower expression in liver due to erroneous splicing. Recent investigations suggest that the amino acid changes contribute complex substrate-dependent effects at the activity level, although data from recombinant systems used by different researchers are not well in agreement with each other. Another important variant, CYP2B6*18 [I328T], occurs predominantly in Africans (4 to 12% and does not express functional protein. A large number of uncharacterized variants are currently emerging from different ethnicities in the course of the 1000 Genomes Project. The CYP2B6 polymorphism is clinically relevant for HIV-infected patients treated with the reverse transcriptase inhibitor efavirenz, but it is increasingly being recognized for other drug substrates. This review summarizes recent advances on the functional and clinical significance of CYP2B6 and its genetic polymorphism, with particular emphasis on the comparison of kinetic data obtained with different substrates for variants expressed in different recombinant

  3. Fusion of the Tumor-Suppressor Gene CHEK2 and the Gene for the Regulatory Subunit B of Protein Phosphatase 2 PPP2R2A in Childhood Teratoma

    Directory of Open Access Journals (Sweden)

    Yuesheng Jin

    2006-05-01

    Full Text Available We characterized the molecular genetic consequences of a balanced chromosome translocation t(8;22(p21; q12, which occurred as the sole cytogenetic aberration in short-term cultured cells from an intrathoracic mature teratoma in a 15-year-old girl. Fluorescence in situ hybridization and reverse transcription- polymerase chain reaction disclosed that t(8;22 resulted in the fusion of the genes PPP2R2A and CHEK2, with an inserted fragment belonging to class I endogenous retrovirus-related sequences at the junction. Sequencing of the two genes did not reveal any additional mutation. None of the three detected PPP2R2A/CHEK2 fusion transcripts resulted in an in-frame PPP2R2A/CHEK2 chimerical open reading frame; however, in all of them, the known open reading frame of CHEK2 was preserved. Thus, promoter swapping leading to deregulated CHEK2 expression would be the most likely oncogenic mechanism. Whereas inactivating mutations of CHEK2 previously have been described in a variety of sporadic tumors and in inherited cancer-predisposing syndromes, PPP2R2A, encoding a regulatory subunit of the multimeric enzyme phosphatase 2, has not been directly implicated in tumorigenesis. Our findings suggest that deregulation of CHEK2 and/or PPP2R2A is of pathogenetic importance in at least a subset of germ cell tumors.

  4. Perilla frutescens leaf extract inhibits mite major allergen Der p 2-induced gene expression of pro-allergic and pro-inflammatory cytokines in human bronchial epithelial cell BEAS-2B.

    Directory of Open Access Journals (Sweden)

    Jer-Yuh Liu

    Full Text Available Perilla frutescens has been used in traditional medicine for respiratory diseases due to its anti-bacterial and anti-inflammatory activity. This study aimed to investigate effects of Perilla frutescens leaf extract (PFE on expression of pro-allergic and pro-inflammatory cytokines in airway epithelial cells exposed to mite major allergen Der p 2 (DP2 and the underlying mechanisms. Our results showed that PFE up to 100 µg/mL had no cytotoxic effect on human bronchial epithelial cell BEAS-2B. Further investigations revealed that PFE dose-dependently diminished mRNA expression of pro-allergic cytokine IL-4, IL-5, IL-13 and GM-CSF, as well as pro-inflammatory cytokine IL-6, IL-8 and MCP-1 in BEAS-2B cells treated with DP2. In parallel to mRNA, the DP-2-elevated levels of the tested cytokines were decreased. Further investigation showed that DP2-indued phosphorylation of p38 MAPK (P38 and JNK, but not Erk1/2, was also suppressed by PFE. In addition, PFE elevated cytosolic IκBα level and decreased nuclear NF-κB level in DP2-stimulated BEAS-2B cells. Taken together, these findings revealed that PFE significantly diminished both mRNA expression and protein levels of pro-allergic and pro-inflammatory cytokines in response to DP2 through inhibition of P38/JNK and NK-κB activation. These findings suggest that PFE should be beneficial to alleviate both allergic and inflammatory responses on airway epithelium in response to aeroallergens.

  5. Substrate Recognition of Histone H2B by DUBm

    Science.gov (United States)

    Henderson, Elizabeth; Berndsen, Christopher; Wolberger, Cynthia

    2011-03-01

    The SAGA complex is a transcriptional coactivator that regulates gene expression in eukaryotes via histone acetylation and deubiquitination, which are crucial for transcription. Our lab is investigating the SAGA-dependent deubiquitination of histone H2B. The deubiquitinating module (DUBm) of SAGA is comprised of a ubiquitin-specific protease, Ubp8, and three other proteins. It is known that Ubp8 cleaves ubiquitin from histone H2B, however, the specific way in which the enzyme binds to the substrate remains elusive. In order to unravel this mechanism, we attempted to determine the crystal structure of the substrate binding complex. We obtained this substrate by exploiting the techniques of intein chemistry to artificially ubiquitinate a histone H2B peptide, which we then co-crystallized with DUBm. Additionally, we synthesized Ub-K63R-linked chains and Ub-K48-linked chains and co-crystallized them with DUBm.

  6. Two thyroid hormone regulated genes, the beta-subunits of nerve growth factor (NGFB) and thyroid stimulating hormone (TSHB), are located less than 310 kb apart in both human and mouse genomes.

    Science.gov (United States)

    Dracopoli, N C; Rose, E; Whitfield, G K; Guidon, P T; Bale, S J; Chance, P A; Kourides, I A; Housman, D E

    1988-08-01

    Two thyroid hormone regulated genes, the beta-subunits of nerve growth factor (NGFB) and thyroid stimulating hormone (TSHB), have been assigned to mouse chromosome 3 and human chromosome 1p22. We have used the techniques of linkage analysis and pulsed field gel electrophoresis to determine the proximity of these two antithetically regulated genes in this conserved linkage group. Four novel restriction fragment length polymorphisms were identified at the human TSHB gene. Two-point linkage analysis between TSHB and NGFB in 46 families, including the Centre d'Etude du Polymorphisme Humain (CEPH) reference panel, demonstrated no recombination (theta = 0.00, Z = 42.8). Analysis of this region by pulsed field gel electrophoresis showed that the genes for TSHB and NGFB are located less than 310 kb apart in man and 220 kb in the mouse.

  7. Genetic predisposition to essential hypertension in a Mongolian population Detecting the C825T polymorphism of the G-protein beta 3 subunit gene

    Institute of Scientific and Technical Information of China (English)

    Chunyu Zhang; Shigang Zhao; Guangming Niu; Rile Hu; Zhiguang Wang; Mingfang Jiang; Rile Hu

    2007-01-01

    BACKGROUND: The prevalences of hypertension, cerebrovascular diseases, etc. are higher in Mongolian population because of the influence of various factors including genetics, geography, diet, etc. Therefore, it is helpful for prevention to develop researches on the genetics of various diseases including hypertension in Mongolian population.OBJECTIVE: To analyze the association between C825T polymorphisms of G-protein beta 3 subunit gene (GNB3), the important candidate gene of various disease of cardiovascular system, and Mongolian patients with essential hypertension.DESIGN: A comparative observation.SETTINGS: Department of Neurology, the First Affiliated Hospital of Inner Mongolia Medical College;Wulate Houqi Red Cross Society.PARTICIPANTS: Totally 267 Mongolian residents, whose blood relations of 3 generations were all Mongolians, were selected from Wulate Houqi, Inner Mongolia. The patients were screened based on the diagnostic standard of hypertension set by WHO in 1999, and the enrolled subjects were divided into two groups according to the level of blood pressure: ① Normal blood pressure group (n =124): 64 males and 60 females, systolic blood pressure (SBP) < 140 mm Hg (1 mm Hg=0.133 kPa), diastolic blood pressure (DBP) <90 mm Hg; ② Essential hypertension group (n =143): 71 males and 72 females, including 60 patients with simple high SBP (SBP ranged 145 to 195 mm Hg, whereas DBP < 90 mm Hg).METHODS: Peripheral venous blood (5 mL) was drawn from all the subjects, the genome DNA was extracted, and the polymorphisms of the GNB3 C825T genotype were detected with the Sequenom system.Polymerase chain reaction (PCR) experiment and SNP detection were performed in Beijing Huada gene laboratory. Then the univariate analysis of variance was applied in the sample comparison among groups, and the chi-square test was used to compare the genotypes and allele frequencies. The odd ratio (OR) and 95% confidence interval (CI)were calculated.MAIN OUTCOME MEASURES: The

  8. The role of NR2B containing NMDA receptor in place preference conditioned with morphine and natural reinforcers in rats.

    Science.gov (United States)

    Ma, Yao-Ying; Guo, Chang-Yong; Yu, Peng; Lee, David Yue-Wei; Han, Ji-Sheng; Cui, Cai-Lian

    2006-08-01

    It has been reported that N-methyl-D-aspartate (NMDA) receptor is implicated in drug addiction and antagonists of the NMDA receptor complex can inhibit the development and expression of conditioned place preference (CPP) induced by several addictive drugs, implying that this class of compounds might be considered as candidate for the treatment of substance abuse. To explore this possibility, it is important to evaluate whether the inhibitory effect of NMDA receptor antagonists would be confined to behaviors produced by drugs of abuse only, but not by natural reinforcers. According to the quantitative changes of NMDA receptor subunits, including NR1, NR2A, and NR2B, induced by diverse types of reinforcers, we chose NR2B subunit as the target of research. Experimental results showed that (1) an augmented expression of NR2B subunit was revealed by Western blotting in the nucleus accumbens (NAc) and the hippocampus in rats with CPP induced by morphine, but not by natural rewards such as food, novel environment and social interaction. (2) Ifenprodil, an antagonist highly selective for NR2B subunit of the NMDA receptor, produced a dose-dependent reduction in CPP induced by morphine and novel environment, but not that by food consumption and social interaction. Taking together, these findings suggested that NR2B containing NMDA receptor may be more involved with morphine reward rather than natural rewards, and that antagonism of NR2B may have a potential for the treatment of morphine abuse.

  9. The rice dynamin-related protein DRP2B mediates membrane trafficking, and thereby plays a critical role in secondary cell wall cellulose biosynthesis.

    Science.gov (United States)

    Xiong, Guangyan; Li, Rui; Qian, Qian; Song, Xueqin; Liu, Xiangling; Yu, Yanchun; Zeng, Dali; Wan, Jianmin; Li, Jiayang; Zhou, Yihua

    2010-10-01

    Membrane trafficking between the plasma membrane (PM) and intracellular compartments is an important process that regulates the deposition and metabolism of cell wall polysaccharides. Dynamin-related proteins (DRPs), which function in membrane tubulation and vesiculation are closely associated with cell wall biogenesis. However, the molecular mechanisms by which DRPs participate in cell wall formation are poorly understood. Here, we report the functional characterization of Brittle Culm3 (BC3), a gene encoding OsDRP2B. Consistent with the expression of BC3 in mechanical tissues, the bc3 mutation reduces mechanical strength, which results from decreased cellulose content and altered secondary wall structure. OsDRP2B, one of three members of the DRP2 subfamily in rice (Oryza sativa L.), was identified as an authentic membrane-associated dynamin via in vitro biochemical analyses. Subcellular localization of fluorescence-tagged OsDRP2B and several compartment markers in protoplast cells showed that this protein not only lies at the PM and the clathrin-mediated vesicles, but also is targeted to the trans-Golgi network (TGN). An FM4-64 uptake assay in transgenic plants that express green fluorescent protein-tagged OsDRP2B verified its involvement in an endocytic pathway. BC3 mutation and overexpression altered the abundance of cellulose synthase catalytic subunit 4 (OsCESA4) in the PM and in the endomembrane systems. All of these findings lead us to conclude that OsDRP2B participates in the endocytic pathway, probably as well as in post-Golgi membrane trafficking. Mutation of OsDRP2B disturbs the membrane trafficking that is essential for normal cellulose biosynthesis of the secondary cell wall, thereby leading to inferior mechanical properties in rice plants.

  10. Noncanonical sortase-mediated assembly of pilus type 2b in group B Streptococcus.

    Science.gov (United States)

    Lazzarin, Maddalena; Cozzi, Roberta; Malito, Enrico; Martinelli, Manuele; D'Onofrio, Mariapina; Maione, Domenico; Margarit, Immaculada; Rinaudo, C Daniela

    2015-11-01

    Group B Streptococcus (GBS) expresses 3 structurally distinct pilus types (1, 2a, and 2b) identified as important virulence factors and vaccine targets. These pili are heterotrimeric polymers, covalently assembled on the cell wall by sortase (Srt) enzymes. We investigated the pilus-2b biogenesis mechanism by using a multidisciplinary approach integrating genetic, biochemical, and structural studies to dissect the role of the 2 pilus-2b-associated Srts. We show that only 1 sortase (SrtC1-2b) is responsible for pilus protein polymerization, whereas the second one (Srt2-2b) does not act as a pilin polymerase, but similarly to the housekeeping class A Srt (SrtA), it is involved in cell-wall pilus anchoring by targeting the minor ancillary subunit. Based on its function and sequence features, Srt2-2b does not belong to class C Srts (SrtCs), nor is it a canonical member of any other known family of Srts. We also report the crystal structure of SrtC1-2b at 1.9 Å resolution. The overall fold resembles the typical structure of SrtCs except for the N-terminal lid region that appears in an open conformation displaced from the active site. Our findings reveal that GBS pilus type 2b biogenesis differs significantly from the current model of pilus assembly in gram-positive pathogens.

  11. Investigation of Class 2b Trucks

    Energy Technology Data Exchange (ETDEWEB)

    Davis, S.C.

    2002-04-03

    The popularity of trucks in the class 2 category--that is, those with a 6,000 to 10,000 pounds (lbs) gross vehicle weight rating (GVWR)--has increased since the late 1970s/early 1980s. The purpose of this research is to identify and examine vehicles in the upper portion of the class 2 weight range (designated as vehicle class 2b) and to assess their impact. Vehicles in class 2b (8,500-10,000 lbs GVWR) include pickup trucks, sport utility vehicles (SUVs), and large vans (i.e., not minivans). Oak Ridge National Laboratory researched each individual truck model to determine which models were class 2b trucks and arrived at four methodologies to derive sales volumes. Two methods--one for calendar year and one for model year sales--were recommended for producing believable and reliable results. The study indicates that 521,000 class 2b trucks were sold in calendar year 1999--6.4% of sales of all trucks under 10,000 lbs. Eighty-two percent of class 2b trucks sold in 1999 were pickups; one third of class 2b trucks sold in 1999 were diesel. There were 5.8 million class 2b trucks on the road in 2000, which amounts to 7.8% of all trucks under 10,000 lbs. Twenty-four percent of the class 2b truck population is diesel. Estimates show that class 2b trucks account for 8% of annual miles traveled by trucks under 10,000 lbs and 9% of fuel use. Data on class 2b trucks are scarce. As the Tier 2 standards, which apply to passenger vehicles in the 8,500-10,000 lb GVWR category, become effective, additional data on class 2b trucks may become available--not only emissions data, but data in all areas. At the moment, distinguishing class 2b trucks from class 2 trucks in general is a substantial task requiring data on an individual model level.

  12. The Cleavage and Polyadenylation Specificity Factor 6 (CPSF6) Subunit of the Capsid-recruited Pre-messenger RNA Cleavage Factor I (CFIm) Complex Mediates HIV-1 Integration into Genes.

    Science.gov (United States)

    Rasheedi, Sheeba; Shun, Ming-Chieh; Serrao, Erik; Sowd, Gregory A; Qian, Juan; Hao, Caili; Dasgupta, Twishasri; Engelman, Alan N; Skowronski, Jacek

    2016-05-27

    HIV-1 favors integration into active genes and gene-enriched regions of host cell chromosomes, thus maximizing the probability of provirus expression immediately after integration. This requires cleavage and polyadenylation specificity factor 6 (CPSF6), a cellular protein involved in pre-mRNA 3' end processing that binds HIV-1 capsid and connects HIV-1 preintegration complexes to intranuclear trafficking pathways that link integration to transcriptionally active chromatin. CPSF6 together with CPSF5 and CPSF7 are known subunits of the cleavage factor I (CFIm) 3' end processing complex; however, CPSF6 could participate in additional protein complexes. The molecular mechanisms underpinning the role of CPSF6 in HIV-1 infection remain to be defined. Here, we show that a majority of cellular CPSF6 is incorporated into the CFIm complex. HIV-1 capsid recruits CFIm in a CPSF6-dependent manner, which suggests that the CFIm complex mediates the known effects of CPSF6 in HIV-1 infection. To dissect the roles of CPSF6 and other CFIm complex subunits in HIV-1 infection, we analyzed virologic and integration site targeting properties of a CPSF6 variant with mutations that prevent its incorporation into CFIm We show, somewhat surprisingly, that CPSF6 incorporation into CFIm is not required for its ability to direct preferential HIV-1 integration into genes. The CPSF5 and CPSF7 subunits appear to have only a minor, if any, role in this process even though they appear to facilitate CPSF6 binding to capsid. Thus, CPSF6 alone controls the key molecular interactions that specify HIV-1 preintegration complex trafficking to active chromatin.

  13. Inverted repeat of Olisthodiscus luteus chloroplast DNA contains genes for both subunits of ribulose-1,5-bisphosphate carboxylase and the 32,000-dalton QB protein: Phylogenetic implications

    Science.gov (United States)

    Reith, Michael; Cattolico, Rose Ann

    1986-01-01

    The chloroplast DNA of the chromophytic alga Olisthodiscus luteus has been physically mapped with four restriction enzymes. An inverted repeat of 22 kilobase pairs is present in this 150-kilobase-pair plastid genome. The inverted repeat contains the genes for the large and small subunit polypeptides of ribulose-1,5-bisphosphate carboxylase (EC 4.1.1.39) and also codes for the 32,000-dalton QB protein. These observations demonstrate that significant differences exist in chloroplast genome structure and organization among major plant taxa. Images PMID:16578794

  14. NR2A- and NR2B-Containing NMDA Receptors in the Prelimbic Medial Prefrontal Cortex Differentially Mediate Trace, Delay, and Contextual Fear Conditioning

    Science.gov (United States)

    Gilmartin, Marieke R.; Kwapis, Janine L.; Helmstetter, Fred J.

    2013-01-01

    Activation of "N"-methyl-D-aspartate receptors (NMDAR) in the prelimbic medial prefrontal cortex (PL mPFC) is necessary for the acquisition of both trace and contextual fear memories, but it is not known how specific NR2 subunits support each association. The NR2B subunit confers unique properties to the NMDAR and may differentially…

  15. Analysis of iron- and sulfur-oxidizing bacteria in a treatment plant of acid rock drainage from a Japanese pyrite mine by use of ribulose-1, 5-bisphosphate carboxylase/oxygenase large-subunit gene.

    Science.gov (United States)

    Kamimura, Kazuo; Okabayashi, Ai; Kikumoto, Mei; Manchur, Mohammed Abul; Wakai, Satoshi; Kanao, Tadayoshi

    2010-03-01

    Iron- and sulfur-oxidizing bacteria in a treatment plant of acid rock drainage (ARD) from a pyrite mine in Yanahara, Okayama prefecture, Japan, were analyzed using the gene (cbbL) encoding the large subunit of ribulose-1, 5-bisphosphate carboxylase/oxygenase (RubisCO). Analyses of partial sequences of cbbL genes from Acidithiobacillus ferrooxidans, Acidithiobacillus thiooxidans and Acidithiobacillus caldus strains revealed the diversity in their cbbL gene sequences. In contrast to the presence of two copies of form I cbbL genes (cbbL1 and cbbL2) in A. ferrooxidans genome, A. thiooxidans and A. caldus had a single copy of form I cbbL gene in their genomes. A phylogenetic analysis based on deduced amino acid sequences from cbbL genes detected in the ARD treatment plant and their close relatives revealed that 89% of the total clones were affiliated with A. ferrooxidans. Clones loosely affiliated with the cbbL from A. thiooxidans NB1-3 or Thiobacillus denitrificans was also detected in the treatment plant. cbbL gene sequences of iron- or sulfur-oxidizing bacteria isolated from the ARD and the ARD treatment plant were not detected in the cbbL libraries from the treatment plant, suggesting the low frequencies of isolates in the samples.

  16. Ubiquitination and sumoylation of the HTLV-2 Tax-2B protein regulate its NF-κB activity: a comparative study with the HTLV-1 Tax-1 protein

    Directory of Open Access Journals (Sweden)

    Turci Marco

    2012-12-01

    Full Text Available Abstract Background Retroviruses HTLV-1 and HTLV-2 have homologous genomic structures but differ significantly in pathogenicity. HTLV-1 is associated with Adult T cell Leukemia (ATL, whereas infection by HTLV-2 has no association with neoplasia. Transformation of T lymphocytes by HTLV-1 is linked to the capacity of its oncoprotein Tax-1 to alter cell survival and cell cycle control mechanisms. Among these functions, Tax-1-mediated activation of cellular gene expression via the NF-κB pathway depends on Tax-1 post-translational modifications by ubiquitination and sumoylation. The Tax-2 protein of HTLV-2B (Tax-2B is also modified by ubiquitination and sumoylation and activates the NF-κB pathway to a level similar to that of Tax-1. The present study aims to understand whether ubiquitination and sumoylation modifications are involved in Tax-2B-mediated activation of the NF-κB pathway. Results The comparison of Tax-1 and Tax-2B lysine to arginine substitution mutants revealed conserved patterns and levels of ubiquitination with notable difference in the lysine usage for sumoylation. Neither Tax-1 nor Tax-2B ubiquitination and sumoylation deficient mutants could activate the NF-κB pathway and fusion of ubiquitin or SUMO-1 to the C-terminus of the ubiquitination and sumoylation deficient Tax-2B mutant strikingly restored transcriptional activity. In addition, ubiquitinated forms of Tax-2B colocalized with RelA and IKKγ in prominent cytoplasmic structures associated with the Golgi apparatus, whereas colocalization of Tax-2B with the RelA subunit of NF-κB and the transcriptional coactivator p300 in punctate nuclear structures was dependent on Tax-2B sumoylation, as previously observed for Tax-1. Conclusions Both Tax-1 and Tax-2 activate the NF-κB pathway via similar mechanisms involving ubiquitination and sumoylation. Therefore, the different transforming potential of HTLV-1 and HTLV-2 is unlikely to be related to different modes of activation of

  17. Expression of mRNA-encoding subunits of N-methyl-D-aspartate receptor in the hypothalamus in sustained monaural block of auditory air-conduction model rats

    Institute of Scientific and Technical Information of China (English)

    Ping Wan; Xiaojian Lai; Cheng Huang; Xinde Sun

    2011-01-01

    A sustained monaural block of auditory air-conduction model was established in rats through subcutaneous suture in the right ear canal. The gene expression levels of hypothalamic N-methyl-D-aspartate receptor NR1, NR2A, NR2B and NR2C mRNA in the auditory central nervous system of Sprague-Dawley rats at postnatal 9, 23, 37 days were determined after an environmental change. Reverse transcription-PCR assay showed that the critical period for the development of NR1, NR2A, and NR2B subunits in the left hypothalamus and NR1- and NR2B-dependent auditory neurons in the right hypothalamus terminated 23 days after the suture in the right ear. The critical period for the development of NR2A subunit-dependent auditory neurons in the right hypothalamus was terminated by postnatal day 37. The results confirmed that N-methyl-D-aspartate receptor subunits in the hypothalamus may be regulated by the auditory environment.

  18. Importance of the GluN2B Carboxy-Terminal Domain for Enhancement of Social Memories

    Science.gov (United States)

    Jacobs, Stephanie; Wei, Wei; Wang, Deheng; Tsien, Joe Z.

    2015-01-01

    The N-methyl-D-aspartate (NMDA) receptor is known to be necessary for many forms of learning and memory, including social recognition memory. Additionally, the GluN2 subunits are known to modulate multiple forms of memory, with a high GluN2A:GluN2B ratio leading to impairments in long-term memory, while a low GluN2A:GluN2B ratio enhances some…

  19. Impaired ventilatory and thermoregulatory responses to hypoxic stress in newborn Phox2b heterozygous knockout mice

    Directory of Open Access Journals (Sweden)

    Nelina eRamanantsoa

    2011-09-01

    Full Text Available The Phox2b gene is necessary for the development of the autonomic nervous system, and especially, of respiratory neuronal circuits. In the present study, we examined the role of Phox2b in ventilatory and thermoregulatory responses to hypoxic stress, which are closely related in the postnatal period. Hypoxic stress was generated by strong thermal stimulus, combined or not with reduced inspired O2. To this end, we exposed 6-day-old Phox2b+/- pups and their wild-type littermates (Phox2b+/+ to hypoxia (10% O2 or hypercapnia (8% CO2 under thermoneutral (33°C or cold (26°C conditions. We found that Phox2b+/- pups showed less normoxic ventilation (VE in the cold than Phox2b+/+ pups. Phox2b+/- pups also showed lower oxygen consumption (VO2 in the cold, reflecting reduced thermogenesis and a lower body temperature. Furthermore, while the cold depressed ventilatory responses to hypoxia and hypercapnia in both genotype groups, this effect was less pronounced in Phox2b+/- pups. Finally, because serotonin (5-HT neurons are pivotal to respiratory and thermoregulatory circuits and depend on Phox2b for their differentiation, we studied 5-HT metabolism using high-pressure liquid chromatography, and found that it was altered in Phox2b+/- pups. We conclude that Phox2b haploinsufficiency alters the ability of newborns to cope with metabolic challenges, possibly due to 5-HT signaling impairments.

  20. The SH2B1 obesity locus and abnormal glucose homeostasis

    DEFF Research Database (Denmark)

    Prudente, S; Copetti, M; Morini, E

    2013-01-01

    affecting risk of obesity and insulin resistance might also modulate risk of T2D. Recently, 32 loci have been associated with body mass index (BMI) by genome-wide studies, including one locus on chromosome 16p11 containing the SH2B1 gene. Animal studies have suggested that SH2B1 is a physiological enhancer...

  1. Genetics Home Reference: RRM2B-related mitochondrial DNA depletion syndrome, encephalomyopathic form with renal ...

    Science.gov (United States)

    ... Munnich A, Rötig A. Mutation of RRM2B, encoding p53-controlled ribonucleotide reductase (p53R2), causes severe mitochondrial DNA depletion. Nat Genet. 2007 Jun;39(6):776-80. Epub 2007 May 7. Citation on PubMed GeneReview: RRM2B-Related Mitochondrial Disease Pontarin G, Ferraro P, Bee L, Reichard P, ...

  2. Early microgliosis precedes neuronal loss and behavioural impairment in mice with a frontotemporal dementia-causing CHMP2B mutation

    DEFF Research Database (Denmark)

    Clayton, Emma L; Mancuso, Renzo; Nielsen, Troels Tolstrup

    2017-01-01

    Frontotemporal dementia (FTD)-causing mutations in the CHMP2B gene lead to the generation of mutant C-terminally truncated CHMP2B. We report that transgenic mice expressing endogenous levels of mutant CHMP2B developed late-onset brain volume loss associated with frank neuronal loss and FTD...

  3. Na+ channel β subunits: Overachievers of the ion channel family

    Directory of Open Access Journals (Sweden)

    William J Brackenbury

    2011-09-01

    Full Text Available Voltage gated Na+ channels (VGSCs in mammals contain a pore-forming α subunit and one or more β subunits. There are five mammalian β subunits in total: β1, β1B, β2, β3, and β4, encoded by four genes: SCN1B-SCN4B. With the exception of the SCN1B splice variant, β1B, the β subunits are type I topology transmembrane proteins. In contrast, β1B lacks a transmembrane domain and is a secreted protein. A growing body of work shows that VGSC β subunits are multifunctional. While they do not form the ion channel pore, β subunits alter gating, voltage-dependence, and kinetics of VGSC α subunits and thus regulate cellular excitability in vivo. In addition to their roles in channel modulation, β subunits are members of the immunoglobulin (Ig superfamily of cell adhesion molecules (CAMs and regulate cell adhesion and migration. β subunits are also substrates for sequential proteolytic cleavage by secretases. An example of the multifunctional nature of β subunits is β1, encoded by SCN1B, that plays a critical role in neuronal migration and pathfinding during brain development, and whose function is dependent on Na+ current and γ-secretase activity. Functional deletion of SCN1B results in Dravet Syndrome, a severe and intractable pediatric epileptic encephalopathy. β subunits are emerging as key players in a wide variety of pathophysiologies, including epilepsy, cardiac arrhythmia, multiple sclerosis, Huntington’s disease, neuropsychiatric disorders, neuropathic and inflammatory pain, and cancer. β subunits mediate multiple signaling pathways on different timescales, regulating electrical excitability, adhesion, migration, pathfinding, and transcription. Importantly, some β subunit functions may operate independent of α subunits. Thus, β subunits perform critical roles during development and disease. As such, they may prove useful in disease diagnosis and therapy.

  4. Reversal of pathology in CHMP2B-mediated frontotemporal dementia patient cells using RNA interference

    DEFF Research Database (Denmark)

    Nielsen, Troels Tolstrup; Mizielinska, Sarah; Hasholt, Lis;

    2012-01-01

    BACKGROUND: Frontotemporal dementia is the second most common form of young-onset dementia after Alzheimer's disease, and several genetic forms of frontotemporal dementia are known. A rare genetic variant is caused by a point mutation in the CHMP2B gene. CHMP2B is a component of the ESCRT...... role in the pathogenesis of the disease. METHODS: In the present study, we used lentiviral vectors to efficiently knockdown CHMP2B by delivering microRNA embedded small hairpin RNAs. RESULTS: We show that CHMP2B can be efficiently knocked down in patient fibroblasts using an RNA interference approach...

  5. Exposure to coplanar PCBs induces endothelial cell inflammation through epigenetic regulation of NF-κB subunit p65

    Science.gov (United States)

    Liu, Dandan; Perkins, Jordan T.; Petriello, Michael C.; Hennig, Bernhard

    2015-01-01

    Epigenetic modifications of DNA and histones alter cellular phenotypes without changing genetic codes. Alterations of epigenetic marks can be induced by exposure to environmental pollutants and may contribute to associated disease risks. Here we test the hypothesis that endothelial cell dysfunction induced by exposure to polychlorinated biphenyls (PCBs) is mediated in part though histone modifications. In this study, human vascular endothelial cells were exposed to physiologically relevant concentrations of several PCBs congeners (e.g., PCBs 77, 118, 126 and 153) followed by quantification of inflammatory gene expression and changes of histone methylation. Only exposure to coplanar PCBs 77 and 126 induced the expression of histone H3K9 trimethyl demethylase jumonji domain-containing protein 2B (JMJD2B) and nuclear factor-kappa B (NF-κB) subunit p65, activated NF-κB signaling as evidenced by nuclear translocation of p65, and up-regulated p65 target inflammatory genes, such as interleukin (IL)-6, C-reactive protein (CRP), intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and IL-1α/β. The increased accumulation of JMJD2B in the p65 promoter led to a depletion of H3K9me3 repression mark, which accounts for the observed up-regulation of p65 and associated inflammatory genes. JMJD2B gene knockdown confirmed a critical role for this histone demethylase in mediating PCB-induced inflammation of the vascular endothelium. Finally, it was determined, via chemical inhibition, that PCB-induced up-regulation of JMJD2B was estrogen receptor-alpha (ER-α) dependent. These data suggest that coplanar PCBs may exert endothelial cell toxicity through changes in histone modifications. PMID:26519613

  6. Regulation of PI-2b Pilus Expression in Hypervirulent Streptococcus agalactiae ST-17 BM110

    Science.gov (United States)

    du Merle, Laurence; Rosinski-Chupin, Isabelle; Gominet, Myriam; Bellais, Samuel; Poyart, Claire; Trieu-Cuot, Patrick

    2017-01-01

    The widely spread Streptococcus agalactiae (also known as Group B Streptococcus, GBS) “hypervirulent” ST17 clone is strongly associated with neonatal meningitis. The PI-2b locus is mainly found in ST17 strains but is also present in a few non ST17 human isolates such as the ST-7 prototype strain A909. Here, we analysed the expression of the PI-2b pilus in the ST17 strain BM110 as compared to the non ST17 A909. Comparative genome analyses revealed the presence of a 43-base pair (bp) hairpin-like structure in the upstream region of PI-2b operon in all 26 ST17 genomes, which was absent in the 8 non-ST17 strains carrying the PI-2b locus. Deletion of this 43-bp sequence in strain BM110 resulted in a 3- to 5-fold increased transcription of PI-2b. Characterization of PI-2b promoter region in A909 and BM110 strains was carried out by RNAseq, primer extension, qRT-PCR and transcriptional fusions with gfp as reporter gene. Our results indicate the presence of a single promoter (Ppi2b) with a transcriptional start site (TSS) mapped 37 bases upstream of the start codon of the first PI-2b gene. The large operon of 16 genes located upstream of PI-2b codes for the group B carbohydrate (also known as antigen B), a major constituent of the bacterial cell wall. We showed that the hairpin sequence located between antigen B and PI-2b operons is a transcriptional terminator. In A909, increased expression of PI-2b probably results from read-through transcription from antigen B operon. In addition, we showed that an extended 5’ promoter region is required for maximal transcription of gfp as a reporter gene in S. agalactiae from Ppi2b promoter. Gene reporter assays performed in Lactococcus lactis strain NZ9000, a related non-pathogenic Gram-positive species, revealed that GBS-specific regulatory factors are required to drive PI-2b transcription. PI-2b expression is up-regulated in the BM110ΔcovR mutant as compared to the parental BM110 strain, but this effect is probably indirect

  7. 2000 Johnston Site 2B-P

    Data.gov (United States)

    US Fish and Wildlife Service, Department of the Interior — Underwater Site 2B-P was established at Johnston Atoll by Dr. James Maragos, U.S. Fish & Wildlife Service, on June 30, 2000. With a start point (meter 0) at...

  8. B2B Pioneer A Millionaire Maker

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    China should soon see its largest group of instant millionaires after Jack Ma,Chief Executive Officer and Chairman of Alibaba Group,announced on July 27 that the China’s preeminent e-commerce company has initiated the listing of its B2B unit alibaba.com

  9. Genetic loss of SH2B3 in acute lymphoblastic leukemia

    Science.gov (United States)

    Perez-Garcia, Arianne; Ambesi-Impiombato, Alberto; Hadler, Michael; Rigo, Isaura; LeDuc, Charles A.; Kelly, Kara; Jalas, Chaim; Paietta, Elisabeth; Racevskis, Janis; Rowe, Jacob M.; Tallman, Martin S.; Paganin, Maddalena; Basso, Giuseppe; Tong, Wei; Chung, Wendy K.

    2013-01-01

    The SH2B adaptor protein 3 (SH2B3) gene encodes a negative regulator of cytokine signaling with a critical role in the homeostasis of hematopoietic stem cells and lymphoid progenitors. Here, we report the identification of germline homozygous SH2B3 mutations in 2 siblings affected with developmental delay and autoimmunity, one in whom B-precursor acute lymphoblastic leukemia (ALL) developed. Mechanistically, loss of SH2B3 increases Janus kinase-signal transducer and activator of transcription signaling, promotes lymphoid cell proliferation, and accelerates leukemia development in a mouse model of NOTCH1-induced ALL. Moreover, extended mutation analysis showed homozygous somatic mutations in SH2B3 in 2 of 167 ALLs analyzed. Overall, these results demonstrate a Knudson tumor suppressor role for SH2B3 in the pathogenesis of ALL and highlight a possible link between genetic predisposition factors in the pathogenesis of autoimmunity and leukemogenesis. PMID:23908464

  10. Functional characterization of cytochromes P450 2B from the desert woodrat Neotoma lepida

    Energy Technology Data Exchange (ETDEWEB)

    Wilderman, P. Ross, E-mail: pwilderman@ucsd.edu [Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, CA (United States); Jang, Hyun-Hee [Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, CA (United States); Malenke, Jael R. [Department of Biology, University of Utah, Salt Lake City, UT (United States); Salib, Mariam; Angermeier, Elisabeth; Lamime, Sonia [Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, CA (United States); Dearing, M. Denise [Department of Biology, University of Utah, Salt Lake City, UT (United States); Halpert, James R. [Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, CA (United States)

    2014-02-01

    Mammalian detoxification processes have been the focus of intense research, but little is known about how wild herbivores process plant secondary compounds, many of which have medicinal value or are drugs. cDNA sequences that code for three enzymes of the cytochrome P450 (CYP) 2B subfamily, here termed 2B35, 2B36, and 2B37 have been recently identified from a wild rodent, the desert woodrat (Malenke et al., 2012). Two variant clones of each enzyme were engineered to increase protein solubility and to facilitate purification, as reported for CYP2B enzymes from multiple species. When expressed in Escherichia coli each of the woodrat proteins gave the characteristic maximum at 450 nm in a reduced carbon monoxide difference spectrum but generally expressed at lower levels than rat CYP2B1. Two enzymes, 2B36 and 2B37, showed dealkylation activity with the model substrates 7-ethoxy-4-(trifluoromethyl)coumarin and 7-benzyloxyresorufin, whereas 2B35 was inactive. Binding of the monoterpene (+)-α-pinene produced a Type I shift in the absorbance spectrum of each enzyme. Mutation of 2B37 at residues 114, 262, or 480, key residues governing ligand interactions with other CYP2B enzymes, did not significantly change expression levels or produce the expected functional changes. In summary, two catalytic and one ligand-binding assay are sufficient to distinguish among CYP2B35, 2B36, and 2B37. Differences in functional profiles between 2B36 and 2B37 are partially explained by changes in substrate recognition site residue 114, but not 480. The results advance our understanding of the mechanisms of detoxification in wild mammalian herbivores and highlight the complexity of this system. - Highlights: • Three CYP2B enzymes from Neotoma lepida were cloned, engineered, and expressed. • A mix of catalytic and binding assays yields unique results for each enzyme. • Mutational analysis indicates CYP{sub 2}B substrate recognition remains to be clarified. • Reported N. lepida gene

  11. H2B ubiquitylation is part of chromatin architecture that marks exon-intron structure in budding yeast

    LENUS (Irish Health Repository)

    Shieh, Grace S.

    2011-12-22

    Abstract Background The packaging of DNA into chromatin regulates transcription from initiation through 3\\' end processing. One aspect of transcription in which chromatin plays a poorly understood role is the co-transcriptional splicing of pre-mRNA. Results Here we provide evidence that H2B monoubiquitylation (H2BK123ub1) marks introns in Saccharomyces cerevisiae. A genome-wide map of H2BK123ub1 in this organism reveals that this modification is enriched in coding regions and that its levels peak at the transcribed regions of two characteristic subgroups of genes. First, long genes are more likely to have higher levels of H2BK123ub1, correlating with the postulated role of this modification in preventing cryptic transcription initiation in ORFs. Second, genes that are highly transcribed also have high levels of H2BK123ub1, including the ribosomal protein genes, which comprise the majority of intron-containing genes in yeast. H2BK123ub1 is also a feature of introns in the yeast genome, and the disruption of this modification alters the intragenic distribution of H3 trimethylation on lysine 36 (H3K36me3), which functionally correlates with alternative RNA splicing in humans. In addition, the deletion of genes encoding the U2 snRNP subunits, Lea1 or Msl1, in combination with an htb-K123R mutation, leads to synthetic lethality. Conclusion These data suggest that H2BK123ub1 facilitates cross talk between chromatin and pre-mRNA splicing by modulating the distribution of intronic and exonic histone modifications.

  12. The gene sml0013 of Synechocystis species strain PCC 6803 encodes for a novel subunit of the NAD(P)H oxidoreductase or complex I that is ubiquitously distributed among Cyanobacteria.

    Science.gov (United States)

    Schwarz, Doreen; Schubert, Hendrik; Georg, Jens; Hess, Wolfgang R; Hagemann, Martin

    2013-11-01

    The NAD(P)H oxidoreductase or complex I (NDH1) complex participates in many processes such as respiration, cyclic electron flow, and inorganic carbon concentration in the cyanobacterial cell. Despite immense progress in our understanding of the structure-function relation of the cyanobacterial NDH1 complex, the subunits catalyzing NAD(P)H docking and oxidation are still missing. The gene sml0013 of Synechocystis 6803 encodes for a small protein of unknown function for which homologs exist in all completely known cyanobacterial genomes. The protein exhibits weak similarities to the NDH-dependent flow6 (NDF6) protein, which was reported from Arabidopsis (Arabidopsis thaliana) chloroplasts as a NDH subunit. An sml0013 inactivation mutant of Synechocystis 6803 was generated and characterized. It showed only weak differences regarding growth and pigmentation in various culture conditions; most remarkably, it exhibited a glucose-sensitive phenotype in the light. The genome-wide expression pattern of the Δsml0013::Km mutant was almost identical to the wild type when grown under high CO2 conditions as well as after shifts to low CO2 conditions. However, measurements of the photosystem I redox kinetic in cells of the Δsml0013::Km mutant revealed differences, such as a decreased capability of cyclic electron flow as well as electron flow into respiration in comparison with the wild type. These results suggest that the Sml0013 protein (named NdhP) represents a novel subunit of the cyanobacterial NDH1 complex, mediating its coupling either to the respiratory or the photosynthetic electron flow.

  13. Lineage-specific fragmentation and nuclear relocation of the mitochondrial cox2 gene in chlorophycean green algae (Chlorophyta).

    Science.gov (United States)

    Rodríguez-Salinas, Elizabeth; Riveros-Rosas, Héctor; Li, Zhongkui; Fucíková, Karolina; Brand, Jerry J; Lewis, Louise A; González-Halphen, Diego

    2012-07-01

    In most eukaryotes the subunit 2 of cytochrome c oxidase (COX2) is encoded in intact mitochondrial genes. Some green algae, however, exhibit split cox2 genes (cox2a and cox2b) encoding two polypeptides (COX2A and COX2B) that form a heterodimeric COX2 subunit. Here, we analyzed the distribution of intact and split cox2 gene sequences in 39 phylogenetically diverse green algae in phylum Chlorophyta obtained from databases (28 sequences from 22 taxa) and from new cox2 data generated in this work (23 sequences from 18 taxa). Our results support previous observations based on a smaller number of taxa, indicating that algae in classes Prasinophyceae, Ulvophyceae, and Trebouxiophyceae contain orthodox, intact mitochondrial cox2 genes. In contrast, all of the algae in Chlorophyceae that we examined exhibited split cox2 genes, and could be separated into two groups: one that has a mitochondrion-localized cox2a gene and a nucleus-localized cox2b gene ("Scenedesmus-like"), and another that has both cox2a and cox2b genes in the nucleus ("Chlamydomonas-like"). The location of the split cox2a and cox2b genes was inferred using five different criteria: differences in amino acid sequences, codon usage (mitochondrial vs. nuclear), codon preference (third position frequencies), presence of nucleotide sequences encoding mitochondrial targeting sequences and presence of spliceosomal introns. Distinct green algae could be grouped according to the form of cox2 gene they contain: intact or fragmented, mitochondrion- or nucleus-localized, and intron-containing or intron-less. We present a model describing the events that led to mitochondrial cox2 gene fragmentation and the independent and sequential migration of cox2a and cox2b genes to the nucleus in chlorophycean green algae. We also suggest that the distribution of the different forms of the cox2 gene provides important insights into the phylogenetic relationships among major groups of Chlorophyceae.

  14. Structure and assembly of group B streptococcus pilus 2b backbone protein.

    Science.gov (United States)

    Cozzi, Roberta; Malito, Enrico; Lazzarin, Maddalena; Nuccitelli, Annalisa; Castagnetti, Andrea; Bottomley, Matthew J; Margarit, Immaculada; Maione, Domenico; Rinaudo, C Daniela

    2015-01-01

    Group B Streptococcus (GBS) is a major cause of invasive disease in infants. Like other Gram-positive bacteria, GBS uses a sortase C-catalyzed transpeptidation mechanism to generate cell surface pili from backbone and ancillary pilin precursor substrates. The three pilus types identified in GBS contain structural subunits that are highly immunogenic and are promising candidates for the development of a broadly-protective vaccine. Here we report the X-ray crystal structure of the backbone protein of pilus 2b (BP-2b) at 1.06Å resolution. The structure reveals a classical IgG-like fold typical of the pilin subunits of other Gram-positive bacteria. The crystallized portion of the protein (residues 185-468) encompasses domains D2 and D3 that together confer high stability to the protein due to the presence of an internal isopeptide bond within each domain. The D2+D3 region, lacking the N-terminal D1 domain, was as potent as the entire protein in conferring protection against GBS challenge in a well-established mouse model. By site-directed mutagenesis and complementation studies in GBS knock-out strains we identified the residues and motives essential for assembly of the BP-2b monomers into high-molecular weight complexes, thus providing new insights into pilus 2b polymerization.

  15. Structure and assembly of group B streptococcus pilus 2b backbone protein.

    Directory of Open Access Journals (Sweden)

    Roberta Cozzi

    Full Text Available Group B Streptococcus (GBS is a major cause of invasive disease in infants. Like other Gram-positive bacteria, GBS uses a sortase C-catalyzed transpeptidation mechanism to generate cell surface pili from backbone and ancillary pilin precursor substrates. The three pilus types identified in GBS contain structural subunits that are highly immunogenic and are promising candidates for the development of a broadly-protective vaccine. Here we report the X-ray crystal structure of the backbone protein of pilus 2b (BP-2b at 1.06Å resolution. The structure reveals a classical IgG-like fold typical of the pilin subunits of other Gram-positive bacteria. The crystallized portion of the protein (residues 185-468 encompasses domains D2 and D3 that together confer high stability to the protein due to the presence of an internal isopeptide bond within each domain. The D2+D3 region, lacking the N-terminal D1 domain, was as potent as the entire protein in conferring protection against GBS challenge in a well-established mouse model. By site-directed mutagenesis and complementation studies in GBS knock-out strains we identified the residues and motives essential for assembly of the BP-2b monomers into high-molecular weight complexes, thus providing new insights into pilus 2b polymerization.

  16. γ-羟丁酸钠对缺氧缺血后新生大鼠海马N-甲基-D-天冬氨酸受体2B亚基mRNA表达的影响%Effect of sodium gamma-hydroxybutyrate on expression of hippocampal N-methyl-D-aspartate receptor 2B subunit mRNA of hypoxic-ischemic neonatal rat

    Institute of Scientific and Technical Information of China (English)

    张研; 马正良; 曾因明; 于常州; 范建伟

    2005-01-01

    目的探讨γ-羟丁酸钠(GHB-Na)对缺氧缺血后脑损伤新生大鼠有无保护作用及N-甲基-D-天冬氨酸受体2B亚基(NR2B)与脑损伤的关系.方法180只7日龄SD乳大鼠,随机分为5组(每组36只),分别为假手术(Sham)对照组、生理盐水(NS)对照组、GHB-Na 50,100和200 mg·kg-1组.按Rice法制备缺氧缺血模型.Sham组仅分离颈总动脉,不结扎,不缺氧.GHB-Na组动物于缺氧缺血后立即腹腔注射GHB-Na.Sham组和NS组动物注射相同体积的生理盐水.以后每8 h一次.在术后2, 6, 12, 24 h, 3和7 d每组处死6只乳大鼠.应用逆转录-聚合酶链反应技术测定左侧海马组织中NR2B mRNA的表达,并进行半定量分析.结果与Sham组相比,NS组术后2 h,NR2B mRNA的表达明显减少(约20%); 在术后6,12,24 h和3 d, NR2B mRNA的表达没有明显变化; 术后7 d,NR2B mRNA的表达显著升高(约94%).与NS组相比, GHB-Na 100 mg·kg-1组在术后6,12,24 h, 3和7 d,NR2B mRNA的表达明显减少(分别减少约27%,45%,19%,31%和37%);而GHB-Na 50,200 mg·kg-1组的NR2B mRNA 在这些时间点虽然也较NS组有不同程度的减少,但差异多无显著意义.结论 GHB-Na能有效抑制新生大鼠缺氧缺血后海马NR2B mRNA的表达,特别是100 mg·kg-1组的抑制作用好于50和200 mg·kg-1组.

  17. Behavior of RNAi suppressor protein 2b of Cucumber mosaic virus in planta in presence and absence of virus.

    Science.gov (United States)

    Praveen, Shelly; Mangrauthia, Satendra K; Singh, Priyanka; Mishra, Anil K

    2008-08-01

    The 2b protein encoded by Cucumber mosaic virus (CMV) has been shown as a virus counter defense factor that interferes with the RNAi pathway. The 2b gene from CMV-banana, New Delhi isolate (CMV-NDLS) was amplified from CMV infected cucumber plants to generate the sense and antisense binary vector constructs for 2b expression and repression in planta. Constitutive expression of 2b gene in healthy Nicotiana tabacum caused phenotypic aberrations during somatic embryogenesis, which were not observed when expressed in CMV infected N. tabacum. Further, the established virus population in CMV infected N. tabacum was not affected by constitutive expression and repression of 2b gene. Thus, indicating its role in initiation of gene silencing, at the early stage of viral infection. This is the first demonstration of differential behavior of 2b suppressor protein in host development in the absence and presence of virus.

  18. Genotypic to expression profiling of bovine calcium channel, voltage-dependent, alpha-2/delta subunit 1 gene, and their association with bovine mastitis among Frieswal (HFX Sahiwal) crossbred cattle of Indian origin.

    Science.gov (United States)

    Deb, Rajib; Singh, Umesh; Kumar, Sushil; Kumar, Arun; Singh, Rani; Sengar, Gyanendra; Mann, Sandeep; Sharma, Arjava

    2014-04-01

    Calcium channel, voltage-dependent, alpha-2/delta subunit 1 (CACNA2D1) gene is considered to be an important noncytokine candidate gene influencing mastitis. Scanty of reports are available until today regarding the role play of CACNA2D1 gene on the susceptibility of bovine mastitis. We interrogated the CACNA2D1 G519663A [A>G] SNP by PCR-RFLP among two hundreds Frieswal (HF X Sahiwal) crossbred cattle of Indian origin. Genotypic frequency of AA (51.5, n=101) was comparatively higher than AG (35, n=70) and GG (14.5, n=29). Association of Somatic cell score (SCS) with genotypes revealed that, GG genotypes showing lesser count (less susceptible to mastitis) compare to AA and AG. Relative expression of CACNA2D1 transcript (in milk samples) was significantly higher among GG than AG and AA. Further we have also isolated blood sample from the all groups and PBMCs were cultured from each blood sample as per the standard protocol. They were treated with Calcium channel blocker and the expression level of the CACNA2D1 gene was evaluated by Real Time PCR. Results show that expression level decline in each genotypic group after treatment and expression level of GG are again significantly higher than AA and AG. Thus, it may be concluded that GG genotypic animals are favorable for selecting disease resistant breeds.

  19. The gene for human U2 snRNP auxiliary factor small 35-kDa subunit (U2AF1) maps to the progressive myoclonus epilepsy (EPM1) critical region on chromosome 21q22.3

    Energy Technology Data Exchange (ETDEWEB)

    Lalioti, M.D.; Rossier, C.; Antonarakis, S.E. [Univ. of Geneva Medical School (Switzerland)] [and others

    1996-04-15

    We used targeted exon trapping to clone portions of genes from human chromosome 21q22.3. One trapped sequence showed complete homology with the cDNA of human U2AF{sup 35} (M96982; HGM-approved nomenclature U2AF1), which encodes for the small 35-kDa subunit of the U2 snRNP auxiliary factor. Using the U2AF1 cDNA as a probe, we mapped this gene to cosmid Q15D2, a P1, and YAC 350F7 of the Chumakov et al. contig, close to the cystathionine-{beta}-synthase gene (CBS) on 21q22.3. This localization was confirmed by PCR using oligonucleotides from the 3{prime} UTR and by FISH. As U2AF1 associated with a number of different factors during mRNA splicing, overexpression in trisomy 21 individuals could contribute to some Down syndrome phenotypes by interfering with the splicing process. Furthermore, because this gene maps in the critical region for the progressive myoclonus epilepsy I locus (EPM1), mutation analysis will be carried out in patients to evaluate the potential role of U2AF1 as a candidate for EPM1. 24 refs., 1 fig.

  20. Ethanol-withdrawal seizures are controlled by tissue plasminogen activator via modulation of NR2B-containing NMDA receptors.

    Science.gov (United States)

    Pawlak, Robert; Melchor, Jerry P; Matys, Tomasz; Skrzypiec, Anna E; Strickland, Sidney

    2005-01-11

    Chronic ethanol abuse causes up-regulation of NMDA receptors, which underlies seizures and brain damage upon ethanol withdrawal (EW). Here we show that tissue-plasminogen activator (tPA), a protease implicated in neuronal plasticity and seizures, is induced in the limbic system by chronic ethanol consumption, temporally coinciding with up-regulation of NMDA receptors. tPA interacts with NR2B-containing NMDA receptors and is required for up-regulation of the NR2B subunit in response to ethanol. As a consequence, tPA-deficient mice have reduced NR2B, extracellular signal-regulated kinase 1/2 phosphorylation, and seizures after EW. tPA-mediated facilitation of EW seizures is abolished by NR2B-specific NMDA antagonist ifenprodil. These results indicate that tPA mediates the development of physical dependence on ethanol by regulating NR2B-containing NMDA receptors.

  1. Comparative Analysis of Eubacterial DNA Polymerase Ⅲ Alpha Subunits

    Institute of Scientific and Technical Information of China (English)

    Xiao-Qian Zhao; Jian-Fei Hu; Jun Yu

    2006-01-01

    DNA polymerase Ⅲ is one of the five eubacterial DNA polymerases that is responsible for the replication of DNA duplex. Among the ten subunits of the DNA polymerase Ⅲ core enzyme, the alpha subunit catalyzes the reaction for polymerizing both DNA strands. In this study, we extracted genomic sequences of the alpha subunit from 159 sequenced eubacterial genomes, and carried out sequencebased phylogenetic and structural analyses. We found that all eubacterial genomes have one or more alpha subunits, which form either homodimers or heterodimers.Phylogenetic and domain structural analyses as well as copy number variations of the alpha subunit in each bacterium indicate the classification of alpha subunit into four basic groups: polC, dnaE1, dnaE2, and dnaE3. This classification is of essence in genome composition analysis. We also consolidated the naming convention to avoid further confusion in gene annotations.

  2. WTS-2 b: Too close for comfort?

    Directory of Open Access Journals (Sweden)

    Kovács G.

    2013-04-01

    Full Text Available We report the discovery of WTS-2 b, a typical hot Jupiter in an unusually close 1.02-day orbit to a K-dwarf star. This is the second planet to be discovered in the infrared light curves of the WFCAM Transit Survey (WTS and is only one-and-a-half times the separation from its host star at which is would be destroyed by Roche lobe overflow. The predicted remaining lifetime of the planet is just 38 Myrs, assuming a tidal dissipation quality factor of Q'* = 106. The magnitude of Q'* is largely unconstrained by observations, thus WTS-2 b provides a useful calibration point for theories describing how frictional processes within a host star affect the tidal orbital evolution of its companion giant planets. It is expected that stars with large convective envelopes are more efficient at dissipating the orbital energy of the planet, and WTS-2 b provides an observational constraint in the sparsely populated K-dwarf regime. In addition, despite its relatively faint magnitude, the favourable size ratio of the WTS-2 star-planet system and the predicted hot equilibrium temperature of the planet will make it possible to characterise the planet's atmosphere via secondary eclipse measurements using existing ground-based instrumentation.

  3. Functional Diversification of Maize RNA Polymerase IV and V subtypes via Alternative Catalytic Subunits

    Energy Technology Data Exchange (ETDEWEB)

    Haag, Jeremy R.; Brower-Toland, Brent; Krieger, Elysia K.; Sidorenko, Lyudmila; Nicora, Carrie D.; Norbeck, Angela D.; Irsigler, Andre; LaRue, Huachun; Brzeski, Jan; Mcginnis, Karen A.; Ivashuta, Sergey; Pasa-Tolic, Ljiljana; Chandler, Vicki L.; Pikaard, Craig S.

    2014-10-01

    Unlike nuclear multisubunit RNA polymerases I, II, and III, whose subunit compositions are conserved throughout eukaryotes, plant RNA polymerases IV and V are nonessential, Pol II-related enzymes whose subunit compositions are still evolving. Whereas Arabidopsis Pols IV and V differ from Pol II in four or five of their 12 subunits, respectively, and differ from one another in three subunits, proteomic ana- lyses show that maize Pols IV and V differ from Pol II in six subunits but differ from each other only in their largest subunits. Use of alternative catalytic second subunits, which are nonredundant for development and paramutation, yields at least two sub- types of Pol IV and three subtypes of Pol V in maize. Pol IV/Pol V associations with MOP1, RMR1, AGO121, Zm_DRD1/CHR127, SHH2a, and SHH2b extend parallels between paramutation in maize and the RNA-directed DNA methylation pathway in Arabidopsis.

  4. Diabetes susceptibility in Mayas: Evidence for the involvement of polymorphisms in HHEX, HNF4α, KCNJ11, PPARγ, CDKN2A/2B, SLC30A8, CDC123/CAMK1D, TCF7L2, ABCA1 and SLC16A11 genes.

    Science.gov (United States)

    Lara-Riegos, J C; Ortiz-López, M G; Peña-Espinoza, B I; Montúfar-Robles, I; Peña-Rico, M A; Sánchez-Pozos, K; Granados-Silvestre, M A; Menjivar, M

    2015-07-01

    Association of type 2 diabetes (T2D) with common variants in HHEX, HNF4α, KCNJ11, PPARγ, CDKN2A/2B, SLC30A8, CDC123/CAMK1D, TCF7L2, ABCA1 and SLC16A11 genes have been reported, mainly in populations of European and Asian ancestry and to a lesser extent in Latin Americans. Thus, we aimed to investigate the contribution of rs1111875 (HHEX), rs1800961 (HNF4α), rs5219 (KCNJ11), rs1801282 (PPARγ), rs10811661 (CDKN2A/2B), rs13266634 (SLC30A8), rs12779790 (CDC123/CAMK1D), rs7903146 (TCF7L2), rs9282541 (ABCA1) and rs13342692 (SLC16A11) polymorphisms in the genetic background of Maya population to associate their susceptibility to develop T2D. This is one of the first studies designed specifically to investigate the inherited component of T2D in the indigenous population of Mexico. SNPs were genotyped by allelic discrimination method in 575 unrelated Maya individuals. Two SNPs rs10811661 and rs928254 were significantly associated with T2D after adjusting for BMI; rs10811661 in a recessive and rs9282541 in a dominant model. Additionally, we found phenotypical alterations associated with genetic variants: HDL to rs9282541 and insulin to rs13342692. In conclusion, these findings support an association of genetic polymorphisms to develop T2D in Maya population.

  5. Identification of Bovine, Pig and Duck Meat Species in Mixtures and in Meat Products on the Basis of the mtDNA Cytochrome Oxidase Subunit I (COI Gene Sequence

    Directory of Open Access Journals (Sweden)

    Spychaj Anita

    2016-03-01

    Full Text Available The aim of this study was to develop a method using PCR and self-designed primers on the basis of the mtDNA cytochrome oxidase subunit I (COI gene sequence to enable direct identification of the meat of three species of animals, i.e. bovines, pigs and ducks, in the single type sample, in meat mixtures and meat products. The mixtures comprised up to six meat species including apart from beef, pork and duck also chicken, turkey and goose meat. The obtained results indicate the possibility of qualitative identification of the aforementioned meat species in all types of investigated food products. The maximum length of PCR products did not exceed 300 bp, which was supposed to favour the amplification of DNA from meat products which are usually thermally processed and/or exposed to high pressure. PCR primers hybridised selectively with bovine, pig and duck DNA, showing total species specificity.

  6. Structural basis for the high Ca2+ affinity of the ubiquitous SERCA2b Ca2+ pump.

    Science.gov (United States)

    Vandecaetsbeek, Ilse; Trekels, Mieke; De Maeyer, Marc; Ceulemans, Hugo; Lescrinier, Eveline; Raeymaekers, Luc; Wuytack, Frank; Vangheluwe, Peter

    2009-11-03

    Sarco(endo)plasmic reticulum Ca(2+) ATPase (SERCA) Ca(2+) transporters pump cytosolic Ca(2+) into the endoplasmic reticulum, maintaining a Ca(2+) gradient that controls vital cell functions ranging from proliferation to death. To meet the physiological demand of the cell, SERCA activity is regulated by adjusting the affinity for Ca(2+) ions. Of all SERCA isoforms, the housekeeping SERCA2b isoform displays the highest Ca(2+) affinity because of a unique C-terminal extension (2b-tail). Here, an extensive structure-function analysis of SERCA2b mutants and SERCA1a2b chimera revealed how the 2b-tail controls Ca(2+) affinity. Its transmembrane (TM) segment (TM11) and luminal extension functionally cooperate and interact with TM7/TM10 and luminal loops of SERCA2b, respectively. This stabilizes the Ca(2+)-bound E1 conformation and alters Ca(2+)-transport kinetics, which provides the rationale for the higher apparent Ca(2+) affinity. Based on our NMR structure of TM11 and guided by mutagenesis results, a structural model was developed for SERCA2b that supports the proposed 2b-tail mechanism and is reminiscent of the interaction between the alpha- and beta-subunits of Na(+),K(+)-ATPase. The 2b-tail interaction site may represent a novel target to increase the Ca(2+) affinity of malfunctioning SERCA2a in the failing heart to improve contractility.

  7. Insights into CYP2B6-mediated drug–drug interactions

    Directory of Open Access Journals (Sweden)

    William D. Hedrich

    2016-09-01

    Full Text Available Mounting evidence demonstrates that CYP2B6 plays a much larger role in human drug metabolism than was previously believed. The discovery of multiple important substrates of CYP2B6 as well as polymorphic differences has sparked increasing interest in the genetic and xenobiotic factors contributing to the expression and function of the enzyme. The expression of CYP2B6 is regulated primarily by the xenobiotic receptors constitutive androstane receptor (CAR and pregnane X receptor (PXR in the liver. In addition to CYP2B6, these receptors also mediate the inductive expression of CYP3A4, and a number of important phase II enzymes and drug transporters. CYP2B6 has been demonstrated to play a role in the metabolism of 2%–10% of clinically used drugs including widely used antineoplastic agents cyclophosphamide and ifosfamide, anesthetics propofol and ketamine, synthetic opioids pethidine and methadone, and the antiretrovirals nevirapine and efavirenz, among others. Significant inter-individual variability in the expression and function of the human CYP2B6 gene exists and can result in altered clinical outcomes in patients receiving treatment with CYP2B6-substrate drugs. These variances arise from a number of sources including genetic polymorphism, and xenobiotic intervention. In this review, we will provide an overview of the key players in CYP2B6 expression and function and highlight recent advances made in assessing clinical ramifications of important CYP2B6-mediated drug–drug interactions.

  8. CDKN2B expression and subcutaneous adipose tissue expandability: Possible influence of the 9p21 atherosclerosis locus

    Energy Technology Data Exchange (ETDEWEB)

    Svensson, Per-Arne; Wahlstrand, Björn; Olsson, Maja [Institute of Medicine, The Sahlgrenska Academy at University of Gothenburg (Sweden); Froguel, Philippe; Falchi, Mario [Department of Genomics of Common Disease, School of Public Health, Imperial College London (United Kingdom); Bergman, Richard N. [Diabetes and Obesity Research Institute, Cedars-Sinai Medical Center, Los Angeles, CA (United States); McTernan, Philip G. [Division of Metabolic and Vascular Health, Warwick Medical School, University of Warwick, Coventry (United Kingdom); Hedner, Thomas; Carlsson, Lena M.S. [Institute of Medicine, The Sahlgrenska Academy at University of Gothenburg (Sweden); Jacobson, Peter, E-mail: peter.jacobson@medfak.gu.se [Institute of Medicine, The Sahlgrenska Academy at University of Gothenburg (Sweden)

    2014-04-18

    Highlights: • The tumor suppressor gene CDKN2B is highly expressed in human adipose tissue. • Risk alleles at the 9p21 locus modify CDKN2B expression in a BMI-dependent fashion. • There is an inverse relationship between expression of CDKN2B and adipogenic genes. • CDKN2B expression influences to postprandial triacylglycerol clearance. • CDKN2B expression in adipose tissue is linked to markers of hepatic steatosis. - Abstract: Risk alleles within a gene desert at the 9p21 locus constitute the most prevalent genetic determinant of cardiovascular disease. Previous research has demonstrated that 9p21 risk variants influence gene expression in vascular tissues, yet the biological mechanisms by which this would mediate atherosclerosis merits further investigation. To investigate possible influences of this locus on other tissues, we explored expression patterns of 9p21-regulated genes in a panel of multiple human tissues and found that the tumor suppressor CDKN2B was highly expressed in subcutaneous adipose tissue (SAT). CDKN2B expression was regulated by obesity status, and this effect was stronger in carriers of 9p21 risk alleles. Covariation between expression of CDKN2B and genes implemented in adipogenesis was consistent with an inhibitory effect of CDKN2B on SAT proliferation. Moreover, studies of postprandial triacylglycerol clearance indicated that CDKN2B is involved in down-regulation of SAT fatty acid trafficking. CDKN2B expression in SAT correlated with indicators of ectopic fat accumulation, including markers of hepatic steatosis. Among genes regulated by 9p21 risk variants, CDKN2B appears to play a significant role in the regulation of SAT expandability, which is a strong determinant of lipotoxicity and therefore might contribute to the development of atherosclerosis.

  9. The upr-1 gene encodes a catalytic subunit of the DNA polymerase zeta which is involved in damage-induced mutagenesis in Neurospora crassa.

    Science.gov (United States)

    Sakai, W; Ishii, C; Inoue, H

    2002-05-01

    The upr-1 mutant was one of the first mutagen-sensitive mutants to be isolated in Neurospora crassa. However, the function of the upr-1 gene has not yet been elucidated, although some genetic and biochemical data have been accumulated. In order to clone the upr-1 gene, we performed a chromosome walk from the mat locus, the closest genetic marker to upr-1 for which a molecular probe was available, towards the centromere, and a chromosomal contig of about 300-400 kb was constructed. Some of these clones complemented the temperature sensitivity of the un-16 mutation, which is located between mat and upr-1. The un-16 gene was sequenced, and localized in the MIPS Neurospora crassa genome database. We then searched the regions flanking un-16 for homologs of known DNA repair genes, and found a gene homologous to the REV3 gene of budding yeast. The phenotype of the upr-1 mutant is similar to that of the yeast rev3 mutant. An ncrev3 mutant carrying mutations in the N. crassa REV3 homolog was constructed using the RIP (repeat-induced point mutation) process. The spectrum of mutagen sensitivity of the ncrev3 mutant was similar to that of the upr-1 mutant. Complementation tests between the upr-1 and ncrev3 mutations indicated that the upr-1 gene is in fact identical to the ncrev3 gene. To clarify the role of the upr-1 gene in DNA repair, the frequency of MMS and 4NQO-induced mutations was assayed using the ad-8 reversion test. The upr-1 mutant was about 10 times less sensitive to both chemicals than the wild type. The expression level of the upr-1 gene is increased on exposure to UV irradiation in the uvs-2 and mus-8 mutants, which belong to postreplication repair group, as well as in the wild type. All these results suggest that the product of the upr-1 gene functions in damage-induced mutagenesis and DNA translesion synthesis in N. crassa.

  10. 大豆7S蛋白β亚基基因RNAi表达载体构建%Construction of the RNAi Expression Vector of Soybean 7S Protein β-subunit Gene

    Institute of Scientific and Technical Information of China (English)

    戴嘉乐; 马建; 付永平; 曲静; 王丕武

    2012-01-01

    In this study we used β-subunit gene of 7S protein as the target genes(Gene number: AB030840) ,tken obtained The current version does not support copying Cyrillic text to the Clipboard. pression vector of 7S protein (β-subunil was successfully constructed. This study provided foundation for the application of RNA interference technology to reduce soybean allergens and improve quality of soybean proteins.%以大豆7S蛋白β亚基基因为靶基因(基因编号:AB030840),利用RT-PCR克隆得到大豆7S蛋白β亚基基因核心序列398 bp,构建了以抗除草剂基因BAR为筛选标记的安全植物RNAi表达载体BAR-7αp-β.分子生物学检测表明7S蛋白β亚基基因的表达载体构建成功.研究结果为应用RNA干扰技术降低大豆过敏原,提高大豆蛋白品质奠定了基础.

  11. Targeted deletion of one or two copies of the G protein β subunit Gβ5 gene has distinct effects on body weight and behavior in mice.

    Science.gov (United States)

    Wang, Qiang; Levay, Konstantin; Chanturiya, Tatyana; Dvoriantchikova, Galina; Anderson, Karen L; Bianco, Suzy D C; Ueta, Cintia B; Molano, R Damaris; Pileggi, Antonello; Gurevich, Eugenia V; Gavrilova, Oksana; Slepak, Vladlen Z

    2011-11-01

    We investigated the physiological role of Gβ5, a unique G protein β subunit that dimerizes with regulators of G protein signaling (RGS) proteins of the R7 family instead of Gγ. Gβ5 is essential for stability of these complexes, so that its knockout (KO)causes degradation of the entire Gβ5-R7 family. We report that the Gβ5-KO mice remain leaner than the wild type (WT) throughout their lifetime and are resistant to a high-fat diet. They have a 5-fold increase in locomotor activity, increased thermogenesis, and lower serum insulin, all of which correlate with a higher level of secreted epinephrine. Heterozygous (HET) mice are 2-fold more active than WT mice. Surprisingly, with respect to body weight, the HET mice display a phenotype opposite to that of the KO mice: by the age of 6 mo, they are ≥ 15% heavier than the WT and have increased adiposity, insulin resistance, and liver steatosis. These changes occur in HET mice fed a normal diet and without apparent hyperphagia, mimicking basic characteristics of human metabolic syndrome. We conclude that even a partial reduction in Gβ5-R7 level can perturb normal animal metabolism and behavior. Our data on Gβ5 haploinsufficient mice may explain earlier observations of genetic linkage between R7 family mutations and obesity in humans.

  12. Diversity of low-molecular-weight-glutenin subunit genes associated with D-genome in Triticum aestivum, Aegilops crassa, A. cylindrica and A. tauschii

    Directory of Open Access Journals (Sweden)

    Fatemeh Vafadar Shamasbi

    2016-09-01

    Full Text Available The 40% of endosperm protein of common wheat is composed of Low Molecular Weight (LMW Glutenin Subunits. To examine variation in the D genome of wheat, 98 accessions from different areas of Iran were studied using the five Glu-D3-specific pair primers. The amplification percentages of all primer pair sets were 80.61%, 92.86%, 79.59%, 90.82% and 67.35%, respectively. In comparison of the four species, the most observed bands of the first primer pair were found in Ae. tauschii samples. For the second primer pair, the most frequency of the amplified product was found in the T. aestivum samples. For the third primer pair, the Ae. cylindrical samples had the most amplified band. For the fourth primer pair, the most amplified band was found in the T. aestivum samples. The Ae. cylindrica samples had the most frequency band for the fifth primer pair. Based on dendrogram analysis, the accessions were divided in to 18 categories; and also 42 accessions had a bond for any PCR reaction. It is hoped that the result will be effective in molecular analysis and breeding of native landrace plants.

  13. PSD95 suppresses dendritic arbor development in mature hippocampal neurons by occluding the clustering of NR2B-NMDA receptors.

    Directory of Open Access Journals (Sweden)

    Fernando J Bustos

    Full Text Available Considerable evidence indicates that the NMDA receptor (NMDAR subunits NR2A and NR2B are critical mediators of synaptic plasticity and dendritogenesis; however, how they differentially regulate these processes is unclear. Here we investigate the roles of the NR2A and NR2B subunits, and of their scaffolding proteins PSD-95 and SAP102, in remodeling the dendritic architecture of developing hippocampal neurons (2-25 DIV. Analysis of the dendritic architecture and the temporal and spatial expression patterns of the NMDARs and anchoring proteins in immature cultures revealed a strong positive correlation between synaptic expression of the NR2B subunit and dendritogenesis. With maturation, the pruning of dendritic branches was paralleled by a strong reduction in overall and synaptic expression of NR2B, and a significant elevation in synaptic expression of NR2A and PSD95. Using constructs that alter the synaptic composition, we found that either over-expression of NR2B or knock-down of PSD95 by shRNA-PSD95 augmented dendritogenesis in immature neurons. Reactivation of dendritogenesis could also be achieved in mature cultured neurons, but required both manipulations simultaneously, and was accompanied by increased dendritic clustering of NR2B. Our results indicate that the developmental increase in synaptic expression of PSD95 obstructs the synaptic clustering of NR2B-NMDARs, and thereby restricts reactivation of dendritic branching. Experiments with shRNA-PSD95 and chimeric NR2A/NR2B constructs further revealed that C-terminus of the NR2B subunit (tail was sufficient to induce robust dendritic branching in mature hippocampal neurons, and suggest that the NR2B tail is important in recruiting calcium-dependent signaling proteins and scaffolding proteins necessary for dendritogenesis.

  14. Microprocessor complex subunit DiGeorge syndrome critical region gene 8 (Dgcr8) is required for schwann cell myelination and myelin maintenance.

    Science.gov (United States)

    Lin, Hsin-Pin; Oksuz, Idil; Hurley, Edward; Wrabetz, Lawrence; Awatramani, Rajeshwar

    2015-10-02

    We investigated the role of a key component of the Microprocessor complex, DGCR8, in the regulation of myelin formation and maintenance. We found that conditionally ablating Dgcr8 in Schwann cells (SCs) during development results in an arrest of SC differentiation. Dgcr8 conditional knock-out (cKO) SCs fail to form 1:1 relationships with axons or, having achieved this, fail to form myelin sheaths. The expression of genes normally found in immature SCs, such as sex-determining region Y-box 2 (Sox2), is increased in Dgcr8 cKO SCs, whereas the expression of myelin-related genes, including the master regulatory transcription factor early growth response 2 (Egr2), is decreased. Additionally, expression of a novel gene expression program involving sonic hedgehog (Shh), activated de novo in injured nerves, is elevated in Dgcr8 cKOs but not in Egr2 null mice, a model of SC differentiation arrest, suggesting that the injury-related gene expression program in Dgcr8 cKOs cannot be attributed to differentiation arrest. Inducible ablation of Dgcr8 in adult SCs results in gene expression changes similar to those found in cKOs, including an increase in the expression of Sox2 and Shh. Analyses of these nerves mainly reveal normal myelin thickness and axon size distribution but some dedifferentiated SCs and increased macrophage infiltration. Together our data suggest that Dgcr8 is responsible for modulation of gene expression programs underlying myelin formation and maintenance as well as suppression of an injury-related gene expression program.

  15. Induction of T helper 1 response by immunization of BALB/c mice with the gene encoding the second subunit of Echinococcus granulosus antigen B (EgAgB8/2

    Directory of Open Access Journals (Sweden)

    Boutennoune H.

    2012-05-01

    Full Text Available A pre-designed plasmid containing the gene encoding the second subunit of Echinococcus granulosus AgB8 (EgAgB8/2 was used to study the effect of the immunization route on the immune response in BALB/c mice. Mice were immunized with pDRIVEEgAgB8/ 2 or pDRIVE empty cassette using the intramuscular (i.m., intranasal (i.n. or the epidermal gene gun (g.g. routes. Analysis of the antibody response and cytokine data revealed that gene immunization by the i.m. route induced a marked bias towards a T helper type 1 (Th1 immune response as characterized by high IFN-γ gene expression and a low IgG1/IgG2a reactivity index (R.I. ratio of 0.04. The i.n. route showed a moderate IFN-γ expression but a higher IgG1/IgG2a R.I. ratio of 0.25 indicating a moderate Th1 response. In contrast, epidermal g.g. immunization induced a Th2 response characterized by high IL-4 expression and the highest IgG1/IgG2a R.I. ratio of 0.58. In conclusion, this study showed the advantage of genetic immunization using the i.m. route and i.n. over the epidermal g.g. routes in the induction of Th1 immunity in response to E. granulosus AgB gene immunization.

  16. Transcriptional regulation of the nuclear gene encoding the alpha-subunit of the mammalian mitochondrial F1F0 ATP synthase complex: role for the orphan nuclear receptor, COUP-TFII/ARP-1.

    Science.gov (United States)

    Jordan, Elzora M; Worley, Teri; Breen, Gail A M

    2003-03-11

    Our laboratory has been studying the transcriptional regulation of the nuclear gene (ATPA) that encodes the alpha-subunit of the mammalian mitochondrial F1F0 ATP synthase complex. We have previously determined that the regulatory factor, upstream stimulatory factor 2 (USF2), can stimulate transcription of the ATPA gene through the cis-acting regulatory element 1 in the upstream promoter of this gene. In this study, we used the yeast one-hybrid screening method to identify another factor, COUP-TFII/ARP-1, which also binds to the ATPA cis-acting regulatory element 1. Binding of the orphan nuclear receptor, COUP-TFII/ARP-1, to the ATPA regulatory element 1 was confirmed using electrophoretic mobility shift experiments, and COUP-TFII/ARP-1-containing complexes were detected in HeLa cell nuclear extracts. A mutational analysis indicated that the binding site for COUP-TFII/ARP-1 in the ATPA regulatory element 1 is an imperfect direct repeat of a nuclear receptor response element (A/GGGTCA) with a spacer of three nucleotides. Functional assays in HeLa cells showed that COUP-TFII/ARP-1 represses the ATPA promoter activity in a dose- and sequence-dependent manner. Furthermore, cotransfection assays demonstrated that COUP-TFII/ARP-1 inhibits the USF2-mediated activation of the wild-type ATPA gene promoter but not a mutant promoter that is defective in COUP-TFII/ARP-1-binding. Overexpression of USF2 reversed the COUP-TFII/ARP-1-mediated repression of the ATPA promoter. Mobility shift assays revealed that COUP-TFII/ARP-1 and USF2 compete for binding to the ATPA regulatory element 1. Thus, the ATPA gene is regulated by a multifunctional binding site through which the transcription factors, COUP-TFII/ARP-1 and USF2, bind and exert their antagonistic effects.

  17. First molecular cloning and gene expression analysis of teleost CD42 (glycoprotein Ib beta chain) GPIb-IX-V subunit from rock bream, Oplegnathus fasciatus.

    Science.gov (United States)

    Jeong, Ji-Min; Kim, Ju-Won; Kim, Do-Hyung; Park, Chan-Il

    2015-04-01

    CD42 is a platelet membrane glycoprotein Ib that plays a key role in haemostasis and thrombin-induced platelet activation. Here, we report the molecular cloning and sequence analysis of the CD42c gene from rock bream (Oplegnathus fasciatus). Rock bream CD42 (RbCD42c) gene expression profiles were determined after infection with Streptococcus iniae, Edwardsiella tarda and red seabream iridovirus (RSIV). The full-length RbCD42c cDNA contained an open reading frame of 624 bp encoding 207 amino acids. The deduced amino acid sequences of the leucine-rich repeat (LRR)-N terminal and LRR-C terminal were conserved between fish and mammals. RbCD42c was highly expressed in red blood cells, spleen, gill, liver and kidney of healthy rock bream. The RbCD42c gene was not significantly up- or downregulated after E. tarda exposure. However, RbCD42c gene expression was upregulated in kidney, spleen and gill after S. iniae infection. RbCD42c was upregulated in spleen, liver and gill, but downregulated in kidney 24 and 48 h after RSIV infection. These results suggest that RbCD42c has different expression patterns after infection with bacterial or viral pathogens. This gene may be directly involved in haemostasis.

  18. To be or not B2B?

    CERN Document Server

    Symons, L J

    2001-01-01

    La question du commerce électronique interentreprises par le web (Business to Business, B2B) est posée actuellement par les grands groupes industriels impliqués dans le commerce mondial. Les prévisions sont imposantes, le B2B atteindra le C.A. de 3000 milliards de dollars en 2003. Les conditions d'accès, la façon de procéder des deux organisateurs (ARIBA et COMMERCE ONE) des plus grandes places de marchés actuelles, sont décrites. La base de l'énorme pyramide est le catalogue électronique multilingue UNSPSC (United Nations Standard Products and Services Classification) et l'organisation ECCMA (Electronic Commerce Code Management Association) qui gère le développement des UNSPSC codes en 8 langues. Dans ce contexte, l'auteur (re)-déclare qu'un des efforts principaux à fournir par le CERN est la création de son propre catalogue électronique. Dans la Division ST, une aide partielle à ce vaste programme pourrait être apportée par la normalisation des codes et désignations des pièces de maint...

  19. What Happened with Spectrometer Magnet 2B

    Energy Technology Data Exchange (ETDEWEB)

    Green, Michael A

    2010-05-27

    The spectrometer solenoid is supposed to be the first magnets installed in MICE [1]-[4]. This report described what happened during the test of the MICE spectrometer solenoid 2B. First, the report describes the temperatures in the magnet, the cooler top plate and the shield during the run where the magnet quenched at 258 A. During this quench, a lead between the bottom of the HTS leads and the diode bank burned out causing the magnet to quench. Second, three methods for measuring the net heat flow into the cold mass are described. Third, there is a discussion of possible resistive heating in the HTS leads between liquid helium temperature and the copper plate, which is at about 50 K. Fourth, there is a discussion of the measured first stage heat loads in the magnet, when there is no current in the magnet. The first stage heat load calculations are based on knowing the first stage temperatures of the three two-stage pulse tube coolers and the single stage GM cooler. Fifth, the estimated heat load to the first stage when the magnet has current in it is discussed. Sixth, there is a comparison of the stage 1 heat loads in magnet 1A [5], magnet 2A [6], and magnet 2B [7]. Finally there is a discussion of recommended changes for improving the spectrometer solenoids so that the coolers can keep them cold.

  20. 微孢子虫核糖体小亚单位RNA(ssUrRNA)基因%Small Subunit Ribosomal RNA Genes of Microsporidia

    Institute of Scientific and Technical Information of China (English)

    王见杨; 黄可威; 毛西成; 赵 昀; 陆长德

    2001-01-01

    微孢子虫是广泛分布于自然界的细胞内原虫类寄生物。它们可寄生于整个生物界。微孢子虫是真核生物,但其核糖体及核糖体RNA(rRNA)为原核生物型。为探讨9种家蚕病原性微孢子虫的种属地位及亲缘关系,对已广泛用于生物进化分类的核糖体小亚单位RNA(asurRNA)基因进行了研究。由微孢子虫ssurRNA基因序列同源性分析所构建的系统进化发育树及Southam杂交分析表明,这9种微孢子虫同为Nosema属,为同属不同种。%Microsporidia are ubiquitous intracellular parasitic protozoa infecting all types of animals. Their ribosomes and rRNAs are of prokaryotic size. In order to better understand their phylogenetic relationship and identify the uncertain species, the sequences of the small subunit ribosomal RNA (ssurRNA, 16 S rRNA) genesof nine microsporidia infectious to the silkworm, Bombyx mori, were determined. The results of phylogenetic trees and Southern blotting suggest all the nine strains of icrosporidia are various species of the genus Nosema.

  1. 2b or Not 2b: Experimental Evolution of Functional Exogenous Sequences in a Plant RNA Virus

    Science.gov (United States)

    Zwart, Mark P.; Ambrós, Silvia; Carrasco, José L.; Elena, Santiago F.

    2017-01-01

    Horizontal gene transfer (HGT) is pervasive in viruses and thought to be a key mechanism in their evolution. On the other hand, strong selective constraints against increasing genome size are an impediment for HGT, rapidly purging horizontally transferred sequences and thereby potentially hindering evolutionary innovation. Here, we explore experimentally the evolutionary fate of viruses with simulated HGT events, using the plant RNA virus Tobacco etch virus (TEV), by separately introducing two functional, exogenous sequences to its genome. One of the events simulates the acquisition of a new function though HGT of a conserved AlkB domain, responsible for the repair of alkylation or methylation damage in many organisms. The other event simulates the acquisition of a sequence that duplicates an existing function, through HGT of the 2b RNA silencing suppressor from Cucumber mosaic virus. We then evolved these two viruses, tracked the maintenance of the horizontally transferred sequences over time, and for the final virus populations, sequenced their genome and measured viral fitness. We found that the AlkB domain was rapidly purged from the TEV genome, restoring fitness to wild-type levels. Conversely, the 2b gene was stably maintained and did not have a major impact on viral fitness. Moreover, we found that 2b is functional in TEV, as it provides a replicative advantage when the RNA silencing suppression domain of HC-Pro is mutated. These observations suggest a potentially interesting role for HGT of short functional sequences in ameliorating evolutionary constraints on viruses, through the duplication of functions. PMID:28137747

  2. The search for mutations in the gene for the beta subunit of the cGMP phosphodiesterase (PDEB) in patients with autosomal recessive retinitis pigmentosa

    DEFF Research Database (Denmark)

    Riess, O; Noerremoelle, A; Weber, B

    1992-01-01

    including 196 bp of the 5' region of the PDEB gene have been assessed for mutations by using single-strand conformational polymorphism analysis in 14 patients from 13 unrelated families with autosomal recessive retinitis pigmentosa (ARRP). No disease-causing mutations were found in this group of affected...... individuals of seven different ancestries. However, a frequent intronic and two exonic polymorphisms (Leu489----Gln and Gly842----Gly) were identified. Segregation analysis using these polymorphic sites excludes linkage of ARRP to the PDEB gene in a family with two affected children....

  3. Molecular basis of positive allosteric modulation of GluN2B NMDA receptors by polyamines.

    Science.gov (United States)

    Mony, Laetitia; Zhu, Shujia; Carvalho, Stéphanie; Paoletti, Pierre

    2011-06-17

    NMDA receptors (NMDARs) form glutamate-gated ion channels that have central roles in neuronal communication and plasticity throughout the brain. Dysfunctions of NMDARs are involved in several central nervous system disorders, including stroke, chronic pain and schizophrenia. One hallmark of NMDARs is that their activity can be allosterically regulated by a variety of extracellular small ligands. While much has been learned recently regarding allosteric inhibition of NMDARs, the structural determinants underlying positive allosteric modulation of these receptors remain poorly defined. Here, we show that polyamines, naturally occurring polycations that selectively enhance NMDARs containing the GluN2B subunit, bind at a dimer interface between GluN1 and GluN2B subunit N-terminal domains (NTDs). Polyamines act by shielding negative charges present on GluN1 and GluN2B NTD lower lobes, allowing their close apposition, an effect that in turn prevents NTD clamshell closure. Our work reveals the mechanistic basis for positive allosteric modulation of NMDARs. It provides the first example of an intersubunit binding site in this class of receptors, a discovery that holds promise for future drug interventions.

  4. The role of transcriptional coactivator ADA2b in Arabidopsis abiotic stress responses.

    Science.gov (United States)

    Vlachonasios, Konstantinos E; Kaldis, Athanasios; Nikoloudi, Adriana; Tsementzi, Despoina

    2011-10-01

    Plant growth and crop production can be greatly affected by common environmental stresses such as drought, high salinity and low temperatures. Gene expression is affected by several abiotic stresses. Stress-inducible genes are regulated by transcription factors and epigenetic mechanisms such as histone modifications. In this Mini-Review, we have explored the role of transcriptional adaptor ADA2b in Arabidopsis responses to abiotic stress. ADA2b is required for the expression of genes involved in abiotic stress either by controlling H3 and H4 acetylation in the case of salt stress or affecting nucleosome occupancy in low temperatures response.

  5. Expression of Kir3 gene and its subunits in human esophageal smooth muscle cells%人食管平滑肌细胞Kir3亚型表达的实验观察

    Institute of Scientific and Technical Information of China (English)

    卢强; 黄立军; 张志培; 刘同刚; 李小飞; 韩勇

    2012-01-01

    OBJECTIVE:To investigate the expression of Kir3 gene and its subunits in esophageal smooth muscles,a series of experiments was designed in this study. The difference of the expression between human esophageal longitudinal muscle(LM) and circular muscle(CM) cells were detected. METHODS:Normal esophageal smooth muscle was selected according to the 24 h esophageal pH testing,esophageal endoscopy.and HE staining. All primers were designed based on human gene sequences. The expression of Kir3. 1 - 3. 4 subunit mRNAs and total proteins were examined in human e-sophageal smooth muscle cells(SMCs) with the methods of reverse transcription polymerase chain reaction(RT-PCR) and western blot. RESULTS:The mRNA expressions of Kir3. 2,Kir3. 3,Kir3. 4 subunits were 0. 121 ±0. 015 and 0. 124± 0. 017,0. 255±0. 018 and 0. 295 ± 0. 028,0. 685 ± 0. 040 and 0. 693 ± 0. 037 respectively. The protein expressions of Kir3. 2.Kir3. 3,Kir3. 4 subunits were 0. 053±0. 010 and 0.068 ± 0.009,0. 160 ± 0.021 and 0. 192 ± 0.032,0.488 + 0.040 and 0. 504 + 0. 033 respectively. The most abundant expression was the Kir3. 4 subunit. The Kir3. 2 and Kir3. 4,Kir3. 3 and Kir3. 4 expression were both significantly differenUP0. 05). CONCLUSIONS:Expression of Kir 3. 2 - 3. 4 subunits was found in human esophageal SMCs except Kir3.1. The Kir3. 4 subunit was the most abundant expression. The expression of Kir3, 2 - 3. 4 was not significantly different in human esophageal LM and CM.%目的:探讨食管下段平滑肌中是否表达Kir3及其各亚型之间表达的差异性,同时明确食管环形平滑肌(CM)与纵行平滑肌(LM)之间表达的区别.方法:根据24 h食管pH值检测、食管镜检查及HE染色结果,分辨、筛选正常人食管平滑肌,按人Kir3.1~3.4序列设计并合成各自的高效引物,利用RT-PCR、蛋白质印迹法检测人食管平滑肌细胞中Kir3的表达及其亚型Kir3.1~3.4表达的差异.结果:在人食管LM、CM层平滑肌细

  6. Differential role of SH2-B and APS in regulating energy and glucose homeostasis.

    Science.gov (United States)

    Li, Minghua; Ren, Decheng; Iseki, Masanori; Takaki, Satoshi; Rui, Liangyou

    2006-05-01

    SH2-B and APS, two members of a pleckstrin homology and SH2 domain-containing adaptor family, promote both insulin and leptin signaling in a similar fashion in cultured cells. In addition, APS mediates insulin-stimulated activation of the c-Cbl/CAP/TC10 pathway in cultured adipocytes. Here we characterized genetically modified mice lacking SH2-B, APS, or both to determine the physiological roles of these two proteins in animals. Disruption of the SH2-B gene resulted in obesity, hyperglycemia, hyperinsulinemia, and glucose intolerance. Conversely, deletion of the APS gene did not alter adiposity, energy balance, and glucose metabolism. Energy intake, energy expenditure, fat content, body weight, and plasma insulin, leptin, glucose, and lipid levels were similar between APS(-/-) and WT littermates fed either normal chow or a high-fat diet. Moreover, deletion of APS failed to alter insulin and glucose tolerance. APS(-/-)/SH2-B(-/-) double knockout mice also developed energy imbalance, obesity, hyperleptinemia, hyperinsulinemia, hyperglycemia, and glucose intolerance; however, plasma leptin and insulin levels were significantly lower in APS(-/-)/SH2-B(-/-) than in SH2-B(-/-) mice. These results suggest that SH2-B, but not APS, is a key positive regulator of energy and glucose metabolism in mice.

  7. Activated CaMKII Couples GluN2B and Casein Kinase 2 to Control Synaptic NMDA Receptors

    Directory of Open Access Journals (Sweden)

    Antonio Sanz-Clemente

    2013-03-01

    Full Text Available Synaptic activity triggers a profound reorganization of the molecular composition of excitatory synapses. For example, NMDA receptors are removed from synapses in an activity- and calcium-dependent manner, via casein kinase 2 (CK2 phosphorylation of the PDZ ligand of the GluN2B subunit (S1480. However, how synaptic activity drives this process remains unclear because CK2 is a constitutively active kinase, which is not directly regulated by calcium. We show here that activated CaMKII couples GluN2B and CK2 to form a trimolecular complex and increases CK2-mediated phosphorylation of GluN2B S1480. In addition, a GluN2B mutant, which contains an insert to mimic the GluN2A sequence and cannot bind to CaMKII, displays reduced S1480 phosphorylation and increased surface expression. We find that although disrupting GluN2B/CaMKII binding reduces synapse number, it increases synaptic-GluN2B content. Therefore, the GluN2B/CaMKII association controls synapse density and PSD composition in an activity-dependent manner, including recruitment of CK2 for the removal of GluN2B from synapses.

  8. Mutations in Nu1, the gene encoding the small subunit of bacteriophage lambda terminase, suppress the postcleavage DNA packaging defect of cosB mutations.

    Science.gov (United States)

    Cai, Z H; Hwang, Y; Cue, D; Catalano, C; Feiss, M

    1997-04-01

    The linear double-stranded DNA molecules in lambda virions are generated by nicking of concatemeric intracellular DNA by terminase, the lambda DNA packaging enzyme. Staggered nicks are introduced at cosN to generate the cohesive ends of virion DNA. After nicking, the cohesive ends are separated by terminase; terminase bound to the left end of the DNA to be packaged then binds the empty protein shell, i.e., the prohead, and translocation of DNA into the prohead occurs. cosB, a site adjacent to cosN, is a terminase binding site. cosB facilitates the rate and fidelity of the cosN cleavage reaction by serving as an anchoring point for gpNu1, the small subunit of terminase. cosB is also crucial for the formation of a stable terminase-DNA complex, called complex I, formed after cosN cleavage. The role of complex I is to bind the prohead. Mutations in cosB affect both cosB functions, causing mild defects in cosN cleavage and severe packaging defects. The lethal cosB R3- R2- R1- mutation contains a transition mutation in each of the three gpNu1 binding sites of cosB. Pseudorevertants of lambda cosB R3- R2- R1- DNA contain suppressor mutations affecting gpNu1. Results of experiments that show that two such suppressors, Nu1ms1 and Nu1ms3, do not suppress the mild cosN cleavage defect caused by the cosB R3- R2- R1- mutation but strongly suppress the DNA packaging defect are presented. It is proposed that the suppressing terminases, unlike the wild-type enzyme, are able to assemble a stable complex I with cosB R3- R2- R1- DNA. Observations on the adenosine triphosphatase activities and protease susceptibilities of gpNu1 of the Nu1ms1 and Nu1ms3 terminases indicate that the conformation of gpNu1 is altered in the suppressing terminases.

  9. Vorticella Linnaeus, 1767 (Ciliophora, Oligohymenophora, Peritrichia) is a grade not a clade: redefinition of Vorticella and the families Vorticellidae and Astylozoidae using molecular characters derived from the gene coding for small subunit ribosomal RNA.

    Science.gov (United States)

    Sun, Ping; Clamp, John; Xu, Dapeng; Kusuoka, Yasushi; Miao, Wei

    2012-01-01

    Recent phylogenetic analyses of the peritrich genus Vorticella have suggested that it might be paraphyletic, with one Vorticella species - Vorticella microstoma grouping with the swimming peritrichs Astylozoon and Opisthonecta in a distant clade. These results were based on very limited taxon sampling and thus could not be accepted as conclusive evidence for revising the generic classification. We tested paraphyly of the genus Vorticella by making a new analysis with a broad range of samples from three continents that yielded 52 new sequences of the gene coding for small subunit rRNA. Our results, together with the available sequences in Genbank, form a comprehensive set of data for the genus Vorticella. Analyses of these data showed that Vorticella microstoma morphotypes, Astylozoon, and Opisthonecta form a well-supported, monophyletic clade, that is distinct from and basal to the family Vorticellidae containing other species of Vorticella. Paraphyly of the genus Vorticella and family Vorticellidae was strongly confirmed by these results. Furthermore, the two clades of Vorticella identified by the SSU rRNA gene are so genetically diverse whereas the genetic distances within the one containing Vorticella microstoma morphotypes, Astylozoon, and Opisthonecta were so slight, which marked it as a separate family that must be defined by molecular characters in the absence of unifying morphological and morphogenetic characters. An emended characterization and status of the genus Vorticella, the families Vorticellidae and Astylozoidae are presented and discussed.

  10. Serotonin 2B receptor: upregulated with age and hearing loss in mouse auditory system.

    Science.gov (United States)

    Tadros, Sherif F; D'Souza, Mary; Zettel, Martha L; Zhu, XiaoXia; Lynch-Erhardt, Martha; Frisina, Robert D

    2007-07-01

    Serotonin (5-HT) is a monoamine neurotransmitter. Serotonin may modulate afferent fiber discharges in the cochlea, inferior colliculus (IC) and auditory cortex. Specific functions of serotonin are exerted upon its interaction with specific receptors; one of those receptors is the serotonin 2B receptor. The aim of this study was to investigate the differences in gene expression of serotonin 2B receptors with age in cochlea and IC, and the possible correlation between gene expression and functional hearing measurements in CBA/CaJ mice. Immunohistochemical examinations of protein expression of IC in mice of different age groups were also performed. Gene expression results showed that serotonin 2B receptor gene was upregulated with age in both cochlea and IC. A significant correlation between gene expression and functional hearing results was established. Immunohistochemical protein expression studies of IC showed more serotonin 2B receptor cells in old mice relative to young adult mice, particularly in the external nucleus. We conclude that serotonin 2B receptors may play a role in the pathogenesis of age-related hearing loss.

  11. ONLINE ACQUISITIONS IN B2B MODEL

    Directory of Open Access Journals (Sweden)

    Constantin SASU

    2016-12-01

    Full Text Available Within the present paper, we propose to review some of the main aspects documented by the academic research so far with regard to the online purchasing behaviour in the B2B online environment (more specifically, in the case of the small and medium-sized companies. Preliminary conclusions reveal that, at least in the case studies, the dominant feature is that the geographic delimitation creates a potential obstacle when it comes to applying a general principle governing the small and medium-sized enterprises. Despite this, we consider that some theoretical elements can be acknowledged, elements from which future research can start to develop theories and hypothesis aimed at better explaining the phenomenon. Furthermore, the fact that the studies are rarely involving the same sector, makes it impossible to generalise the process.

  12. Fragile X mental retardation protein interactions with a G quadruplex structure in the 3'-untranslated region of NR2B mRNA.

    Science.gov (United States)

    Stefanovic, Snezana; DeMarco, Brett A; Underwood, Ayana; Williams, Kathryn R; Bassell, Gary J; Mihailescu, Mihaela Rita

    2015-12-01

    Fragile X syndrome, the most common cause of inherited intellectual disability, is caused by a trinucleotide CGG expansion in the 5'-untranslated region of the FMR1 gene, which leads to the loss of expression of the fragile X mental retardation protein (FMRP). FMRP, an RNA-binding protein that regulates the translation of specific mRNAs, has been shown to bind a subset of its mRNA targets by recognizing G quadruplex structures. It has been suggested that FMRP controls the local protein synthesis of several protein components of the post synaptic density (PSD) in response to specific cellular needs. We have previously shown that the interactions between FMRP and mRNAs of the PSD scaffold proteins PSD-95 and Shank1 are mediated via stable G-quadruplex structures formed within the 3'-untranslated regions of these mRNAs. In this study we used biophysical methods to show that a comparable G quadruplex structure forms in the 3'-untranslated region of the glutamate receptor subunit NR2B mRNA encoding for a subunit of N-methyl-d-aspartate (NMDA) receptors that is recognized specifically by FMRP, suggesting a common theme for FMRP recognition of its dendritic mRNA targets.

  13. Evidence for multiple promoters of the human IL-5 receptor alpha subunit gene: a novel 6-base pair element determines cell-specific promoter function.

    Science.gov (United States)

    Zhang, J; Kuvelkar, R; Cheewatrakoolpong, B; Williams, S; Egan, R W; Billah, M M

    1997-12-01

    In addition to a previously characterized promoter (P1), we now show the existence of a second promoter for the human IL-5Ralpha gene. Initially, a genomic region (P2) 5' upstream of human IL-5Ralpha exon 2 was cloned by an inverted PCR. The transcriptional start site was then mapped to a deoxycytidine (C) residue within P2 by analyzing cellular mRNA with both the 5' rapid amplification of cDNA end-PCR and S1 nuclease protection assays. Transfection of eosinophilic HL-60 cells with reporter gene constructs in which either P1 or P2 was linked to the bacterial chloramphenicol acetyltransferase (CAT) gene resulted in CAT expression; little or no CAT expression occurred in other myeloid and nonmyeloid cell lines. Deletion studies showed that a 66-bp region, ranging from -31 to +35, was sufficient to promote CAT expression in eosinophilic HL-60 cells. Analysis of linker-scanning mutants identified a novel 6-bp element (5' CTAATT 3') spanning -19 to -14 that was essential for P2 promoter activity. In electrophoretic mobility shift assays, a P2 region from -31 to +1 containing the unique 6-bp element, when used as a probe, formed a complex with a protein(s) that was found only in the eosinophilic cell line. This binding activity was lost upon replacement of the 6-bp element with a 6-bp linker, suggesting that this element likely serves as the binding site for an eosinophilic HL-60 cell-specific transcription factor(s). Together, these data suggest an important role for P2 promoter in the regulation of eosinophil-specific expression of the human IL-5 receptor alpha gene.

  14. Mutation in cpsf6/CFIm68 (Cleavage and Polyadenylation Specificity Factor Subunit 6) causes short 3'UTRs and disturbs gene expression in developing embryos, as revealed by an analysis of primordial germ cell migration using the medaka mutant naruto.

    Science.gov (United States)

    Sasado, Takao; Kondoh, Hisato; Furutani-Seiki, Makoto; Naruse, Kiyoshi

    2017-01-01

    Our previous studies analyzing medaka mutants defective in primordial germ cell (PGC) migration identified cxcr4b and cxcr7, which are both receptors of the chemokine sdf1/cxcl12, as key regulators of PGC migration. Among PGC migration mutants, naruto (nar) is unique in that the mutant phenotype includes gross morphological abnormalities of embryos, suggesting that the mutation affects a broader range of processes. A fine genetic linkage mapping and genome sequencing showed the nar gene encodes Cleavage and Polyadenylation Specificity Factor subunit 6 (CPSF6/CFIm68). CPSF6 is a component of the Cleavage Factor Im complex (CFIm) which plays a key role in pre-mRNA 3'-cleavage and polyadenylation. 3'RACE of sdf1a/b and cxcr7 transcripts in the mutant embryos indicated shorter 3'UTRs with poly A additions occurring at more upstream positions than wild-type embryos, suggesting CPSF6 functions to prevent premature 3'UTR cleavage. In addition, expression of the coding region sequences of sdf1a/b in nar mutants was more anteriorly extended in somites than wild-type embryos, accounting for the abnormally extended distribution of PGCs in nar mutants. An expected consequence of shortening 3'UTR is the escape from the degradation mechanism mediated by microRNAs interacting with distal 3'UTR sequence. The abnormal expression pattern of sdf1a coding sequence may be at least partially accounted for by this mechanism. Given the pleiotropic effects of nar mutation, further analysis using the nar mutant will reveal processes in which CPSF6 plays essential regulatory roles in poly A site selection and involvement of 3'UTRs in posttranscriptional gene regulation in various genes in vivo.

  15. Heterogeneity in hand veins responses to acetylcholine is not associated with polymorphisms in the G-protein beta3-subunit (C825T) and endothelial nitric oxide synthase (G894T) genes but with serum low density lipoprotein cholesterol.

    Science.gov (United States)

    Grossmann, M; Dobrev, D; Siffert, W; Kirch, W

    2001-06-01

    Vascular responses to acetylcholine (ACh) are notoriously variable, the reason for this phenomenon is unknown. We tested the hypothesis that the variability in venous response to acetylcholine may be associated with two recently identified genetic polymorphisms for proteins involved in the signal transduction pathway, i.e. the G-protein beta3-subunit (GNB3) and endothelial nitric oxide synthase (eNOS). The dorsal hand vein technique was used in 37 healthy subjects. Hand veins were preconstricted with the alpha1-adrenoceptor agonist phenylephrine and the venodilator response to local ACh infusion was measured with and without comedication of acetylsalicylic acid or co-infusion of N(G)-monomethyl-L-arginine (L-NMMA). In addition, all subjects received routine laboratory tests and 26 of them were genotyped for the C825T polymorphism of the GNB3 gene and for the G894T polymorphism of the eNOS gene. A striking variability in venous response to ACh was found with dilation observed in the low ACh concentration range and reduced dilation or even constriction at high concentrations. ACh-induced venodilation was mediated by muscarinic receptors and abolished in the presence of both acetylsalicylic acid and L-NMMA suggesting dependence on endothelium. We did not find any association of the variability in ACh response with GNB3 or eNOS allele status. On the other hand, a significant positive correlation between ACh responsiveness and low density lipoprotein-cholesterol status was detected. Two recently discovered gene polymorphisms are not responsible for the profound heterogeneity in venodilator response to ACh. Surprisingly, this variability appears to relate to the lipid status of the subjects. The exact nature of this new finding requires further study.

  16. Mutation in cpsf6/CFIm68 (Cleavage and Polyadenylation Specificity Factor Subunit 6) causes short 3'UTRs and disturbs gene expression in developing embryos, as revealed by an analysis of primordial germ cell migration using the medaka mutant naruto

    Science.gov (United States)

    Kondoh, Hisato; Furutani-Seiki, Makoto; Naruse, Kiyoshi

    2017-01-01

    Our previous studies analyzing medaka mutants defective in primordial germ cell (PGC) migration identified cxcr4b and cxcr7, which are both receptors of the chemokine sdf1/cxcl12, as key regulators of PGC migration. Among PGC migration mutants, naruto (nar) is unique in that the mutant phenotype includes gross morphological abnormalities of embryos, suggesting that the mutation affects a broader range of processes. A fine genetic linkage mapping and genome sequencing showed the nar gene encodes Cleavage and Polyadenylation Specificity Factor subunit 6 (CPSF6/CFIm68). CPSF6 is a component of the Cleavage Factor Im complex (CFIm) which plays a key role in pre-mRNA 3'-cleavage and polyadenylation. 3'RACE of sdf1a/b and cxcr7 transcripts in the mutant embryos indicated shorter 3’UTRs with poly A additions occurring at more upstream positions than wild-type embryos, suggesting CPSF6 functions to prevent premature 3’UTR cleavage. In addition, expression of the coding region sequences of sdf1a/b in nar mutants was more anteriorly extended in somites than wild-type embryos, accounting for the abnormally extended distribution of PGCs in nar mutants. An expected consequence of shortening 3'UTR is the escape from the degradation mechanism mediated by microRNAs interacting with distal 3’UTR sequence. The abnormal expression pattern of sdf1a coding sequence may be at least partially accounted for by this mechanism. Given the pleiotropic effects of nar mutation, further analysis using the nar mutant will reveal processes in which CPSF6 plays essential regulatory roles in poly A site selection and involvement of 3'UTRs in posttranscriptional gene regulation in various genes in vivo. PMID:28253363

  17. Visual recognition memory is related to basic expression level of NMDA receptor NR1/NR2B subtype in hippocampus and striatum of rats

    Institute of Scientific and Technical Information of China (English)

    Shu-jun XU; Zhong CHEN; Li-jun ZHU; Hai-qing SHEN; Jian-hong LUO

    2005-01-01

    Aim: To examine the basic expression levels of N-methyl-D-aspartate (NMDA) receptor NR1 and NR2B subunits in six brain regions of Sprague-Dawley (SD) rats with different visual recognition memory. Methods: Rats were tested by a novelobject-recognition model and grouped into the high and the low visual recognition memory groups. The expression levels of NR1 and NR2B subunits in the cortex, hippocampus, striatum, amygdala, diencephalon, and olfactory bulb were measured by semiquantitative immunoblotting. Results: The NR1 and NR2B subunit protein levels in the hippocampus of the high visual recognition memory group were 35.9% (P<0.01) and 53.4% (P<0.05) higher respectively than those in the low group. In addition, the NR2B level in the striatum in the high visual recognition memory group was 25.0% (P<0.05) higher than that in the low one. However, no significant difference was found in the levels of the subunits between the two groups in other brain regions. Conclusion: The visual recognition memory in rats is related to the basic expression level of NMDA receptor NR1/NR2B subtype in the hippocampus and striatum.

  18. Mutation screening of GRIN2B in schizophrenia and autism spectrum disorder in a Japanese population

    Science.gov (United States)

    Takasaki, Yuto; Koide, Takayoshi; Wang, Chenyao; Kimura, Hiroki; Xing, Jingrui; Kushima, Itaru; Ishizuka, Kanako; Mori, Daisuke; Sekiguchi, Mariko; Ikeda, Masashi; Aizawa, Miki; Tsurumaru, Naoko; Iwayama, Yoshimi; Yoshimi, Akira; Arioka, Yuko; Yoshida, Mami; Noma, Hiromi; Oya-Ito, Tomoko; Nakamura, Yukako; Kunimoto, Shohko; Aleksic, Branko; Uno, Yota; Okada, Takashi; Ujike, Hiroshi; Egawa, Jun; Kuwabara, Hitoshi; Someya, Toshiyuki; Yoshikawa, Takeo; Iwata, Nakao; Ozaki, Norio

    2016-01-01

    N-methyl-d-aspartate receptors (NMDARs) play a critical role in excitatory synaptic transmission and plasticity in the central nervous systems. Recent genetics studies in schizophrenia (SCZ) show that SCZ is susceptible to NMDARs and the NMDAR signaling complex. In autism spectrum disorder (ASD), several studies report dysregulation of NMDARs as a risk factor for ASD. To further examine the association between NMDARs and SCZ/ASD development, we conducted a mutation screening study of GRIN2B which encodes NR2B subunit of NMDARs, to identify rare mutations that potentially cause diseases, in SCZ and ASD patients (n = 574 and 152, respectively). This was followed by an association study in a large sample set of SCZ, ASD, and normal healthy controls (n = 4145, 381, and 4432, respectively). We identified five rare missense mutations through the mutation screening of GRIN2B. Although no statistically significant association between any single mutation and SCZ or ASD was found, one of its variant, K1292R, is found only in the patient group. To further examine the association between mutations in GRIN2B and SCZ/ASD development, a larger sample size and functional experiments are needed. PMID:27616045

  19. Genetic analysis of the cytoplasmic dynein subunit families.

    Directory of Open Access Journals (Sweden)

    K Kevin Pfister

    2006-01-01

    Full Text Available Cytoplasmic dyneins, the principal microtubule minus-end-directed motor proteins of the cell, are involved in many essential cellular processes. The major form of this enzyme is a complex of at least six protein subunits, and in mammals all but one of the subunits are encoded by at least two genes. Here we review current knowledge concerning the subunits, their interactions, and their functional roles as derived from biochemical and genetic analyses. We also carried out extensive database searches to look for new genes and to clarify anomalies in the databases. Our analysis documents evolutionary relationships among the dynein subunits of mammals and other model organisms, and sheds new light on the role of this diverse group of proteins, highlighting the existence of two cytoplasmic dynein complexes with distinct cellular roles.

  20. Encephalomyocarditis virus viroporin 2B activates NLRP3 inflammasome.

    Directory of Open Access Journals (Sweden)

    Minako Ito

    Full Text Available Nod-like receptors (NLRs comprise a large family of intracellular pattern- recognition receptors. Members of the NLR family assemble into large multiprotein complexes, termed the inflammasomes. The NLR family, pyrin domain-containing 3 (NLRP3 is triggered by a diverse set of molecules and signals, and forms the NLRP3 inflammasome. Recent studies have indicated that both DNA and RNA viruses stimulate the NLRP3 inflammasome, leading to the secretion of interleukin 1 beta (IL-1β and IL-18 following the activation of caspase-1. We previously demonstrated that the proton-selective ion channel M2 protein of influenza virus activates the NLRP3 inflammasome. However, the precise mechanism by which NLRP3 recognizes viral infections remains to be defined. Here, we demonstrate that encephalomyocarditis virus (EMCV, a positive strand RNA virus of the family Picornaviridae, activates the NLRP3 inflammasome in mouse dendritic cells and macrophages. Although transfection with RNA from EMCV virions or EMCV-infected cells induced robust expression of type I interferons in macrophages, it failed to stimulate secretion of IL-1β. Instead, the EMCV viroporin 2B was sufficient to cause inflammasome activation in lipopolysaccharide-primed macrophages. While cells untransfected or transfected with the gene encoding the EMCV non-structural protein 2A or 2C expressed NLRP3 uniformly throughout the cytoplasm, NLRP3 was redistributed to the perinuclear space in cells transfected with the gene encoding the EMCV 2B or influenza virus M2 protein. 2B proteins of other picornaviruses, poliovirus and enterovirus 71, also caused the NLRP3 redistribution. Elevation of the intracellular Ca(2+ level, but not mitochondrial reactive oxygen species and lysosomal cathepsin B, was important in EMCV-induced NLRP3 inflammasome activation. Chelation of extracellular Ca(2+ did not reduce virus-induced IL-1β secretion. These results indicate that EMCV activates the NLRP3 inflammasome by

  1. Identification of a novel HMW glutenin subunit and comparison of its amino acid sequence with those of homologous subunits

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Aegilops tauschii is the donor of the D genome of common wheat (Triticum aestivum). Genetic variation of HMW glutenin subunits encoded by the Glu-1Dt locus of Ae. tauschii has been found to be higher than that specified by the Glu-1D locus in common wheat. In the present note, we report the identification of a novel HMW glutenin subunit, Dy13t, from Ae. tauschii. The newly identified subunit possessed an electrophoretic mobility that was faster than that of the Dy12 subunit of common wheat. The complete ORF of encoding the Dy13t subunit contained 624 codons (excluding the stop codons). The amino acid sequence deduced from the Dy13t gene ORF was the shortest among those of the previously reported subunits derived by the D genome. A further comparison of Dy13t amino acid sequence with those of the subunits characterized from the A, B, D, R genomes of Triticeae showed that the smaller size of the Dy13t subunit was associated with a reduction in the size of its repetitive domain.

  2. Study of genetic diversity of eukaryotic picoplankton in different oceanic regions by small-subunit rRNA gene cloning and sequencing.

    Science.gov (United States)

    Díez, B; Pedrós-Alió, C; Massana, R

    2001-07-01

    Very small eukaryotic organisms (picoeukaryotes) are fundamental components of marine planktonic systems, often accounting for a significant fraction of the biomass and activity in a system. Their identity, however, has remained elusive, since the small cells lack morphological features for identification. We determined the diversity of marine picoeukaryotes by sequencing cloned 18S rRNA genes in five genetic libraries from North Atlantic, Southern Ocean, and Mediterranean Sea surface waters. Picoplankton were obtained by filter size fractionation, a step that excluded most large eukaryotes and recovered most picoeukaryotes. Genetic libraries of eukaryotic ribosomal DNA were screened by restriction fragment length polymorphism analysis, and at least one clone of each operational taxonomic unit (OTU) was partially sequenced. In general, the phylogenetic diversity in each library was rather great, and each library included many different OTUs and members of very distantly related phylogenetic groups. Of 225 eukaryotic clones, 126 were affiliated with algal classes, especially the Prasinophyceae, the Prymnesiophyceae, the Bacillariophyceae, and the Dinophyceae. A minor fraction (27 clones) was affiliated with clearly heterotrophic organisms, such as ciliates, the chrysomonad Paraphysomonas, cercomonads, and fungi. There were two relatively abundant novel lineages, novel stramenopiles (53 clones) and novel alveolates (19 clones). These lineages are very different from any organism that has been isolated, suggesting that there are previously unknown picoeukaryotes. Prasinophytes and novel stramenopile clones were very abundant in all of the libraries analyzed. These findings underscore the importance of attempts to grow the small eukaryotic plankton in pure culture.

  3. 5-HT7 receptor is coupled to G alpha subunits of heterotrimeric G12-protein to regulate gene transcription and neuronal morphology.

    Science.gov (United States)

    Kvachnina, Elena; Liu, Guoquan; Dityatev, Alexander; Renner, Ute; Dumuis, Aline; Richter, Diethelm W; Dityateva, Galina; Schachner, Melitta; Voyno-Yasenetskaya, Tatyana A; Ponimaskin, Evgeni G

    2005-08-24

    The neurotransmitter serotonin (5-HT) plays an important role in the regulation of multiple events in the CNS. We demonstrated recently a coupling between the 5-HT4 receptor and the heterotrimeric G13-protein resulting in RhoA-dependent neurite retraction and cell rounding (Ponimaskin et al., 2002). In the present study, we identified G12 as an additional G-protein that can be activated by another member of serotonin receptors, the 5-HT7 receptor. Expression of 5-HT7 receptor induced constitutive and agonist-dependent activation of a serum response element-mediated gene transcription through G12-mediated activation of small GTPases. In NIH3T3 cells, activation of the 5-HT7 receptor induced filopodia formation via a Cdc42-mediated pathway correlating with RhoA-dependent cell rounding. In mouse hippocampal neurons, activation of the endogenous 5-HT7 receptors significantly increased neurite length, whereas stimulation of 5-HT4 receptors led to a decrease in the length and number of neurites. These data demonstrate distinct roles for 5-HT7R/G12 and 5-HT4R/G13 signaling pathways in neurite outgrowth and retraction, suggesting that serotonin plays a prominent role in regulating the neuronal cytoarchitecture in addition to its classical role as neurotransmitter.

  4. Association study of polymorphisms in the alpha 7 nicotinic acetylcholine receptor subunit and catechol-o-methyl transferase genes with sensory gating in first-episode schizophrenia.

    Science.gov (United States)

    Liu, Xia; Hong, Xiaohong; Chan, Raymond C K; Kong, Fanzhi; Peng, Zhizhen; Wan, Xiaona; Wang, Changqing; Cheng, Lu

    2013-10-30

    The purpose of the current study was to explore the association of auditory P50 sensory gating (P50) and prepulse inhibition (PPI) of schizophrenia with polymorphisms in the CHRNA7 and COMT genes. One hundred and fourty patients with schizophrenia participated in this study. They were administered the tests P50 and PPI. Moreover, three single nucleotide polymorphisms (SNPs) (rs2337980, rs1909884 and rs883473) in CHRNA7 and three SNPs (rs4680, rs737865 and rs165599) in COMT were selected to be genotyped by polyacrylamide gel microarray techniques. P50 index showed significant reduction in S2 amplitude between wild-type and mutation groups in the COMT rs4680. S1 amplitude of mutation group in the COMT rs737865 was also lower compared to wild-type group. PPI index revealed a shorter pulse latency of mutation group in the rs4680. The suppression ratio of mutation group was lower in COMT rs165599. Negative findings were shown between comparisons in all the CHRNA7 SNPs. We find that P50 and PPI may be influenced by COMT rs4680 polymorphisms in schizophrenia; more excitingly, we find that P50 might be influenced by COMT rs737865 polymorphisms and PPI may be influenced by COMT rs165599 polymorphisms in schizophrenia, and their mutations are associated with the reduction of the risk of P50 or PPI defects in schizophrenia. Futher studies with a larger number of subjects are needed to verify the present findings.

  5. Morphology, morphogenesis and small subunit rRNA gene sequence of a soil hypotrichous ciliate, Perisincirra paucicirrata (Ciliophora, Kahliellidae), from the shoreline of the Yellow River, North China.

    Science.gov (United States)

    Li, Fengchao; Xing, Yi; Li, Jiamei; Al-Rasheid, Khaled A S; He, Songke; Shao, Chen

    2013-01-01

    The morphology, morphogenesis, and 18S rRNA gene sequence of a soil hypotrichous ciliate Perisincirra paucicirrata, isolated from north China, were investigated. Perisincirra paucicirrata differs from its congeners in: (1) having a body length to width ratio in vivo of 4:1, (2) its adoral zone occupying between 15% and 25% of the total body length, and (3) the presence of two parabuccal cirri, three left (with 10-16 cirri each) and two right marginal rows (with 14-24 cirri each), and three dorsal kineties. Our study offers a first attempt to begin to map the morphogenetic processes of the genus, which are mainly characterised by the following: the formation of four frontal ventral transverse anlagens for each daughter cell, with the proter's anlage I originating from the reorganised anterior part of the parental paroral; the paroral and endoral anlage developed from the reorganised old endoral and do not contribute the first frontal cirrus; the frontoventral transverse anlage I contributing the left frontal cirrus; anlage II generating the middle frontal and the buccal cirri; anlage III developing the right frontal cirrus and the anterior parabuccal cirrus; and anlage IV contributing the posterior parabuccal cirrus. As an additional contribution, we judge that the inner one or the two right rows of P. kahli and P. longicirrata are marginal rows. Phylogenetic analysis based on SSU rDNA sequences suggests that Perisincirra is related to sporadotrichids, but provides no credible evidence for its taxonomic position.

  6. Prevalence of microsporidiosis due to Enterocytozoon bieneusi and Encephalitozoon (Septata) intestinalis among patients with AIDS-related diarrhea: determination by polymerase chain reaction to the microsporidian small-subunit rRNA gene.

    Science.gov (United States)

    Coyle, C M; Wittner, M; Kotler, D P; Noyer, C; Orenstein, J M; Tanowitz, H B; Weiss, L M

    1996-11-01

    Microsporidia are emerging as opportunistic pathogens in patients with AIDS. Enterocytozoon bieneusi and Encephalitozoon (Septata) intestinalis have been implicated in enteric infections in AIDS patients with chronic diarrhea, a wasting syndrome, and malabsorption. We used the polymerase chain reaction (PCR) and primers that amplify the conserved regions of the small-subunit rRNA (SSU-rRNA) gene of E. bieneusi and E. intestinalis in tissue specimens from HIV-infected patients with and without diarrhea to examine the association between microsporidia and diarrhea in patients with AIDS. Tissue specimens were obtained from 68 patients with AIDS and diarrhea (mean CD4 lymphocyte count, 21/mm3) and 43 AIDS patients without diarrhea (mean CD4 lymphocyte count, 60/mm3). By means of PCR with use of the SSU-rRNA primers specific for E. bieneusi and E. intestinalis, we found that 44% of patients with diarrhea were infected with microsporidia, whereas only 2.3% of the patients without diarrhea were infected with microsporidia (P < .001). There was a clear association between the presence of microsporidia and diarrhea. In addition, the SSU-rRNA primers proved to be sensitive and specific when used in this clinical setting.

  7. The Arabidopsis gene DIG6 encodes a large 60S subunit nuclear export GTPase 1 that is involved in ribosome biogenesis and affects multiple auxin-regulated development processes

    KAUST Repository

    Zhao, Huayan

    2015-08-13

    The circularly permuted GTPase large subunit GTPase 1 (LSG1) is involved in the maturation step of the 60S ribosome and is essential for cell viability in yeast. Here, an Arabidopsis mutant dig6 (drought inhibited growth of lateral roots) was isolated. The mutant exhibited multiple auxin-related phenotypes, which included reduced lateral root number, altered leaf veins, and shorter roots. Genetic mapping combined with next-generation DNA sequencing identified that the mutation occurred in AtLSG1-2. This gene was highly expressed in regions of auxin accumulation. Ribosome profiling revealed that a loss of function of AtLSG1-2 led to decreased levels of monosomes, further demonstrating its role in ribosome biogenesis. Quantitative proteomics showed that the expression of certain proteins involved in ribosome biogenesis was differentially regulated, indicating that ribosome biogenesis processes were impaired in the mutant. Further investigations showed that an AtLSG1-2 deficiency caused the alteration of auxin distribution, response, and transport in plants. It is concluded that AtLSG1-2 is integral to ribosome biogenesis, consequently affecting auxin homeostasis and plant development.

  8. Molecular and functional characterization of seven Na+/K+-ATPase β subunit paralogs in Senegalese sole (Solea senegalensis Kaup, 1858).

    Science.gov (United States)

    Armesto, Paula; Infante, Carlos; Cousin, Xavier; Ponce, Marian; Manchado, Manuel

    2015-04-01

    In the present work, seven genes encoding Na(+),K(+)-ATPase (NKA) β-subunits in the teleost Solea senegalensis are described for the first time. Sequence analysis of the predicted polypeptides revealed a high degree of conservation with those of other vertebrate species and maintenance of important motifs involved in structure and function. Phylogenetic analysis clustered the seven genes into four main clades: β1 (atp1b1a and atp1b1b), β2 (atp1b2a and atp1b2b), β3 (atp1b3a and atp1b3b) and β4 (atp1b4). In juveniles, all paralogous transcripts were detected in the nine tissues examined albeit with different expression patterns. The most ubiquitous expressed gene was atp1b1a whereas atp1b1b was mainly detected in osmoregulatory organs (gill, kidney and intestine), and atp1b2a, atp1b2b, atp1b3a, atp1b3b and atp1b4 in brain. An expression analysis in three brain regions and pituitary revealed that β1-type transcripts were more abundant in pituitary than the other β paralogs with slight differences between brain regions. Quantification of mRNA abundance in gills after a salinity challenge showed an activation of atp1b1a and atp1b1b at high salinity water (60 ppt) and atp1b3a and atp1b3b in response to low salinity (5 ppt). Transcriptional analysis during larval development showed specific expression patterns for each paralog. Moreover, no differences in the expression profiles between larvae cultivated at 10 and 35 ppt were observed except for atp1b4 with higher mRNA levels at 10 than 35 ppt at 18 days post hatch. Whole-mount in situ hybridization analysis revealed that atp1b1b was mainly localized in gut, pronephric tubule, gill, otic vesicle, and chordacentrum of newly hatched larvae. All these data suggest distinct roles of NKA β subunits in tissues, during development and osmoregulation with β1 subunits involved in the adaptation to hyperosmotic conditions and β3 subunits to hypoosmotic environments.

  9. Knockdown of BNST GluN2B-containing NMDA receptors mimics the actions of ketamine on novelty-induced hypophagia.

    Science.gov (United States)

    Louderback, K M; Wills, T A; Muglia, L J; Winder, D G

    2013-12-03

    Administration of a single low dose of the N-methyl-D-aspartate (NMDA) receptor antagonist ketamine has been demonstrated to elicit long-lasting antidepressant effects in humans with depression, as well as in rodent models of depression. Although pharmacological studies have implicated the GluN2B subunit of the NMDA receptor in these effects, drugs targeting this subunit have off-target actions, and systemic administration of these compounds does not allow for delineation of specific brain regions involved. In this study, we assessed the role of GluN2B in the bed nucleus of the stria terminalis (BNST) in novelty-induced hypophagia (NIH) in mice. First, we verified that ketamine, as well as the GluN2B antagonist Ro25-6981, decreased the latency to consume food in a novel environment in a version of the NIH test. We then hypothesized that GluN2B-containing receptors within the BNST may be a target of systemic ketamine and contribute to behavioral effects. Through the combination of a GluN2B-floxed mouse line and stereotaxic delivery of lentiviral Cre recombinase, we found that targeted knockdown of this subunit within the BNST mimicked the reduction in affective behavior observed with systemic ketamine or Ro25-6981 in the NIH test. These data suggest a role for GluN2B-containing NMDARs within the BNST in the affective effects of systemic ketamine.

  10. TRUST IN B2B E-MARKETPLACES

    Directory of Open Access Journals (Sweden)

    SEBASTIAN KOT

    2011-01-01

    Full Text Available The paper presents background of B2B exchanges and review of their forms and functionalities. The benefits and fails reasons are noticed. European enterprises interest in B2B trade is next aspect of consideration. Finally, the trust barriers of B2B exchanges are presented.

  11. 7 CFR 301.80-2b - Exempted articles. 1

    Science.gov (United States)

    2010-01-01

    ... 7 Agriculture 5 2010-01-01 2010-01-01 false Exempted articles. 1 301.80-2b Section 301.80-2b....80-2b Exempted articles. 1 1 The articles hereby exempted remain subject to applicable restrictions under other quarantines. (a) The following articles are exempt from the certification and permit...

  12. 7 CFR 301.85-2b - Exempted articles. 1

    Science.gov (United States)

    2010-01-01

    ... 7 Agriculture 5 2010-01-01 2010-01-01 false Exempted articles. 1 301.85-2b Section 301.85-2b... § 301.85-2b Exempted articles. 1 1 The articles hereby exempted remain subject to applicable restrictions under other quarantines and other provisions of this subpart. (a) The following articles...

  13. Genetics Home Reference: CHMP2B-related frontotemporal dementia

    Science.gov (United States)

    ... Conditions CHMP2B-related frontotemporal dementia CHMP2B-related frontotemporal dementia Enable Javascript to view the expand/collapse boxes. ... Open All Close All Description CHMP2B -related frontotemporal dementia is a progressive brain disorder that affects personality, ...

  14. Cadmium treatment suppresses DNA polymerase δ catalytic subunit gene expression by acting on the p53 and Sp1 regulatory axis.

    Science.gov (United States)

    Antoniali, Giulia; Marcuzzi, Federica; Casarano, Elena; Tell, Gianluca

    2015-11-01

    Cadmium (Cd) is a carcinogenic and neurotoxic environmental pollutant. Among the proposed mechanisms for Cd toxic effects, its ability to promote oxidative stress and to inhibit, in vitro, the activities of some Base Excision DNA Repair (BER) enzymes, such as hOGG1, XRCC1 and APE1, have been already established. However, the molecular mechanisms at the basis of these processes are largely unknown especially at sub-lethal doses of Cd and no information is available on the effect of Cd on the expression levels of BER enzymes. Here, we show that non-toxic treatment of neuronal cell lines, with pro-mitogenic doses of Cd, promotes a significant time- and dose-dependent down-regulation of DNA polymerase δ (POLD1) expression through a transcriptional mechanism with a modest effect on Polβ, XRCC1 and APE1. We further elucidated that the observed transcriptional repression on Polδ is acted by through competition by activated p53 on Sp1 at POLD1 promoter and by a squelching effect. We further proved the positive effect of Sp1 not only on POLD1 expression but also on Polβ, XRCC1 and APE1 expression, suggesting that Sp1 has pleiotropic effects on the whole BER pathway. Our results indicated that Cd-mediated impairment of BER pathway, besides acting on the enzymatic functions of some key proteins, is also exerted at the gene expression level of Polδ by acting on the p53-Sp1 regulatory axis. These data may explain not only the Cd-induced neurotoxic effects but also the potential carcinogenicity of this heavy metal.

  15. Complementation of Escherichia coli unc mutant strains by chloroplast and cyanobacterial F1-ATPase subunits.

    Science.gov (United States)

    Lill, H; Burkovski, A; Altendorf, K; Junge, W; Engelbrecht, S

    1993-10-04

    The genes encoding the five subunits of the F1 portion of the ATPases from both spinach chloroplasts and the cyanobacterium Synechocystis sp. PCC 6803 were cloned into expression vectors and expressed in Escherichia coli. The recombinant subunits formed inclusion bodies within the cells. Each particular subunit was expressed in the respective unc mutant, each unable to grow on non-fermentable carbon sources. The following subunits restored growth under conditions of oxidative phosphorylation: alpha (both sources, cyanobacterial subunit more than spinach subunit), beta (cyanobacterial subunit only), delta (both spinach and Synechocystis), and epsilon (both sources), whereas no growth was achieved with the gamma subunits from both sources. Despite a high degree of sequence homology the large subunits alpha and beta of spinach and cyanobacterial F1 were not as effective in the substitution of their E. coli counterparts. On the other hand, the two smallest subunits of the E. coli ATPase could be more effectively replaced by their cyanobacterial or chloroplast counterparts, although the sequence identity or even similarity is very low. We attribute these findings to the different roles of these subunits in F1: The large alpha and beta subunits contribute to the catalytic centers of the enzyme, a function rendering them very sensitive to even minor changes. For the smaller delta and epsilon subunits it was sufficient to maintain a certain tertiary structure during evolution, with little emphasis on the conservation of particular amino acids.

  16. Quantitative analysis of interferon alpha receptor subunit 1 and suppressor of cytokine signaling 1 gene transcription in blood cells of patients with chronic hepatitis C

    Directory of Open Access Journals (Sweden)

    Sedeño-Monge Virginia

    2010-09-01

    Full Text Available Abstract Background Interferon (IFN-α receptor 1 (ifnar1 and suppressor of cytokine signaling 1 (socs1 transcription levels were quantified in peripheral blood mononuclear cells (PBMC of 59 patients infected with hepatitis C virus (HCV and 17 non-infected individuals. Samples were obtained from patients infected with HCV that were either untreated or treated with IFN-α2 plus ribavirin for 1 year and divided into responders and non-responders based on viral load reduction 6 months after treatment. Ifnar1 and socs1 transcription was quantified by real-time RT-PCR, and the fold difference (2-ΔΔCT with respect to hprt housekeeping gene was calculated. Results Ifnar1 transcription increased significantly in HCV-infected patients either untreated (3.26 ± 0.31, responders (3.1 ± 0.23 and non-responders (2.18 ± 0.23 with respect to non-infected individuals (1 ± 0.34; P = 0.005. Ifnar1 transcription increased significantly (P = 0.003 in patients infected with HCV genotypes 1a (4.74 ± 0.25 and 1b (2.81 ± 0.25 but not in 1a1b (1.58 ± 0.21. No association was found of Ifnar1 transcription with disease progress, initial viral load or other clinical factors. With respect to socs1 transcription, values were similar for non-infected individuals (1 ± 0.28 and untreated patients (0.99 ± 0.41 but increased in responders (2.81 ± 0.17 and non-responder patients (1.67 ± 0.41. Difference between responder and non-responder patients was not statistically significant. Socs1 transcription increased in patients infected with HCV genotypes 1a and 1b (2.87 ± 0.45 and 2.22 ± 0.17, respectively but not in 1a1b (1.28 ± 0.40. Socs1 transcript was absent in three patients infected with HCV genotype 1b. A weak correlation between ifnar1 and socs1 transcription was found, when Spearman's correlation coefficient was calculated. Conclusion Our results suggest that HCV infection may up-regulate ifnar1 transcription. HCV genotypes differ in their capacity to affect

  17. Mapping of a conformational epitope on the cashew allergen Ana o 2: a discontinuous large subunit epitope dependent upon homologous or heterologous small subunit association.

    Science.gov (United States)

    Xia, Lixin; Willison, LeAnna N; Porter, Lauren; Robotham, Jason M; Teuber, Suzanne S; Sathe, Shridhar K; Roux, Kenneth H

    2010-05-01

    The 11S globulins are members of the cupin protein superfamily and represent an important class of tree nut allergens for which a number of linear epitopes have been mapped. However, specific conformational epitopes for these allergens have yet to be described. We have recently reported a cashew Ana o 2 conformational epitope defined by murine mAb 2B5 and competitively inhibited by a subset of patient IgE antibodies. The 2B5 epitope appears to reside on the large (acidic) subunit, is dependent upon small (basic) subunit association for expression, and is highly susceptible to denaturation. Here we fine map the epitope using a combination of recombinant chimeric cashew Ana o 2-soybean Gly m 6 chimeras, deletion and point mutations, molecular modeling, and electron microscopy of 2B5-Ana o 2 immune complexes. Key residues appear confined to a 24 amino acid segment near the N-terminus of the large subunit peptide, a portion of which makes direct contact with the small subunit. These data provide an explanation for both the small subunit dependence and the structurally labile nature of the epitope.

  18. Mutations in SH3PXD2B cause Borrone dermato-cardio-skeletal syndrome.

    Science.gov (United States)

    Wilson, Gabrielle R; Sunley, Jasmine; Smith, Katherine R; Pope, Kate; Bromhead, Catherine J; Fitzpatrick, Elizabeth; Di Rocco, Maja; van Steensel, Maurice; Coman, David J; Leventer, Richard J; Delatycki, Martin B; Amor, David J; Bahlo, Melanie; Lockhart, Paul J

    2014-06-01

    Borrone Dermato-Cardio-Skeletal (BDCS) syndrome is a severe progressive autosomal recessive disorder characterized by coarse facies, thick skin, acne conglobata, dysmorphic facies, vertebral abnormalities and mitral valve prolapse. We identified a consanguineous kindred with a child clinically diagnosed with BDCS. Linkage analysis of this family (BDCS1) identified five regions homozygous by descent with a maximum LOD score of 1.75. Linkage analysis of the family that originally defined BDCS (BDCS3) identified an overlapping linkage peak at chromosome 5q35.1. Sequence analysis identified two different homozygous mutations in BDCS1 and BDCS3, affecting the gene encoding the protein SH3 and PX domains 2B (SH3PXD2B), which localizes to 5q35.1. Western blot analysis of patient fibroblasts deri