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Sample records for nr1 subunit gene

  1. PRODUCTION AND PURIFICATION OF IgY ANTIBODIES AS A NOVEL TOOL TO PURIFY THE NR1 SUBUNIT OF NMDA RECEPTO

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    Edgar Antonio Reyes Montaño

    2011-12-01

    Full Text Available Producing polyclonal antibodies (IgY inchickens has advantages over those obtainedin other animal models, since theyhave been used as a tool for studyingdifferent proteins (NMDA glutamate receptorin our case, specifically the NR1subunit. We produced specific antibodiesagainst expression products by thealternative splicing of the gene encodingNMDA receptor NR1 subunit in adult ratbrain. Three peptides corresponding tothe splicing sites (N1, C1 and C2’ cassetteswere designed, synthesised and usedindividually as antigens in hens. Specificimmunoglobulins were purified fromyolks. The antibodies were then used forpurifying the NMDA receptor NR1 subunitusing affinity chromatography couplingthe three antibodies to the support.R

  2. Spinal D1-like dopamine receptors modulate NMDA receptor-induced hyperexcitability and NR1 subunit phosphorylation at serine 889.

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    Aira, Zigor; Barrenetxea, Teresa; Buesa, Itsaso; Martínez, Endika; Azkue, Jon Jatsu

    2016-04-01

    Activation of the N-methyl-d-aspartate receptor (NMDAR) in dorsal horn neurons is recognized as a fundamental mechanism of central sensitization and pathologic pain. This study assessed the influence of dopaminergic, D1-like receptor-mediated input to the spinal dorsal horn on NMDAR function. Spinal superfusion with selective NMDAR agonist cis-ACPD significantly increased C-fiber-evoked field potentials in rats subjected to spinal nerve ligation (SNL), but not in sham-operated rats. Simultaneous application of D1LR antagonist SCH 23390 dramatically reduced hyperexcitability induced by cis-ACPD. Furthermore, cis-ACPD-induced hyperexcitability seen in nerve-ligated rats could be mimicked in unin-jured rats during stimulation of D1LRs by agonist SKF 38393 at subthreshold concentration. Phosphorylation of NMDAR subunit NR1 at serine 889 at postsynaptic sites was found to be increased in dorsal horn neurons 90 min after SNL, as assessed by increased co-localization with postsynaptic marker PSD-95. Increased NR1 phosphorylation was attenuated in the presence of SCH 23390 in the spinal superfusate. The present results support that D1LRs regulate most basic determinants of NMDAR function in dorsal horn neurons, suggesting a potential mechanism whereby dopaminergic input to the dorsal horn can modulate central sensitization and pathologic pain.

  3. Postsynaptic density levels of the NMDA receptor NR1 subunit and PSD-95 protein in prefrontal cortex from people with schizophrenia.

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    Catts, Vibeke Sørensen; Derminio, Dominique Suzanne; Hahn, Chang-Gyu; Weickert, Cynthia Shannon

    2015-01-01

    There is converging evidence of involvement of N-methyl-d-aspartate (NMDA) receptor hypofunction in the pathophysiology of schizophrenia. Our group recently identified a decrease in total NR1 mRNA and protein expression in the dorsolateral prefrontal cortex in a case-control study of individuals with schizophrenia (n=37/group). The NR1 subunit is critical to NMDA receptor function at the postsynaptic density, a cellular structure rich in the scaffolding protein, PSD-95. The extent to which the NMDA receptor NR1 subunit is altered at the site of action, in the postsynaptic density, is not clear. To extend our previous results by measuring levels of NR1 and PSD-95 protein in postsynaptic density-enriched fractions of prefrontal cortex from the same individuals in the case-control study noted above. Postsynaptic density-enriched fractions were isolated from fresh-frozen prefrontal cortex (BA10) and subjected to western blot analysis for NR1 and PSD-95. We found a 20% decrease in NR1 protein (t(66)=-2.874, P=0.006) and a 30% decrease in PSD-95 protein (t(63)=-2.668, P=0.010) in postsynaptic density-enriched fractions from individuals with schizophrenia relative to unaffected controls. Individuals with schizophrenia have less NR1 protein, and therefore potentially fewer functional NMDA receptors, at the postsynaptic density. The associated decrease in PSD-95 protein at the postsynaptic density suggests that not only are glutamate receptors compromised in individuals with schizophrenia, but the overall spine architecture and downstream signaling supported by PSD-95 may also be deficient.

  4. Association of NR1I2 gene polymorphisms and time of progression to AIDS

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    de Medeiros, Rúbia Marília; Menti, Carolina Fialho; Benelli, Jéssica Louise; Matte, Maria Cristina Cotta; de Melo, Marineide Gonçalves; Almeida, Sabrina Esteves de Matos; Fiegenbaum, Marilu

    2017-01-01

    BACKGROUND The time of progression towards AIDS can vary greatly among seropositive patients, and may be associated with host genetic variation. The NR1I2 (PXR) gene, a ligand-activated transcription factor, regulates the transcription immune pathway genes and can therefore be targets of viral replication mechanisms influencing time of progression to AIDS. OBJECTIVE To verify the association of single nucleotide polymorphisms (SNPs) rs3814057, rs6785049, rs7643645, and rs2461817 in the NR1I2 (PXR) gene with progression to AIDS in HIV-1 infected patients. METHODS Blood samples were obtained from 96 HIV-1 positive individuals following informed consent. DNA was isolated and genotyped through real time polymerase chain reaction (PCR) for the presence of SNPs in the NR1I2. Questionnaires on socio-demographic features and behaviors were answered and time of progression to AIDS was estimated based on medical chart analysis. FINDINGS Patients with the GG genotype for rs7643645 were shown to be related with a more rapid disease progression when compared to GA and AA genotypes. This result was maintained by the Multivariate Cox Regression considering sex, ethnicity, and presence of HLA-B*57, HLA-B*27, and CCR5del32 polymorphisms. MAIN CONCLUSIONS Recent studies reported the expression of the nuclear receptors in T-Lymphocytes, suggesting their possible role in the immune response. In addition, nuclear receptors have been shown to inhibit the HIV replication, although no such mechanism has been thoroughly elucidated to date. This is the first time an association between NR1I2 polymorphism and time of progression to AIDS is reported and supports an apparent relationship between the gene in the immune response and identifies another genetic factor influencing AIDS progression. PMID:28327790

  5. Investigation of associations between NR1D1, RORA and RORB genes and bipolar disorder.

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    Yin-Chieh Lai

    Full Text Available Several genes that are involved in the regulation of circadian rhythms are implicated in the susceptibility to bipolar disorder (BD. The current study aimed to investigate the relationships between genetic variants in NR1D1 RORA, and RORB genes and BD in the Han Chinese population. We conducted a case-control genetic association study with two samples of BD patients and healthy controls. Sample I consisted of 280 BD patients and 200 controls. Sample II consisted of 448 BD patients and 1770 healthy controls. 27 single nucleotide polymorphisms in the NR1D1, RORA, and RORB genes were genotyped using GoldenGate VeraCode assays in sample I, and 492 markers in the three genes were genotyped using Affymetrix Genome-Wide CHB Array in sample II. Single marker and gene-based association analyses were performed using PLINK. A combined p-value for the joining effects of all markers within a gene was calculated using the rank truncated product method. Multifactor dimensionality reduction (MDR method was also applied to test gene-gene interactions in sample I. All markers were in Hardy-Weinberg equilibrium (P>0.001. In sample I, the associations with BD were observed for rs4774388 in RORA (OR = 1.53, empirical p-value, P = 0.024, and rs1327836 in RORB (OR = 1.75, P = 0.003. In Sample II, there were 45 SNPs showed associations with BD, and the most significant marker in RORA was rs11639084 (OR = 0.69, P = 0.002, and in RORB was rs17611535 (OR = 3.15, P = 0.027. A combined p-value of 1.6×10-6, 0.7, and 1.0 was obtained for RORA, RORB and NR1D1, respectively, indicting a strong association for RORA with the risk of developing BD. A four way interaction was found among markers in NR1D1, RORA, and RORB with the testing accuracy 53.25% and a cross-validation consistency of 8 out of 10. In sample II, 45 markers had empirical p-values less than 0.05. The most significant markers in RORA and RORB genes were rs11639084 (OR = 0.69, P = 0.002, and rs17611535 (OR = 3

  6. Bovine NR1I3 gene polymorphisms and its association with feed efficiency traits in Nellore cattle

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    Pâmela A. Alexandre

    2014-12-01

    Full Text Available The Nuclear receptor 1 family I member 3 (NR1I3, also known as the Constitutive Androstane Receptor (CAR, was initially characterized as a key regulator of xenobiotic metabolism. However, recent biochemical and structural data suggest that NR1I3 is activated in response to metabolic and nutritional stress in a ligand-independent manner. Thus, we prospected the Bovine NR1I3 gene for polymorphisms and studied their association with feed efficiency traits in Nellore cattle. First, 155 purebred Nellore bulls were individually measured for Residual Feed Intake (RFI and the 25 best (High Feed Efficiency group, HFE and the 25 worst animals (Low Feed Efficiency group, LFE were selected for DNA extraction. The entire Bovine NR1I3 gene was amplified and polymorphisms were identified by sequencing. Then, one SNP different between HFE and LFE groups was genotyped in all the 155 animals and in another 288 animals totalizing 443 Nellore bulls genotyped for association of NR1I3 SNPs with feed efficiency traits. We found 24 SNPs in the NR1I3 gene and choose a statistically different SNP between HFE and LFE groups for further analysis. Genotyping of the 155 animals showed a significant association within SNP and RFI (p = 0.04, Residual Intake and BW Gain (p = 0.04 and Dry Matter Intake (p = 0.01. This SNP is located in the 5′flanking promoter region of NR1I3 gene and different alleles alter the binding site for predicted transcriptional factors as HNF4alpha, CREM and c-MYB, leading us to conclude that NR1I3 expression and regulation might be important to feed efficiency.

  7. Role of a circadian-relevant gene NR1D1 in brain development: possible involvement in the pathophysiology of autism spectrum disorders

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    Goto, Masahide; Mizuno, Makoto; Matsumoto, Ayumi; Yang, Zhiliang; Jimbo, Eriko F.; Tabata, Hidenori; Yamagata, Takanori; Nagata, Koh-ichi

    2017-01-01

    In our previous study, we screened autism spectrum disorder (ASD) patients with and without sleep disorders for mutations in the coding regions of circadian-relevant genes, and detected mutations in several clock genes including NR1D1. Here, we further screened ASD patients for NR1D1 mutations and identified three novel mutations including a de novo heterozygous one c.1499 G > A (p.R500H). We then analyzed the role of Nr1d1 in the development of the cerebral cortex in mice. Acute knockdown of mouse Nr1d1 with in utero electroporation caused abnormal positioning of cortical neurons during corticogenesis. This aberrant phenotype was rescued by wild type Nr1d1, but not by the c.1499 G > A mutant. Time-lapse imaging revealed characteristic abnormal migration phenotypes in Nr1d1-deficient cortical neurons. When Nr1d1 was knocked down, axon extension and dendritic arbor formation of cortical neurons were also suppressed while proliferation of neuronal progenitors and stem cells at the ventricular zone was not affected. Taken together, Nr1d1 was found to play a pivotal role in corticogenesis via regulation of excitatory neuron migration and synaptic network formation. These results suggest that functional defects in NR1D1 may be related to ASD etiology and pathophysiology. PMID:28262759

  8. A neuroligin-1-derived peptide stimulates phosphorylation of the NMDA receptor NR1 subunit and rescues MK-801-induced decrease in long-term potentiation and memory impairment

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    Korshunova, Irina; Gjørlund, Michelle D; Jacobsen, Sylwia Owczarek

    2015-01-01

    neurolide-1 effects on short- and long-term social and spatial memory in social recognition, Morris water-maze, and Y-maze tests. We found that subcutaneous neurolide-1 administration, restored hippocampal LTP compromised by NMDA receptor inhibitor MK-801. It counteracted MK-801-induced memory deficit...... in the water-maze and Y-maze tests after long-term treatment (24 h and 1-2 h before the test), but not after short-term exposure (1-2 h). Long-term exposure to neurolide-1 also facilitated social recognition memory. In addition, neurolide-1-induced phosphorylation of the NMDA receptor NR1 subunit on a site...... receptor phosphorylation after treatment with NL1 or a mimetic peptide, neurolide-1, was quantified by immunoblotting. Subsequently, we investigated effects of neurolide-1 on long-term potentiation (LTP) induction in hippocampal slices compromised by NMDA receptor inhibitor MK-801. Finally, we investigated...

  9. Mechanisms responsible for the effect of median nerve electrical stimulation on traumatic brain injury-induced coma: orexin-A-mediated N-methyl-D-aspartate receptor subunit NR1 upregulation

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    Zhen Feng

    2016-01-01

    Full Text Available Electrical stimulation of the median nerve is a noninvasive technique that facilitates awakening from coma. In rats with traumatic brain injury-induced coma, median nerve stimulation markedly enhances prefrontal cortex expression of orexin-A and its receptor, orexin receptor 1. To further understand the mechanism underlying wakefulness mediated by electrical stimulation of the median nerve, we evaluated its effects on the expression of the N-methyl-D-aspartate receptor subunit NR1 in the prefrontal cortex in rat models of traumatic brain injury-induced coma, using immunohistochemistry and western blot assays. In rats with traumatic brain injury, NR1 expression increased with time after injury. Rats that underwent electrical stimulation of the median nerve (30 Hz, 0.5 ms, 1.0 mA for 15 minutes showed elevated NR1 expression and greater recovery of consciousness than those without stimulation. These effects were reduced by intracerebroventricular injection of the orexin receptor 1 antagonist SB334867. Our results indicate that electrical stimulation of the median nerve promotes recovery from traumatic brain injury-induced coma by increasing prefrontal cortex NR1 expression via an orexin-A-mediated pathway.

  10. Modifier genes as therapeutics: the nuclear hormone receptor Rev Erb alpha (Nr1d1 rescues Nr2e3 associated retinal disease.

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    Nelly M Cruz

    Full Text Available Nuclear hormone receptors play a major role in many important biological processes. Most nuclear hormone receptors are ubiquitously expressed and regulate processes such as metabolism, circadian function, and development. They function in these processes to maintain homeostasis through modulation of transcriptional gene networks. In this study we evaluate the effectiveness of a nuclear hormone receptor gene to modulate retinal degeneration and restore the integrity of the retina. Currently, there are no effective treatment options for retinal degenerative diseases leading to progressive and irreversible blindness. In this study we demonstrate that the nuclear hormone receptor gene Nr1d1 (Rev-Erbα rescues Nr2e3-associated retinal degeneration in the rd7 mouse, which lacks a functional Nr2e3 gene. Mutations in human NR2E3 are associated with several retinal degenerations including enhanced S cone syndrome and retinitis pigmentosa. The rd7 mouse, lacking Nr2e3, exhibits an increase in S cones and slow, progressive retinal degeneration. A traditional genetic mapping approach previously identified candidate modifier loci. Here, we demonstrate that in vivo delivery of the candidate modifier gene, Nr1d1 rescues Nr2e3 associated retinal degeneration. We observed clinical, histological, functional, and molecular restoration of the rd7 retina. Furthermore, we demonstrate that the mechanism of rescue at the molecular and functional level is through the re-regulation of key genes within the Nr2e3-directed transcriptional network. Together, these findings reveal the potency of nuclear receptors as modulators of disease and specifically of NR1D1 as a novel therapeutic for retinal degenerations.

  11. Selective up-regulation of NMDA-NR1 receptor expression in myenteric plexus after TNBS induced colitis in rats

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    Price Donald D

    2006-01-01

    Full Text Available Abstract Background N-methyl-D-aspartic acid (NMDA spinal cord receptors play an important role in the development of hyperalgesia following inflammation. It is unclear, however, if changes in NMDA subunit receptor gene expression in the colonic myenteric plexus are associated with colonic inflammation. We investigated regulation of NMDA-NR1 receptor gene expression in TNBS induced colitis in rats. Male Sprague-Dawley rats (150 g–250 g were treated with 20 mg trinitrobenzene sulfonic acid (TNBS diluted in 50% ethanol. The agents were delivered with a 24 gauge catheter inserted into the lumen of the colon. The animals were sacrificed at 2, 7, 14, 21, and 28 days after induction of the colitis, their descending colon was retrieved for reverse transcription-polymerase chain reaction; a subset of animals' distal colon was used for two-dimensional (2-D western analysis and immunocytochemistry. Results NR1-exon 5 (N1 and NR1-exon 21 (C1 appeared 14, 21 and 28 days after TNBS treatment. NR1 pan mRNA was up-regulated at 14, 21, and 28 days. The NR1-exon 22 (C2 mRNA did not show significant changes. Using 2-D western analysis, untreated control rats were found to express only NR1001 whereas TNBS treated rats expressed NR1001, NR1011, and NR1111. Immunocytochemistry demonstrated NR1-N1 and NR1-C1 to be present in the myenteric plexus of TNBS treated rats. Conclusion These results suggest a role for colonic myenteric plexus NMDA receptors in the development of neuronal plasticity and visceral hypersensitivity in the colon. Up-regulation of NMDA receptor subunits may reflect part of the basis for chronic visceral hypersensitivity in conditions such as post-infectious irritable bowel syndrome.

  12. NMDA受体亚单位NR1在离体人神经干细胞中的表达%The expression of N-methyl-D-aspartate receptors subunit 1 in the human hippocampus neural stem cells in vitro

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    胡忠浩; 王姗姗; 徐铁军

    2011-01-01

    目的 研究离体培养的人海马神经干细胞(NSCs)中NMDA受体亚单位NR1的表达.方法 分离培养胎龄8~12周人胚脑海马神经干细胞,对NSCs进行Nestin和分化鉴定.用免疫细胞化学、Western blot免疫印迹和RT-PCR等方法检测原代、传代1次、传代2次的人胚胎海马NSCs中NR1蛋白和mRNA的表达.结果 在孕8~12周人胚脑海马分离培养的NSCs中,NMDA受体亚单位NR1免疫细胞化学反应呈阳性,该受体亚单位的蛋白和mRNA均被检测到.结论 体外培养的早期人胚胎海马NSCs能稳定表达NMDA受体亚单位NR1.%Objective To investigate the expression of N - methyl - D - aspartate (NMDA) receptor subunit 1 in the cuhured human hippocampus neural stem cells (NSCs). Methods NSCs from hippocampus of abortion human embryonic tissue (8 -12 weeks post conception) were cultured and passaged in suspension. The neurospheres were identified by Nestin immunocytochemistry and the differentiation potential of NSCs were also detected. The protein and mRNA expression of NMDA receptor subunit 1 in the human hippocampus NSCs of primitive, passage 1 and 2 were measured by immunocytochemistry, Western blot and reverse transcription polymerase chain reaction (RT -PCR). Results NRI immunostaining positive cells could be found in the human hippocampus NSCs. The protein and mRNA of NR1 were identified in the same cells. Conclusion NSCs from the human embryonic hippocampus can express NMDA receptor subunitl in vitro.

  13. Genome-wide identification of nuclear receptor (NR) genes and the evolutionary significance of the NR1O subfamily in the monogonont rotifer Brachionus spp.

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    Kim, Duck-Hyun; Kim, Hui-Su; Hwang, Dae-Sik; Kim, Hee-Jin; Hagiwara, Atsushi; Lee, Jae-Seong; Jeong, Chang-Bum

    2017-10-01

    Nuclear receptors (NRs) are a large family of transcription factors that are involved in many fundamental biological processes. NRs are considered to have originated from a common ancestor, and are highly conserved throughout the whole animal taxa. Therefore, the genome-wide identification of NR genes in an animal taxon can provide insight into the evolutionary tendencies of NRs. Here, we identified all the NR genes in the monogonont rotifer Brachionus spp., which are considered an ecologically key species due to their abundance and world-wide distribution. The NR family was composed of 40, 32, 29, and 32 genes in the genomes of the rotifers B. calyciflorus, B. koreanus, B. plicatilis, and B. rotundiformis, respectively, which were classified into seven distinct subfamilies. The composition of each subfamily was highly conserved between species, except for NR1O genes, suggesting that they have undergone sporadic evolutionary processes for adaptation to their different environmental pressures. In addition, despite the dynamics of NR evolution, the significance of the conserved endocrine system, particularly for estrogen receptor (ER)-signaling, in rotifers was discussed on the basis of phylogenetic analyses. The results of this study may help provide a better understanding the evolution of NRs, and expand our knowledge of rotifer endocrine systems. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. Gene targeting of CK2 catalytic subunits.

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    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.

  15. Gene targeting of CK2 catalytic subunits

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    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

  16. Conserved expression of the glutamate NMDA receptor 1 subunit splice variants during the development of the Siberian hamster suprachiasmatic nucleus.

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    Giles E Duffield

    Full Text Available Glutamate neurotransmission and the N-methyl-D-aspartate receptor (NMDAR are central to photic signaling to the master circadian pacemaker located in the hypothalamic suprachiasmatic nucleus (SCN. NMDARs also play important roles in brain development including visual input circuits. The functional NMDAR is comprised of multiple subunits, but each requiring the NR1 subunit for normal activity. The NR1 can be alternatively spliced to produce isoforms that confer different functional properties on the NMDAR. The SCN undergoes extensive developmental changes during postnatal life, including synaptogenesis and acquisition of photic signaling. These changes are especially important in the highly photoperiodic Siberian hamster, in which development of sensitivity to photic cues within the SCN could impact early physiological programming. In this study we examined the expression of NR1 isoforms in the hamster at different developmental ages. Gene expression in the forebrain was quantified by in situ hybridization using oligonucleotide probes specific to alternatively spliced regions of the NR1 heteronuclear mRNA, including examination of anterior hypothalamus, piriform cortex, caudate-putamen, thalamus and hippocampus. Gene expression analysis within the SCN revealed the absence of the N1 cassette, the presence of the C2 cassette alone and the combined absence of C1 and C2 cassettes, indicating that the dominant splice variants are NR1-2a and NR1-4a. Whilst we observe changes at different developmental ages in levels of NR1 isoform probe hybridization in various forebrain structures, we find no significant changes within the SCN. This suggests that a switch in NR1 isoform does not underlie or is not produced by developmental changes within the hamster SCN. Consistency of the NR1 isoforms would ensure that the response of the SCN cells to photic signals remains stable throughout life, an important aspect of the function of the SCN as a responder to

  17. Conserved expression of the glutamate NMDA receptor 1 subunit splice variants during the development of the Siberian hamster suprachiasmatic nucleus.

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    Duffield, Giles E; Mikkelsen, Jens D; Ebling, Francis J P

    2012-01-01

    Glutamate neurotransmission and the N-methyl-D-aspartate receptor (NMDAR) are central to photic signaling to the master circadian pacemaker located in the hypothalamic suprachiasmatic nucleus (SCN). NMDARs also play important roles in brain development including visual input circuits. The functional NMDAR is comprised of multiple subunits, but each requiring the NR1 subunit for normal activity. The NR1 can be alternatively spliced to produce isoforms that confer different functional properties on the NMDAR. The SCN undergoes extensive developmental changes during postnatal life, including synaptogenesis and acquisition of photic signaling. These changes are especially important in the highly photoperiodic Siberian hamster, in which development of sensitivity to photic cues within the SCN could impact early physiological programming. In this study we examined the expression of NR1 isoforms in the hamster at different developmental ages. Gene expression in the forebrain was quantified by in situ hybridization using oligonucleotide probes specific to alternatively spliced regions of the NR1 heteronuclear mRNA, including examination of anterior hypothalamus, piriform cortex, caudate-putamen, thalamus and hippocampus. Gene expression analysis within the SCN revealed the absence of the N1 cassette, the presence of the C2 cassette alone and the combined absence of C1 and C2 cassettes, indicating that the dominant splice variants are NR1-2a and NR1-4a. Whilst we observe changes at different developmental ages in levels of NR1 isoform probe hybridization in various forebrain structures, we find no significant changes within the SCN. This suggests that a switch in NR1 isoform does not underlie or is not produced by developmental changes within the hamster SCN. Consistency of the NR1 isoforms would ensure that the response of the SCN cells to photic signals remains stable throughout life, an important aspect of the function of the SCN as a responder to environmental changes

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

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    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.

  19. Nuclear receptor NR1H3 in familial multiple sclerosis

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    Wang, Zhe; Sadovnick, A. Dessa; Traboulsee, Anthony L.; Ross, Jay P.; Bernales, Cecily Q.; Encarnacion, Mary; Yee, Irene M.; de Lemos, Madonna; Greenwood, Talitha; Lee, Joshua D.; Wright, Galen; Ross, Colin J.; Zhang, Si; Song, Weihong; Vilariño-Güell, Carles

    2016-01-01

    SUMMARY Multiple sclerosis (MS) is an inflammatory disease characterized by myelin loss and neuronal dysfunction. Despite the aggregation observed in some families, pathogenic mutations have remained elusive. In this study we describe the identification of NR1H3 p.Arg415Gln in seven MS patients from two multi-incident families presenting severe and progressive disease, with an average age at onset of 34 years. Additionally, association analysis of common variants in NR1H3 identified rs2279238 conferring a 1.35-fold increased risk of developing progressive MS. The p.Arg415Gln position is highly conserved in orthologs and paralogs, and disrupts NR1H3 heterodimerization and transcriptional activation of target genes. Protein expression analysis revealed that mutant NR1H3 (LXRA) alters gene expression profiles, suggesting a disruption in transcriptional regulation as one of the mechanisms underlying MS pathogenesis. Our study indicates that pharmacological activation of LXRA or its targets may lead to effective treatments for the highly debilitating and currently untreatable progressive phase of MS. PMID:27253448

  20. Repeated ketamine administration alters N-methyl-d-aspartic acid receptor subunit gene expression: Implication of genetic vulnerability for ketamine abuse and ketamine psychosis in humans

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    Lipsky, Robert H

    2015-01-01

    For more than 40 years following its approval by the Food and Drug Administration (FDA) as an anesthetic, ketamine, a non-competitive N-methyl-d-aspartic acid (NMDA) receptor antagonist, has been used as a tool of psychiatric research. As a psychedelic drug, ketamine induces psychotic symptoms, cognitive impairment, and mood elevation, which resemble some symptoms of schizophrenia. Recreational use of ketamine has been increasing in recent years. However, little is known of the underlying molecular mechanisms responsible for ketamine-associated psychosis. Recent animal studies have shown that repeated ketamine administration significantly increases NMDA receptor subunit gene expression, in particular subunit 1 (NR1 or GluN1) levels. This results in neurodegeneration, supporting a potential mechanism where up-regulation of NMDA receptors could produce cognitive deficits in chronic ketamine abuse patients. In other studies, NMDA receptor gene variants are associated with addictive behavior. Here, we focus on the roles of NMDA receptor gene subunits in ketamine abuse and ketamine psychosis and propose that full sequencing of NMDA receptor genes may help explain individual vulnerability to ketamine abuse and ketamine-associated psychosis. PMID:25245072

  1. A common polymorphism in NR1H2 (LXRbeta is associated with preeclampsia

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    Brouillet Jean-Paul

    2011-10-01

    Full Text Available Abstract Background Preeclampsia is a frequent complication of pregnancy and a leading cause of perinatal mortality. Both genetic and environmental risk factors have been identified. Lipid metabolism, particularly cholesterol metabolism, is associated with this disease. Liver X receptors alpha (NR1H3, also known as LXRalpha and beta (NR1H2, also known as LXRbeta play a key role in lipid metabolism. They belong to the nuclear receptor superfamily and are activated by cholesterol derivatives. They have been implicated in preeclampsia because they modulate trophoblast invasion and regulate the expression of the endoglin (CD105 gene, a marker of preeclampsia. The aim of this study was to investigate associations between the NR1H3 and NR1H2 genes and preeclampsia. Methods We assessed associations between single nucleotide polymorphisms of NR1H3 (rs2279238 and rs7120118 and NR1H2 (rs35463555 and rs2695121 and the disease in 155 individuals with preeclampsia and 305 controls. Genotypes were determined by high-resolution melting analysis. We then used a logistic regression model to analyze the different alleles and genotypes for those polymorphisms as a function of case/control status. Results We found no association between NR1H3 SNPs and the disease, but the NR1H2 polymorphism rs2695121 was found to be strongly associated with preeclampsia (genotype C/C: adjusted odds ratio, 2.05; 95% CI, 1.04-4.05; p = 0.039 and genotype T/C: adjusted odds ratio, 1.85; 95% CI, 1.01-3.42; p = 0.049. Conclusions This study provides the first evidence of an association between the NR1H2 gene and preeclampsia, adding to our understanding of the links between cholesterol metabolism and this disease.

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

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    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

  3. 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.

  4. Allotopic Expression of a Gene Encoding FLAG Tagged-subunit 8 of Yeast Mitochondrial ATP Synthase

    Directory of Open Access Journals (Sweden)

    I MADE ARTIKA

    2006-03-01

    Full Text Available Subunit 8 of yeast mitochondrial ATP synthase is a polypeptide of 48 amino acids encoded by the mitochondrial ATP8 gene. A nuclear version of subunit 8 gene has been designed to encode FLAG tagged-subunit 8 fused with a mitochondrial signal peptide. The gene has been cloned into a yeast expression vector and then expressed in a yeast strain lacking endogenous subunit 8. Results showed that the gene was successfully expressed and the synthesized FLAG tagged-subunit 8 protein was imported into mitochondria. Following import, the FLAG tagged-subunit 8 protein assembled into functional mitochondrial ATP synthase complex. Furthermore, the subunit 8 protein could be detected using anti-FLAG tag monoclonal antibody.

  5. Analysis list: Nr1d2 [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Nr1d2 Liver + mm9 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Nr1d2.1.tsv... http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Nr1d2.5.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Nr1d...2.10.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/colo/Nr1d2.Liver.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/colo/Liver.gml ...

  6. Analysis list: Nr1h3 [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Nr1h3 Liver + mm9 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Nr1h3.1.tsv... http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Nr1h3.5.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Nr1h...3.10.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/colo/Nr1h3.Liver.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/colo/Liver.gml ...

  7. Analysis list: Nr1h2 [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Nr1h2 Blood + mm9 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Nr1h2.1.tsv... http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Nr1h2.5.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Nr1h...2.10.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/colo/Nr1h2.Blood.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/colo/Blood.gml ...

  8. Analysis list: Nr1d1 [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Nr1d1 Adipocyte,Cardiovascular,Liver,Neural + mm9 http://dbarchive.biosciencedbc.jp.../kyushu-u/mm9/target/Nr1d1.1.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/target/Nr1d1.5.tsv http://db...archive.biosciencedbc.jp/kyushu-u/mm9/target/Nr1d1.10.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/colo/Nr1d1....Adipocyte.tsv,http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/colo/Nr1d1.C...ardiovascular.tsv,http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/colo/Nr1d1.Liver.tsv,http://dbarchive.biosc

  9. Potentiation of glycine-gated NR1/NR3A NMDA receptors relieves Ca2+-dependent outward rectification

    Directory of Open Access Journals (Sweden)

    Christian Madry

    2010-03-01

    Full Text Available Glycine has diverse functions within the mammalian central nervous system. It inhibits postsynaptic neurons via strychnine-sensitive glycine receptors (GlyRs and enhances neuronal excitation through co-activation of N-methyl-D-aspartate (NMDA receptors. Classical Ca2+-permeable NMDA receptors are composed of glycine-binding NR1 and glutamate-binding NR2 subunits, and hence require both glutamate and glycine for efficient activation. In contrast, recombinant receptors composed of NR1 and the glycine binding NR3A and/or NR3B subunits lack glutamate binding sites and can be activated by glycine alone. Therefore these receptors are also named excitatory glycine receptors. Co-application of antagonists of the NR1 glycine-binding site or of the divalent cation Zn2+ markedly enhances the glycine responses of these receptors. To gain further insight into the properties of these glycine-gated NMDA receptors, we investigated their current-voltage (I-V dependence. Whole-cell current-voltage relations of glycine currents recorded from NR1/NR3B and NR1/NR3A/NR3B expressing oocytes were found to be linear under our recording conditions. In contrast, NR1/NR3A receptors displayed a strong outwardly rectifying I-V relation. Interestingly, the voltage-dependent inward current block was abolished in the presence of NR1 antagonists, Zn2+ or a combination of both. Further analysis revealed that Ca2+ (1.8 mM present in our recording solutions was responsible for the voltage-dependent inhibition of ion flux through NR1/NR3A receptors. Since physiological concentrations of the divalent cation Mg2+ did not affect the I-V dependence, our data suggest that relief of the voltage-dependent Ca2+ block of NR1/NR3A receptors by Zn2+ may be important for the regulation of excitatory glycinergic transmission, according to the Mg2+-block of conventional NR1/NR2 NMDA receptors.

  10. Expression of NR1I3 in mouse lung tumors induced by the tobacco-specific nitrosamine 4-(methylnitrosamino)-4-(3-pyridyl)-1-butanone

    Energy Technology Data Exchange (ETDEWEB)

    Fukumasu, H.; Cordeiro, Y.G.; Rochetti, A.L.; Barra, C.N.; Sámora, T.S.; Strefezzi, R.F. [Laboratório de Oncologia Comparada e Translacional, Departmento de Medicina Veterinária, Faculdade de Zootecnia e Engenharia de Alimentos, Universidade de São Paulo, Pirassununga, SP (Brazil); Dagli, M.L.Z. [Laboratório de Oncologia Experimental e Comparada, Departmento de Patologia, Faculdade de Medicina Veterinária e Zootecnia, Universidade de São Paulo, São Paulo, SP (Brazil)

    2015-02-13

    Nuclear receptor subfamily 1, group I, member 3 (NR1I3) is reported to be a possible novel therapeutic target for some cancers, including lung, brain and hematopoietic tumors. Here, we characterized expression of NR1I3 in a mouse model of lung carcinogenesis induced by 4-(methylnitrosamino)-4-(3-pyridyl)-1-butanone (NNK), the most potent tobacco carcinogen. Lung tumors were collected from mice treated with NNK (400 mg/kg) and euthanized after 52 weeks. Benign and malignant lesions were formalin-fixed and paraffin-embedded for histology and immunohistochemistry, with samples snap-frozen for mRNA analysis. Immunohistochemically, we found that most macrophages and type I and II pneumocytes expressed NR1I3, whereas fibroblasts and endothelial cells were NR1I3{sup −}. Compared with benign lesions, malignant lesions had less NR1I3{sup +} tumor cells. Gene expression analysis also showed an inverse correlation between NR1I3 mRNA expression and tumor size (P=0.0061), suggesting that bigger tumors expressed less NR1I3 transcripts, in accordance with our immunohistochemical NR1I3 tests. Our results indicate that NR1I3 expression decreased during progression of malignant lung tumors induced by NNK in mice.

  11. Immunolocalization of NR1, NR2A, and PSD-95 in rat hippocampal subregions during postnatal development.

    Science.gov (United States)

    Ling, Wei; Chang, Lirong; Song, Yizhi; Lu, Tao; Jiang, Yuhua; Li, Youxiang; Wu, Yan

    2012-05-01

    Although the expression of NMDARs and synaptic-associated proteins has been widely studied, the temporospatial distribution of NMDAR subunits and synaptic proteins in different hippocampal subregions during postnatal development still lacks detailed information, and the relationship between NR1 or NR2 subunits and PSD-95 family proteins is controversial. In this study, we used immunofluorescent staining to assess NR1 or NR2A and PSD-95 expressions and the relationship between them in CA1, CA3, and DG of rat hippocampus on postnatal (P) days: P0, P4, P7, P10, P14, P21, P28, P56. The results showed that from P0 to P56, NR1, NR2A, and PSD-95 expressions increased gradually, and the time points of their expression peak differed in CA1, CA3, and DG during postnatal development. Interestingly, although the expression of PSD-95 was positively correlated to both NR1 and NR2A, the NR1 and PSD-95 coexpressed puncta were greatest in CA3, while NR2A and PSD-95 coexpressed puncta were greatest in CA1, compared to other subregions. Surprisingly, at P21, among different strata of CA1, the area of highest expression of NR2A was dramatically changed from stratum pyramidale to stratum polymorphum and stratum moleculare, and returned to stratum pyramidale gradually on the later observed days again, indicating that P21 may be one critical timepoint during postnatal development in CA1. The specific temporospatial distribution pattern of NR1, NR2A, and PSD-95 might be related to the different physiological functions during postnatal development. Discovering the alteration of the relationship between PSD-95 and NMDAR subunits expression may be helpful for understanding mechanisms and therapy of neurodegenerative diseases. Copyright © 2011 Elsevier GmbH. All rights reserved.

  12. 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.

  13. 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.

  14. Self-subunit swapping occurs in another gene type of cobalt nitrile hydratase.

    Directory of Open Access Journals (Sweden)

    Yi Liu

    Full Text Available Self-subunit swapping is one of the post-translational maturation of the cobalt-containing nitrile hydratase (Co-NHase family of enzymes. All of these NHases possess a gene organization of , which allows the activator protein to easily form a mediatory complex with the α-subunit of the NHase after translation. Here, we discovered that the incorporation of cobalt into another type of Co-NHase, with a gene organization of , was also dependent on self-subunit swapping. We successfully isolated a recombinant NHase activator protein (P14K of Pseudomonas putida NRRL-18668 by adding a Strep-tag N-terminal to the P14K gene. P14K was found to form a complex [α(StrepP14K(2] with the α-subunit of the NHase. The incorporation of cobalt into the NHase of P. putida was confirmed to be dependent on the α-subunit substitution between the cobalt-containing α(StrepP14K(2 and the cobalt-free NHase. Cobalt was inserted into cobalt-free α(StrepP14K(2 but not into cobalt-free NHase, suggesting that P14K functions not only as a self-subunit swapping chaperone but also as a metallochaperone. In addition, NHase from P. putida was also expressed by a mutant gene that was designed with a order. Our findings expand the general features of self-subunit swapping maturation.

  15. Self-Subunit Swapping Occurs in Another Gene Type of Cobalt Nitrile Hydratase

    Science.gov (United States)

    Xia, Yuanyuan; Cui, Youtian; Kobayashi, Michihiko; Zhou, Zhemin

    2012-01-01

    Self-subunit swapping is one of the post-translational maturation of the cobalt-containing nitrile hydratase (Co-NHase) family of enzymes. All of these NHases possess a gene organization of , which allows the activator protein to easily form a mediatory complex with the α-subunit of the NHase after translation. Here, we discovered that the incorporation of cobalt into another type of Co-NHase, with a gene organization of , was also dependent on self-subunit swapping. We successfully isolated a recombinant NHase activator protein (P14K) of Pseudomonas putida NRRL-18668 by adding a Strep-tag N-terminal to the P14K gene. P14K was found to form a complex [α(StrepP14K)2] with the α-subunit of the NHase. The incorporation of cobalt into the NHase of P. putida was confirmed to be dependent on the α-subunit substitution between the cobalt-containing α(StrepP14K)2 and the cobalt-free NHase. Cobalt was inserted into cobalt-free α(StrepP14K)2 but not into cobalt-free NHase, suggesting that P14K functions not only as a self-subunit swapping chaperone but also as a metallochaperone. In addition, NHase from P. putida was also expressed by a mutant gene that was designed with a order. Our findings expand the general features of self-subunit swapping maturation. PMID:23226397

  16. A mollusk VDR/PXR/CAR-like (NR1J) nuclear receptor provides insight into ancient detoxification mechanisms

    Energy Technology Data Exchange (ETDEWEB)

    Cruzeiro, Catarina, E-mail: catarinarcruzeiro@hotmail.com [ICBAS - Institute of Biomedical Sciences Abel Salazar, U. Porto - University of Porto (Portugal); CIIMAR/CIMAR - Interdisciplinary Center of Marine and Environmental Research, U. Porto (Portugal); Lopes-Marques, Mónica, E-mail: monicaslm@hotmail.com [ICBAS - Institute of Biomedical Sciences Abel Salazar, U. Porto - University of Porto (Portugal); CIIMAR/CIMAR - Interdisciplinary Center of Marine and Environmental Research, U. Porto (Portugal); Ruivo, Raquel, E-mail: ruivo.raquel@gmail.com [CIIMAR/CIMAR - Interdisciplinary Center of Marine and Environmental Research, U. Porto (Portugal); Rodrigues-Oliveira, Nádia, E-mail: nadia.oliveira@ciimar.up.pt [CIIMAR/CIMAR - Interdisciplinary Center of Marine and Environmental Research, U. Porto (Portugal); Santos, Miguel M., E-mail: santos@ciimar.up.pt [CIIMAR/CIMAR - Interdisciplinary Center of Marine and Environmental Research, U. Porto (Portugal); FCUP - Faculty of Sciences, Department of Biology, U. Porto (Portugal); Rocha, Maria João, E-mail: mjsrocha@netcabo.pt [ICBAS - Institute of Biomedical Sciences Abel Salazar, U. Porto - University of Porto (Portugal); CIIMAR/CIMAR - Interdisciplinary Center of Marine and Environmental Research, U. Porto (Portugal); Rocha, Eduardo, E-mail: erocha@icbas.up.pt [ICBAS - Institute of Biomedical Sciences Abel Salazar, U. Porto - University of Porto (Portugal); CIIMAR/CIMAR - Interdisciplinary Center of Marine and Environmental Research, U. Porto (Portugal); Castro, L. Filipe C., E-mail: filipe.castro@ciimar.up.pt [CIIMAR/CIMAR - Interdisciplinary Center of Marine and Environmental Research, U. Porto (Portugal); FCUP - Faculty of Sciences, Department of Biology, U. Porto (Portugal)

    2016-05-15

    Highlights: • A nuclear receptor orthologue of the NR1J group is isolated from a mollusc. • The molluscan NR1J transactivates gene expression upon exposure to okadaic acid but not a pesticide, esfenvarelate and triclosan. • Lineage specific gene duplications and gene loss have occurred in the NR1J of protostomes with likely impacts on detoxification mechanisms. - Abstract: The origin and diversification of the metazoan endocrine systems represents a fundamental research issue in biology. Nuclear receptors are critical components of these systems. A particular group named VDR/PXR/CAR (NR1I/J) is central in the mediation of detoxification responses. While orthologues have been thoroughly characterized in vertebrates, a sparse representation is currently available for invertebrates. Here, we provide the first isolation and characterization of a lophotrochozoan protostome VDR/PXR/CAR nuclear receptor (NR1J), in the estuarine bivalve the peppery furrow shell (Scrobicularia plana). Using a reporter gene assay, we evaluated the xenobiotic receptor plasticity comparing the human PXR with the S. plana NR1Jβ. Our results show that the molluscan receptor responds to a natural toxin (okadaic acid) in a similar fashion to that reported for other invertebrates. In contrast, the pesticide esfenvalerate displayed a unique response, since it down regulated transactivation at higher concentrations, while for triclosan no response was observed. Additionally, we uncovered lineage specific gene duplications and gene loss in the gene group encoding NRs in protostomes with likely impacts on the complexity of detoxification mechanisms across different phyla. Our findings pave the way for the development of multi-specific sensor tools to screen xenobiotic compounds acting via the NR1I/J group.

  17. >RESTAURANT Nr 1 (building 501 - Meyrin site)

    CERN Multimedia

    2004-01-01

    Customers are kindly requested to note the modified opening times of restaurant nr. 1 and the adjoining newspaper stand from Monday, January 5 to Sunday February 29, 2004: - Kiosque from Monday to Friday 07:30 - 17:00 hrs - Restaurant from Monday to Friday Saturday / Sunday 07:00 - 23:00 hrs08:00 - 21:00 hrs Hot meals will be served between 11:30 and 14:00 hrs, then from 18:00 to 19:30 hrs. Restaurant Supervisory Committee

  18. RESTAURANT Nr 1 (building 501 - Meyrin site)

    CERN Multimedia

    2003-01-01

    OPENING TIMES in JANUARY-FEBRUARY 2004 Customers are kindly requested to note the modified opening times of restaurant nr. 1 and the adjoining newspaper stand from Monday, January 5 to Sunday February 29, 2004: Kiosquefrom Monday to Friday07:30 - 17:00 hrs Restaurant from Monday to FridaySaturday / Sunday 07:00 - 23:00 hrs08:00 - 21:00 hrs Hot meals will be served between 11:30 and 14:00 hrs, then from 18:00 to 19:30 hrs. Restaurant Supervisory Committee

  19. Analysis list: NR1H3 [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available NR1H3 Adipocyte,Blood + hg19 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/target.../NR1H3.1.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/target/NR1H3.5.tsv http://dbarchive.bioscienced...bc.jp/kyushu-u/hg19/target/NR1H3.10.tsv http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/colo/NR1H3.Adipocyte....tsv,http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/colo/NR1H3.Blood.tsv http://dbarchive.bioscience...dbc.jp/kyushu-u/hg19/colo/Adipocyte.gml,http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/colo/Blood.gml ...

  20. Nuclear receptor Rev-erb alpha (Nr1d1 functions in concert with Nr2e3 to regulate transcriptional networks in the retina.

    Directory of Open Access Journals (Sweden)

    Nissa J Mollema

    Full Text Available The majority of diseases in the retina are caused by genetic mutations affecting the development and function of photoreceptor cells. The transcriptional networks directing these processes are regulated by genes such as nuclear hormone receptors. The nuclear hormone receptor gene Rev-erb alpha/Nr1d1 has been widely studied for its role in the circadian cycle and cell metabolism, however its role in the retina is unknown. In order to understand the role of Rev-erb alpha/Nr1d1 in the retina, we evaluated the effects of loss of Nr1d1 to the developing retina and its co-regulation with the photoreceptor-specific nuclear receptor gene Nr2e3 in the developing and mature retina. Knock-down of Nr1d1 expression in the developing retina results in pan-retinal spotting and reduced retinal function by electroretinogram. Our studies show that NR1D1 protein is co-expressed with NR2E3 in the outer neuroblastic layer of the developing mouse retina. In the adult retina, NR1D1 is expressed in the ganglion cell layer and is co-expressed with NR2E3 in the outer nuclear layer, within rods and cones. Several genes co-targeted by NR2E3 and NR1D1 were identified that include: Nr2c1, Recoverin, Rgr, Rarres2, Pde8a, and Nupr1. We examined the cyclic expression of Nr1d1 and Nr2e3 over a twenty-four hour period and observed that both nuclear receptors cycle in a similar manner. Taken together, these studies reveal a novel role for Nr1d1, in conjunction with its cofactor Nr2e3, in regulating transcriptional networks critical for photoreceptor development and function.

  1. Nuclear receptor Rev-erb alpha (Nr1d1) functions in concert with Nr2e3 to regulate transcriptional networks in the retina.

    Science.gov (United States)

    Mollema, Nissa J; Yuan, Yang; Jelcick, Austin S; Sachs, Andrew J; von Alpen, Désirée; Schorderet, Daniel; Escher, Pascal; Haider, Neena B

    2011-03-08

    The majority of diseases in the retina are caused by genetic mutations affecting the development and function of photoreceptor cells. The transcriptional networks directing these processes are regulated by genes such as nuclear hormone receptors. The nuclear hormone receptor gene Rev-erb alpha/Nr1d1 has been widely studied for its role in the circadian cycle and cell metabolism, however its role in the retina is unknown. In order to understand the role of Rev-erb alpha/Nr1d1 in the retina, we evaluated the effects of loss of Nr1d1 to the developing retina and its co-regulation with the photoreceptor-specific nuclear receptor gene Nr2e3 in the developing and mature retina. Knock-down of Nr1d1 expression in the developing retina results in pan-retinal spotting and reduced retinal function by electroretinogram. Our studies show that NR1D1 protein is co-expressed with NR2E3 in the outer neuroblastic layer of the developing mouse retina. In the adult retina, NR1D1 is expressed in the ganglion cell layer and is co-expressed with NR2E3 in the outer nuclear layer, within rods and cones. Several genes co-targeted by NR2E3 and NR1D1 were identified that include: Nr2c1, Recoverin, Rgr, Rarres2, Pde8a, and Nupr1. We examined the cyclic expression of Nr1d1 and Nr2e3 over a twenty-four hour period and observed that both nuclear receptors cycle in a similar manner. Taken together, these studies reveal a novel role for Nr1d1, in conjunction with its cofactor Nr2e3, in regulating transcriptional networks critical for photoreceptor development and function.

  2. 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.

  3. 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...

  4. 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

  5. Molecular cloning and expression of heteromeric ACCase subunit genes from Jatropha curcas.

    Science.gov (United States)

    Gu, Keyu; Chiam, Huihui; Tian, Dongsheng; Yin, Zhongchao

    2011-04-01

    Acetyl-CoA carboxylase (ACCase) catalyzes the biotin-dependent carboxylation of acetyl-CoA to produce malonyl-CoA, which is the essential first step in the biosynthesis of long-chain fatty acids. ACCase exists as a multi-subunit enzyme in most prokaryotes and the chloroplasts of most plants and algae, while it is present as a multi-domain enzyme in the endoplasmic reticulum of most eukaryotes. The heteromeric ACCase of higher plants consists of four subunits: an α-subunit of carboxyltransferase (α-CT, encoded by accA gene), a biotin carboxyl carrier protein (BCCP, encoded by accB gene), a biotin carboxylase (BC, encoded by accC gene) and a β-subunit of carboxyltransferase (β-CT, encoded by accD gene). In this study, we cloned and characterized the genes accA, accB1, accC and accD that encode the subunits of heteromeric ACCase in Jatropha (Jatropha curcas), a potential biofuel plant. The full-length cDNAs of the four subunit genes were isolated from a Jatropha cDNA library and by using 5' RACE, whereas the genomic clones were obtained from a Jatropha BAC library. They encode a 771 amino acid (aa) α-CT, a 286-aa BCCP1, a 537-aa BC and a 494-aa β-CT, respectively. The single-copy accA, accB1 and accC genes are nuclear genes, while the accD gene is located in chloroplast genome. Jatropha α-CT, BCCP1, BC and β-CT show high identity to their homologues in other higher plants at amino acid level and contain all conserved domains for ACCase activity. The accA, accB1, accC and accD genes are temporally and spatially expressed in the leaves and endosperm of Jatropha plants, which are regulated by plant development and environmental factors. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  6. 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.

  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. 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.

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

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    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.

  13. Caffeine increases Nr1i3 expression and potentiates the effects of its ligand, TCPOBOP, in mice liver

    Directory of Open Access Journals (Sweden)

    Heidge Fukumasu

    2015-06-01

    Full Text Available Caffeine is one of the world's most consumed substances. It is present in coffee, green tea and guarana, among others. The xenobiotic-sensing nuclear receptor subfamily 1, group I, member 3 (Nr1i3, also known as the Constitutive Androstane Receptor (Car is a key regulator of drug metabolism and excretion. No consistent description of caffeine effects on this receptor has been described. Thus, to unravel the effects of caffeine on this receptor, we performed experiments in mice. First, C57Bl/6 mice that were treated daily with caffeine (50 mg/kg for 15 days presented a slight but significant increase in Nr1i3 and Cyp2b10 gene expression. A second experiment was then performed to verify the effects of caffeine on TCPOBOP (1,4-bis-[2-(3,5-dichloropyridyloxy]benzene, 3,3′,5,5′-tetrachloro-1,4-bis(pyridyloxybenzene, the most potent agonist known for mice Nr1i3. Interestingly, caffeine potentiated TCPOBOP pleiotropic effects in mice liver, such as hepatomegaly, hepatotoxicity, hepatocyte proliferation and loss of cell-to-cell communication through gap junctions. In addition, caffeine plus TCPOBOP treatment increased liver gene expression of Nr1i3 and Cyp2b10 comparing with only caffeine or TCPOBOP treatments. Together, these results indicate that caffeine increases the expression of Nr1i3 in mice liver, although at this point it is not possible to determine if Nr1i3 directly or indirectly mediates this effect.

  14. 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

  15. A mollusk VDR/PXR/CAR-like (NR1J) nuclear receptor provides insight into ancient detoxification mechanisms.

    Science.gov (United States)

    Cruzeiro, Catarina; Lopes-Marques, Mónica; Ruivo, Raquel; Rodrigues-Oliveira, Nádia; Santos, Miguel M; Rocha, Maria João; Rocha, Eduardo; Castro, L Filipe C

    2016-05-01

    The origin and diversification of the metazoan endocrine systems represents a fundamental research issue in biology. Nuclear receptors are critical components of these systems. A particular group named VDR/PXR/CAR (NR1I/J) is central in the mediation of detoxification responses. While orthologues have been thoroughly characterized in vertebrates, a sparse representation is currently available for invertebrates. Here, we provide the first isolation and characterization of a lophotrochozoan protostome VDR/PXR/CAR nuclear receptor (NR1J), in the estuarine bivalve the peppery furrow shell (Scrobicularia plana). Using a reporter gene assay, we evaluated the xenobiotic receptor plasticity comparing the human PXR with the S. plana NR1Jβ. Our results show that the molluscan receptor responds to a natural toxin (okadaic acid) in a similar fashion to that reported for other invertebrates. In contrast, the pesticide esfenvalerate displayed a unique response, since it down regulated transactivation at higher concentrations, while for triclosan no response was observed. Additionally, we uncovered lineage specific gene duplications and gene loss in the gene group encoding NRs in protostomes with likely impacts on the complexity of detoxification mechanisms across different phyla. Our findings pave the way for the development of multi-specific sensor tools to screen xenobiotic compounds acting via the NR1I/J group.

  16. GABA{sub A} receptor beta 3 subunit gene is possibly paternally imprinted in humans

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1994-02-15

    As the gene for GABA{sub A} receptor beta 3 subunit (GABRB3) is encompassed by a small molecular deletion in chromosome 15q11-q13, which is the critical region for Angelman syndrome(AS), the GABRB3 gene could be a candidate gene for AS. The abnormal phenotype of AS is manifested only when the deletion is inherited from the mother, not from the father. Therefore, a candidate gene for AS should be paternally imprinted. Although it was reported that the GABRB3 gene was expressed equally from either the maternal or paternal chromosome in mouse brain (i.e., not imprinted), it is well known that imprinting shows tissue specificity, and it remains to be determined if all genes imprinted in the mouse are also imprinted in humans. 4 refs., 1 fig.

  17. 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.

  18. 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

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

    Directory of Open Access Journals (Sweden)

    Ben-Wen Li

    2015-12-01

    Full Text Available 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 Vas 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

  20. 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.

  1. 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.

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

    Science.gov (United States)

    Zhao, Yan; Gentekaki, Eleni; Yi, Zhenzhen; Lin, Xiaofeng

    2013-01-01

    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. 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. Our results suggest that the COI gene is a suitable marker for resolving inter- and intra-specific relationships of Paramecium spp.

  3. 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...

  4. 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...

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

    Directory of Open Access Journals (Sweden)

    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.

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

    Directory of Open Access Journals (Sweden)

    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.

  7. Evolutionary Analysis of the B56 Gene Family of PP2A Regulatory Subunits

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    Lauren M. Sommer

    2015-05-01

    Full Text Available Protein phosphatase 2A (PP2A is an abundant serine/threonine phosphatase that functions as a tumor suppressor in numerous cell-cell signaling pathways, including Wnt, myc, and ras. The B56 subunit of PP2A regulates its activity, and is encoded by five genes in humans. B56 proteins share a central core domain, but have divergent amino- and carboxy-termini, which are thought to provide isoform specificity. We performed phylogenetic analyses to better understand the evolution of the B56 gene family. We found that B56 was present as a single gene in eukaryotes prior to the divergence of animals, fungi, protists, and plants, and that B56 gene duplication prior to the divergence of protostomes and deuterostomes led to the origin of two B56 subfamilies, B56αβε and B56γδ. Further duplications led to three B56αβε genes and two B56γδ in vertebrates. Several nonvertebrate B56 gene names are based on distinct vertebrate isoform names, and would best be renamed. B56 subfamily genes lack significant divergence within primitive chordates, but each became distinct in complex vertebrates. Two vertebrate lineages have undergone B56 gene loss, Xenopus and Aves. In Xenopus, B56δ function may be compensated for by an alternatively spliced transcript, B56δ/γ, encoding a B56δ-like amino-terminal region and a B56γ core.

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

    Directory of Open Access Journals (Sweden)

    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.

  9. 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.

  10. Exercise induced upregulation of glutamate-cysteine ligase catalytic subunit and glutamate-cysteine ligase modifier subunit gene expression in Thoroughbred horses

    Directory of Open Access Journals (Sweden)

    Jeong-Woong Park

    2017-05-01

    Full Text Available Objective This study was performed to reveal the molecular structure and expression patterns of horse glutamate-cysteine ligase catalytic subunit (GCLC and glutamate-cysteine ligase modifier subunit (GCLM genes whose products form glutamate cysteine ligase, which were identified as differentially expressed genes in the previous study. Methods We performed bioinformatics analyses, and gene expression assay with quantitative polymerase chain reaction (qPCR for horse GCLC and GCLM genes in muscle and blood leukocytes of Thoroughbred horses Results Expression of GCLC showed the same pattern in both blood and muscle tissues after exercise. Expression of GCLC increased in the muscle and blood of Thoroughbreds, suggesting a tissue-specific regulatory mechanism for the expression of GCLC. In addition, expression of the GCLM gene increased after exercise in both the blood and muscle of Thoroughbreds. Conclusion We established the expression patterns of GCLC and GCLM in the skeletal muscle and blood of Thoroughbred horses in response to exercise. Further study is now warranted to uncover the functional importance of these genes in exercise and recovery in racehorses.

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

    Directory of Open Access Journals (Sweden)

    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.

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

    Science.gov (United States)

    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.

  13. 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.

  14. Characterization of a Low-Molecular-Weight Glutenin Subunit Gene from Bread Wheat and the Corresponding Protein That Represents a Major Subunit of the Glutenin Polymer1

    Science.gov (United States)

    Masci, Stefania; D'Ovidio, Renato; Lafiandra, Domenico; Kasarda, Donald D.

    1998-01-01

    Both high- and low-molecular-weight glutenin subunits (LMW-GS) play the major role in determining the viscoelastic properties of wheat (Triticum aestivum L.) flour. To date there has been no clear correspondence between the amino acid sequences of LMW-GS derived from DNA sequencing and those of actual LMW-GS present in the endosperm. We have characterized a particular LMW-GS from hexaploid bread wheat, a major component of the glutenin polymer, which we call the 42K LMW-GS, and have isolated and sequenced the putative corresponding gene. Extensive amino acid sequences obtained directly for this 42K LMW-GS indicate correspondence between this protein and the putative corresponding gene. This subunit did not show a cysteine (Cys) at position 5, in contrast to what has frequently been reported for nucleotide-based sequences of LMW-GS. This Cys has been replaced by one occurring in the repeated-sequence domain, leaving the total number of Cys residues in the molecule the same as in various other LMW-GS. On the basis of the deduced amino acid sequence and literature-based assignment of disulfide linkages, a computer-generated molecular model of the 42K subunit was constructed. PMID:9847089

  15. Gene Regulation, Alternative Splicing, and Posttranslational Modification of Troponin Subunits in Cardiac Development and Adaptation: A Focused Review

    Directory of Open Access Journals (Sweden)

    Juan-Juan eSheng

    2014-04-01

    Full Text Available 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.

  16. 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.

  17. 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.

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

    Directory of Open Access Journals (Sweden)

    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

  19. 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

  20. 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

  1. 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.

  2. 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 biosynthesis of carbamoylphosphate is catalysed by the heterodimeric enzyme carbamoylphosphate synthetase (CPSase). The genes encoding the two subunits in procaryotes are normally transcribed as an operon, whereas in Lactococcus lactis, the gene encoding the large subunit (carB) is shown...... 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...... 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...

  3. 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.

  4. 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.

  5. Small-subunit ribosomal RNA gene sequences of Phaeodarea challenge the monophyly of Haeckel's Radiolaria.

    Science.gov (United States)

    Polet, Stephane; Berney, Cédric; Fahrni, José; Pawlowski, Jan

    2004-03-01

    In his grand monograph of Radiolaria, Ernst Haeckel originally included Phaeodarea together with Acantharea and Polycystinea, all three taxa characterized by the presence of a central capsule and the possession of axopodia. Cytological and ultrastructural studies, however, questioned the monophyly of Radiolaria, suggesting an independent evolutionary origin of the three taxa, and the first molecular data on Acantharea and Polycystinea brought controversial results. To test further the monophyly of Radiolaria, we sequenced the complete small subunit ribosomal RNA gene of three phaeodarians and three polycystines. Our analyses reveal that phaeodarians clearly branch among the recently described phylum Cercozoa, separately from Acantharea and Polycystinea. This result enhances the morphological variability within the phylum Cercozoa, which already contains very heterogeneous groups of protists. Our study also confirms the common origin of Acantharea and Polycystinea, which form a sister-group to the Cercozoa, and allows a phylogenetic reinterpretation of the morphological features of the three radiolarian groups.

  6. Promoter Structure of the RNA Polymerase II Large Subunit Gene in Caenorhabditis elegans and C. briggsae.

    Science.gov (United States)

    Bird, D M; Kaloshian, I; Molinari, S

    1997-06-01

    The 5'-end of the Caenorhabditis elegans ama-1 gene transcript, which encodes the largest subunit of RNA polymerase II, was cloned. Sequencing revealed that the message is trans-spliced. To characterize the Ce-ama-1 promoter, DNA sequence spanning 3 kb upstream from the initiation codon was determined. Typical elements, such as TATA and Spl sites, were absent. The homologue of ama-1 in C. briggsae, Cb-ama-1, was isolated and its 5' flanking sequence compared with that of Ce-ama-1, revealing only limited similarity, although both sequences included a potential initiator-class transcriptional regulator and phased repeats of an ATC motif. The latter elements are postulated to facilitate DNA bending and may play a role in transcription regulation.

  7. Utility of the cytochrome c oxidase subunit I gene for the diagnosis of toxoplasmosis using PCR.

    Science.gov (United States)

    Feng, Xue; Norose, Kazumi; Li, Kexin; Hikosaka, Kenji

    2017-10-01

    Toxoplasmosis is caused by the protozoan parasite Toxoplasma gondii, which belongs to the phylum Apicomplexa. Since this parasite causes severe clinical symptoms in immunocompromised patients, early diagnosis of toxoplasmosis is essential. PCR is currently used for early diagnosis, but there is no consensus regarding the most effective method for amplifying Toxoplasma DNA. In this study, we considered the utility of the cytochrome c subunit I (cox1) gene, which is encoded in the mitochondrial DNA of this parasite, as a novel target of PCR for the diagnosis of toxoplasmosis. To do this, we compared its copy number per haploid nuclear genome and the detection sensitivity of cox1-PCR with the previously reported target genes B1 and 18S rRNA and the AF146527 repeat element. We found that the copy number of cox1 was high and that the PCR using cox1 primers was more efficient at amplifying Toxoplasma DNA than the other PCR targets examined. In addition, PCR using clinical samples indicated that the cox1 gene would be useful for the diagnosis of toxoplasmosis. These findings suggest that use of cox1-PCR would facilitate the diagnosis of toxoplasmosis in clinical laboratories. Copyright © 2017 Elsevier B.V. All rights reserved.

  8. 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.

  9. 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.

  10. Targeting Activation of Specific NF-κB Subunits Prevents Stress-Dependent Atherothrombotic Gene Expression

    Science.gov (United States)

    Djuric, Zdenka; Kashif, Muhammed; Fleming, Thomas; Muhammad, Sajjad; Piel, David; von Bauer, Rüdiger; Bea, Florian; Herzig, Stephan; Zeier, Martin; Pizzi, Marina; Isermann, Berend; Hecker, Markus; Schwaninger, Markus; Bierhaus, Angelika; Nawroth, Peter P

    2012-01-01

    Psychosocial stress has been shown to be a contributing factor in the development of atherosclerosis. Although the underlying mechanisms have not been elucidated entirely, it has been shown previously that the transcription factor nuclear factor-κB (NF-κB) is an important component of stress-activated signaling pathway. In this study, we aimed to decipher the mechanisms of stress-induced NF-κB-mediated gene expression, using an in vitro and in vivo model of psychosocial stress. Induction of stress led to NF-κB-dependent expression of proinflammatory (tissue factor, intracellular adhesive molecule 1 [ICAM-1]) and protective genes (manganese superoxide dismutase [MnSOD]) via p50, p65 or cRel. Selective inhibition of the different subunits and the respective kinases showed that inhibition of cRel leads to the reduction of atherosclerotic lesions in apolipoprotein−/− (ApoE−/−) mice via suppression of proinflammatory gene expression. This observation may therefore provide a possible explanation for ineffectiveness of antioxidant therapies and suggests that selective targeting of cRel activation may provide a novel approach for the treatment of stress-related inflammatory vascular disease. PMID:23114885

  11. 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.

  12. 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.

  13. 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.

  14. 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

  15. 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

  16. 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...

  17. 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.

  18. 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.

  19. 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

  20. 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

  1. 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

  2. Influence of a threonine residue in the S2 ligand binding domain in determining agonist potency and deactivation rate of recombinant NR1a/NR2D NMDA receptors.

    Science.gov (United States)

    Chen, Philip E; Johnston, Alexander R; Mok, M H Selina; Schoepfer, Ralf; Wyllie, David J A

    2004-07-01

    NR1/NR2D NMDA receptors display unusually slow deactivation kinetics which may be critical for their role as extrasynaptic receptors. A threonine to alanine point mutation has been inserted at amino acid position 692 of the NR2D subunit (T692A). Recombinant NR1a/NR2D(T692A) NMDA receptors have been expressed in Xenopus laevis oocytes and their pharmacological and single-channel properties examined using two-electrode voltage-clamp and patch-clamp recording techniques. Glutamate dose-response curves from NR1a/NR2D(T692A) receptor channels produced an approximately 1600-fold reduction in glutamate potency compared to wild-type NR1a/NR2D receptors. There was no change in Hill slopes or gross reduction in mean maximal currents recorded in oocytes expressing either wild-type or mutant receptors. The mutation did not affect the potency of the co-agonist glycine. The shifts in potency produced by NR2D(T692A) containing receptors when activated by other glutamate-site agonists such as aspartate or NMDA were 30- to 60-fold compared to wild-type. Single-channel conductance levels of NR1a/NR2D(T692A) mutant receptors were indistinguishable from wild-type NR2D-containing channels. Additionally NR1a/NR2D(T692A) receptors showed the transitional asymmetry that is characteristic of NR2D-containing NMDA receptors. Rapid applications of glutamate on outside-out patches containing NR1a/NR2D(T692A) receptors produced macroscopic current deactivations that were about 60-fold faster than wild-type NR1a/NR2D receptors. Our results suggest that this conserved threonine residue plays a crucial role in ligand binding to NMDA NR2 receptor subunits and supports the idea that the slow decay kinetics associated with NR1a/NR2D NMDA receptors can be explained by the slow dissociation of glutamate from this NMDA receptor subtype.

  3. 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.

  4. 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.

  5. 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.

  6. 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).

  7. 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.

  8. 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.

  9. 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

  10. 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.

  11. 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.

  12. 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

  13. 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.

  14. The epithelial sodium channel γ-subunit gene and blood pressure: family based association, renal gene expression, and physiological analyses.

    Science.gov (United States)

    Büsst, Cara J; Bloomer, Lisa D S; Scurrah, Katrina J; Ellis, Justine A; Barnes, Timothy A; Charchar, Fadi J; Braund, Peter; Hopkins, Paul N; Samani, Nilesh J; Hunt, Steven C; Tomaszewski, Maciej; Harrap, Stephen B

    2011-12-01

    Variants in the gene encoding the γ-subunit of the epithelial sodium channel (SCNN1G) are associated with both Mendelian and quantitative effects on blood pressure. Here, in 4 cohorts of 1611 white European families composed of a total of 8199 individuals, we undertook staged testing of candidate single-nucleotide polymorphisms for SCNN1G (supplemented with imputation based on data from the 1000 Genomes Project) followed by a meta-analysis in all of the families of the strongest candidate. We also examined relationships between the genotypes and relevant intermediate renal phenotypes, as well as expression of SCNN1G in human kidneys. We found that an intronic single-nucleotide polymorphism of SCNN1G (rs13331086) was significantly associated with age-, sex-, and body mass index-adjusted blood pressure in each of the 4 populations (Ppressure and 0.52-mm Hg increase in diastolic blood pressure (SE=0.33, P=0.002 for systolic blood pressure; SE=0.21, P=0.011 for diastolic blood pressure). The same allele was also associated with higher 12-hour overnight urinary potassium excretion (P=0.04), consistent with increased epithelial sodium channel activity. Renal samples from hypertensive subjects showed a nonsignificant (P=0.07) 1.7-fold higher expression of SCNN1G compared with normotensive controls. These data provide genetic and phenotypic evidence in support of a role for a common genetic variant of SCNN1G in blood pressure determination.

  15. 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.

  16. Genetic interaction of an origin recognition complex subunit and the Polycomb group gene MEDEA during seed development.

    Science.gov (United States)

    Collinge, Margaret A; Spillane, Charles; Köhler, Claudia; Gheyselinck, Jacqueline; Grossniklaus, Ueli

    2004-04-01

    The eukaryotic origin recognition complex (ORC) is made up of six subunits and functions in nuclear DNA replication, chromatin structure, and gene silencing in both fungi and metazoans. We demonstrate that disruption of a plant ORC subunit homolog, AtORC2 of Arabidopsis (Arabidopsis thaliana), causes a zygotic lethal mutant phenotype (orc2). Seeds of orc2 abort early, typically producing embryos with up to eight cells. Nuclear division in the endosperm is arrested at an earlier developmental stage: only approximately four nuclei are detected in orc2 endosperm. The endosperm nuclei in orc2 are dramatically enlarged, a phenotype that is most similar to class B titan mutants, which include mutants in structural maintenance of chromosomes (SMC) cohesins. The highest levels of ORC2 gene expression were found in preglobular embryos, coinciding with the stage at which homozygous orc2 mutant seeds arrest. The homologs of the other five Arabidopsis ORC subunits are also expressed at this developmental stage. The orc2 mutant phenotype is partly suppressed by a mutation in the Polycomb group gene MEDEA. In double mutants between orc2 and medea (mea), orc2 homozygotes arrest later with a phenotype intermediate between those of mea and orc2 single mutants. Either alterations in chromatin structure or the release of cell cycle checkpoints by the mea mutation may allow more cell and nuclear divisions to occur in orc2 homozygous seeds.

  17. The carB Gene Encoding the Large Subunit of Carbamoylphosphate Synthetase from Lactococcus lactis Is Transcribed Monocistronically

    Science.gov (United States)

    Martinussen, Jan; Hammer, Karin

    1998-01-01

    The biosynthesis of carbamoylphosphate is catalyzed by the heterodimeric enzyme carbamoylphosphate synthetase. The genes encoding the two subunits of this enzyme in procaryotes are normally transcribed as an operon, but the gene encoding the large subunit (carB) in Lactococcus lactis is shown to be transcribed as an isolated 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 the requirement for carbamoylphosphate in pyrimidine biosynthesis through degradation by means of 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 showing a high degree of similarity to those of glutathione peroxidases from other organisms was identified. PMID:9721272

  18. Prodynorphin gene deletion increased anxiety-like behaviours, impaired the anxiolytic effect of bromazepam and altered GABAA receptor subunits gene expression in the amygdala.

    Science.gov (United States)

    Femenía, Teresa; Pérez-Rial, Sandra; Urigüen, Leyre; Manzanares, Jorge

    2011-01-01

    This study evaluated the role of prodynorphin gene in the regulation of anxiety and associated molecular mechanisms. Emotional responses were assessed using the light-dark test, elevated plus maze and social interaction tests in prodynorphin knockout and wild-type mice. Corticotrophin releasing factor and proopiomelanocortin gene expressions in the hypothalamus were evaluated after restraint stress using in situ hybridization. The anxiolytic efficacy of bromazepam and GABA(A) receptor subunits gene expression in the amygdala were also assessed in both genotypes. The deletion of prodynorphin increased anxiety-like behaviours and proopiomelanocortin gene expression in the arcuate nucleus (two-fold). Moreover, the anxiolytic action of bromazepam was significantly attenuated in the mutant mice. Decreased GABA(A)γ(2) and increased GABA(A)β(2) gene expression receptor subunits were found in the amygdala of prodynorphin knockout mice. These results indicate that deletion of prodynorphin gene is associated with increased anxiety-like behaviours, enhanced sensibility response to stress stimuli, reduced anxiolytic efficacy of bromazepam and altered expression of the GABA(A) receptor subunits.

  19. 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.

  20. 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.

  1. 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.

  2. 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.

  3. 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.

  4. File list: Oth.ALL.10.Nr1d2.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  5. File list: Oth.ALL.05.Nr1d1.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  6. File list: Oth.ALL.20.Nr1d1.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  7. File list: Oth.ALL.50.Nr1d2.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  8. File list: Oth.ALL.20.Nr1d2.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  9. 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

  10. 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.

  11. Bilaterian phylogeny based on analyses of a region of the sodium-potassium ATPase beta-subunit gene.

    Science.gov (United States)

    Anderson, Frank E; Córdoba, Alonso J; Thollesson, Mikael

    2004-03-01

    Molecular investigations of deep-level relationships within and among the animal phyla have been hampered by a lack of slowly evolving genes that are amenable to study by molecular systematists. To provide new data for use in deep-level metazoan phylogenetic studies, primers were developed to amplify a 1.3-kb region of the alpha subunit of the nuclear-encoded sodium-potassium ATPase gene from 31 bilaterians representing several phyla. Maximum parsimony, maximum likelihood, and Bayesian analyses of these sequences (combined with ATPase sequences for 23 taxa downloaded from GenBank) yield congruent trees that corroborate recent findings based on analyses of other data sets (e.g., the 18S ribosomal RNA gene). The ATPase-based trees support monophyly for several clades (including Lophotrochozoa, a form of Ecdysozoa, Vertebrata, Mollusca, Bivalvia, Gastropoda, Arachnida, Hexapoda, Coleoptera, and Diptera) but do not support monophyly for Deuterostomia, Arthropoda, or Nemertea. Parametric bootstrapping tests reject monophyly for Arthropoda and Nemertea but are unable to reject deuterostome monophyly. Overall, the sodium-potassium ATPase alpha-subunit gene appears to be useful for deep-level studies of metazoan phylogeny.

  12. 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

  13. Genetic mapping of the LMP2 proteasome subunit gene to the BoLA class IIb region

    Energy Technology Data Exchange (ETDEWEB)

    Shalhevet, D.; Da, Y.; Beever, J.E.; Eijk, M.J.T. van; Ma, R.; Lewin, H.A.; Gaskins, H.R. [Univ. of Illinois, Champaign, IL (United States)

    1995-01-01

    Recent identification of four tightly-linked genes within the class II region of the major histocompatibility complex (MHC) in humans and rodents has led to a better understanding of class I antigen processing mechanisms. Two of these genes, LMP2 and LMP7, encode subunits of a low molecular mass poypeptide (LMP) complex. Several observations suggest that the LMP complex may be the proteolytic system responsible for generating the size-restricted peptides required for MHC class I assembly. For example, the LMP complex is a large cytoplasmic structure that is antigenically and biochemically related to the proteasome, a proteolytic complex that mediates degradation of ubiquitinated substrates. Data regarding proteolytic specificity indicates that the LMP complex may specifically produce nonamers, the appropriate peptide size for class I binding. In addition, similar to all components of the class I assembly process, intra-MHC LMP genes are regulated by IFN{gamma}. 26 refs., 2 figs., 1 tab.

  14. Mitochondrial genome from the facultative anaerobe and petite-positive yeast Dekkera bruxellensis contains the NADH dehydrogenase subunit genes.

    Science.gov (United States)

    Procházka, Emanuel; Poláková, Silvia; Piskur, Jure; Sulo, Pavol

    2010-08-01

    The progenitor of the Dekkera/Brettanomyces clade separated from the Saccharomyces/Kluyveromyces clade over 200 million years ago. However, within both clades, several lineages developed similar physiological traits. Both Saccharomyces cerevisiae and Dekkera bruxellensis are facultative anaerobes; in the presence of excess oxygen and sugars, they accumulate ethanol (Crabtree effect) and they both spontaneously generate respiratory-deficient mutants (petites). In order to understand the role of respiratory metabolism, the mitochondrial DNA (mtDNA) molecules of two Dekkera/Brettanomyces species were analysed. Dekkera bruxellensis mtDNA shares several properties with S. cerevisiae, such as the large genome size (76 453 bp), and the organization of the intergenic sequences consisting of spacious AT-rich regions containing a number of hairpin GC-rich cluster-like elements. In addition to a basic set of the mitochondrial genes coding for the components of cytochrome oxidase, cytochrome b, subunits of ATPase, two rRNA subunits and 25 tRNAs, D. bruxellensis also carries genes for the NADH dehydrogenase complex. Apparently, in yeast, the loss of this complex is not a precondition to develop a petite-positive, Crabtree-positive and anaerobic nature. On the other hand, mtDNA from a petite-negative Brettanomyces custersianus is much smaller (30 058 bp); it contains a similar gene set and has only short intergenic sequences.

  15. 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.

  16. 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.

  17. 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

  18. 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.

  19. Hyperkalemic periodic paralysis caused by recurring mutation in the adult muscle sodium channel alpha-subunit gene.

    Science.gov (United States)

    Sillén, A; Wadelius, C; Sundvall, M; Ahlsten, G; Gustavson, K H

    1996-01-01

    Linkage studies and mutation analysis were performed in two Swedish families with hyperkalemic periodic paralysis (HYPP), an autosomal dominant inherited disorder characterized by episodic muscle weakness associated with increasing or high levels of serum potassium. The gene for HYPP is the gene encoding the alpha-subunit of the sodium channel of adult human skeletal muscle (SCN4A). SCN4A has been localized on chromosome 17 q closely linked to the human growth hormone gene. Linkage between a microsatellite polymorphism in the SCN4A gene and the disease was shown in two Swedish families (Z = 12.10 theta = 0). Sequence analysis revealed that the two Swedish families have got a C to T transition at position 2188 in the cDNA. At the protein level this Thr 704 to Met mutation is located in the fifth membrane spanning segment of domain II of the protein, as previously described (28). The mutation was linked to different microsatellite alleles regarding both a (GT)n and a (GA)n repeat in the gene. Either the families are related and new mutations have occurred in both microsatellites when the pedigrees were separated or the mutation has arisen independently in the two families analysed. From the mutant alleles characterized so far it seems as if a limited number of mutations is present in this gene.

  20. 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.

  1. 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....

  2. 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

  3. 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

  4. 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.

  5. 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.

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

  7. 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.

  8. 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.

  9. 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.

  10. 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.

  11. Modeling RNA polymerase competition: the effect of σ-subunit knockout and heat shock on gene transcription level

    Directory of Open Access Journals (Sweden)

    Seliverstov Alexandr V

    2011-01-01

    type of σ-subunit, the amount of transcription initiation aborts, etc. The model can be used to make functional predictions, e.g., heat shock response in isolated chloroplasts and changes of gene transcription levels under knockout of different σ-subunits or RNA polymerases or due to gene expression regulation. Reviewers This article was reviewed by Dr. Anthony Almudevar, Dr. Aniko Szabo, Dr. Yuri Wolf (nominated by Dr. Peter Olofsson and Prof. Marek Kimmel.

  12. 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.

  13. 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.

  14. File list: Oth.Liv.05.Nr1d1.AllCell [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Oth.Liv.05.Nr1d1.AllCell mm9 TFs and others Nr1d1 Liver SRX128176,SRX997762,SRX9977...63,SRX997761,SRX997760,SRX997765,SRX997764,SRX109461,SRX100296,SRX994725,SRX994726,SRX100297 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Oth.Liv.05.Nr1d1.AllCell.bed ...

  15. 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.

  16. IDENTIFICATION OF POLYMORPHISM OF FSH BETA-SUBUNIT GENE AS SPERM QUALITY MARKER IN BALI CATTLE USING PCR-RFLP

    Directory of Open Access Journals (Sweden)

    A.B.L. Ishak

    2014-10-01

    Full Text Available The aim of study was to identify the association of FSH beta-subunit gene polymorphisms withsperm quality traits. A total of 470 samples of normal mature bull from several breeds were used forpopulation study and 127 bulls from National and Regional AI centre of Indonesia for association study.To amplify, a PCR-RFLP method was used and digested with Pst1 restriction enzyme. The allelefrequency of the A and B in Bali cattle were (0.000 and (1.000, respectively. The absence of otherallele A suggested that the Bali cattle was monomorphic, while Brahman, FH, Simmental and Limousinewere polymorphic. The highest observed heterozygosity were found in Limousine (0.318 and thehighest expected heterozygosity were in Simmental (0.420. The higher incident of percentage of spermabnormalities were found in Simmental, Limousin, Brahman compared to Bali and FH. Among all typesof sperm abnormalities, the abaxial and microcephalus were found in highest number.

  17. 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.

  18. 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.

  19. 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.

  20. 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

  1. 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.

  2. 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...

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

    Directory of Open Access Journals (Sweden)

    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

  4. Glycine activated ion channel subunits encoded by ctenophore glutamate receptor genes.

    Science.gov (United States)

    Alberstein, Robert; Grey, Richard; Zimmet, Austin; Simmons, David K; Mayer, Mark L

    2015-11-03

    Recent genome projects for ctenophores have revealed the presence of numerous ionotropic glutamate receptors (iGluRs) in Mnemiopsis leidyi and Pleurobrachia bachei, among our earliest metazoan ancestors. Sequence alignments and phylogenetic analysis show that these form a distinct clade from the well-characterized AMPA, kainate, and NMDA iGluR subtypes found in vertebrates. Although annotated as glutamate and kainate receptors, crystal structures of the ML032222a and PbiGluR3 ligand-binding domains (LBDs) reveal endogenous glycine in the binding pocket, whereas ligand-binding assays show that glycine binds with nanomolar affinity; biochemical assays and structural analysis establish that glutamate is occluded from the binding cavity. Further analysis reveals ctenophore-specific features, such as an interdomain Arg-Glu salt bridge, present only in subunits that bind glycine, but also a conserved disulfide in loop 1 of the LBD that is found in all vertebrate NMDA but not AMPA or kainate receptors. We hypothesize that ctenophore iGluRs are related to an early ancestor of NMDA receptors, suggesting a common evolutionary path for ctenophores and bilaterian species, and suggest that future work should consider both glycine and glutamate as candidate neurotransmitters in ctenophore species.

  5. A linkage study between the GABAA beta2 and GABAA gamma2 subunit genes and major psychoses.

    Science.gov (United States)

    Ambrósio, Alda M; Kennedy, James L; Macciardi, Fabio; King, Nicole; Azevedo, Maria H; Oliveira, Catarina R; Pato, Carlos N

    2005-01-01

    Alterations of the gamma-aminobutyric acid (GABA) system have been implicated in the pathophysiology of major psychoses. Restriction fragment length polymorphisms associated with the human gamma-aminobutyric acid type A (GABAA) beta2 and GABAA gamma2 subunit genes on chromosome 5q32-q35 were tested to determine whether they confer susceptibility to major psychoses. Thirty-two schizophrenic families and 25 bipolar families were tested for linkage. Nonparametric linkage (NPL) analysis performed by GENEHUNTER showed no significant NPL scores for both genes in schizophrenia (GABAA beta2: NPL narrow= -0.450; NPL broad= -0.808; GABAA gamma2: NPL narrow=0.177; NPL broad= -0.051) or bipolar disorder (GABAA beta2: NPL narrow=0.834; NPL broad=0.783; GABAA gamma2: NPL narrow= -0.159; NPL broad=0.070). Linkage analysis does not support the hypothesis that variants within the GABAA beta2 and GABAA gamma2 genes are significantly linked to major psychoses in a Portuguese population.

  6. 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.

  7. 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.

  8. Mediator subunit MED1 is a T3-dependent and T3-independent coactivator on the thyrotropin β gene promoter

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    Matsui, Keiji; Oda, Kasumi; Mizuta, Shumpei; Ishino, Ruri; Urahama, Norinaga; Hasegawa, Natsumi [Laboratory of Hematology, Division of Medical Biophysics, Kobe University Graduate School of Health Sciences, 7-10-2 Tomogaoka, Suma-ku, Kobe 654-0142 (Japan); Roeder, Robert G. [Laboratory of Biochemistry and Molecular Biology, The Rockefeller University, 1230 York Avenue, New York, NY 10065 (United States); Ito, Mitsuhiro, E-mail: itomi@med.kobe-u.ac.jp [Laboratory of Hematology, Division of Medical Biophysics, Kobe University Graduate School of Health Sciences, 7-10-2 Tomogaoka, Suma-ku, Kobe 654-0142 (Japan); Laboratory of Biochemistry and Molecular Biology, The Rockefeller University, 1230 York Avenue, New York, NY 10065 (United States); Department of Family and Community Medicine, Kobe University Graduate School of Medicine, 7-5-1 Kusunoki-cho, Chuo-ku, Kobe 654-0142 (Japan); Consolidated Research Institute for Advanced Science and Medical Care, Waseda University, 3-4-1 Okubo, Shinjuku-ku, Tokyo 159-8555 (Japan)

    2013-10-11

    Highlights: •MED1 is a bona fide T3-dependent coactivator on TSHB promoter. •Mice with LxxLL-mutant MED1 have attenuated TSHβ mRNA and thyroid hormone levels. •MED1 activates TSHB promoter T3-dependently in cultured cells. •T3-dependent MED1 action is enhanced when SRC1/SRC2 or HDAC2 is downregulated. •MED1 is also a T3-independent GATA2/Pit1 coactivator on TSHB promoter. -- Abstract: The MED1 subunit of the Mediator transcriptional coregulator complex is a nuclear receptor-specific coactivator. A negative feedback mechanism of thyroid-stimulating hormone (TSH, or thyrotropin) expression in the thyrotroph in the presence of triiodothyronine (T3) is employed by liganded thyroid hormone receptor β (TRβ) on the TSHβ gene promoter, where conventional histone-modifying coactivators act as corepressors. We now provide evidence that MED1 is a ligand-dependent positive cofactor on this promoter. TSHβ gene transcription was attenuated in MED1 mutant mice in which the nuclear receptor-binding ability of MED1 was specifically disrupted. MED1 stimulated GATA2- and Pit1-mediated TSHβ gene promoter activity in a ligand-independent manner in cultured cells. MED1 also stimulated transcription from the TSHβ gene promoter in a T3-dependent manner. The transcription was further enhanced when the T3-dependent corepressors SRC1, SRC2, and HDAC2 were downregulated. Hence, MED1 is a T3-dependent and -independent coactivator on the TSHβ gene promoter.

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

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    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.

  10. 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.

  11. 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

  12. 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.

  13. Sequence and structure of the mouse gene coding for the largest neurofilament subunit.

    NARCIS (Netherlands)

    J-P. Julien (Jean-Pierre); F. Cote; L. Beaudet (Lucille); M. Sidky (Malak); D. Flavell (David); F.G. Grosveld (Frank); W. Mushynski (Walter)

    1988-01-01

    textabstractWe have determined the complete nucleotide sequence of the mouse gene encoding the neurofilament NF-H protein. The C-terminal domain of NF-H is very rich in charged amino acids (aa) and contains a 3-aa sequence, Lys-Ser-Pro, that is repeated 51 times within a stretch of 368 aa. The

  14. 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

  15. Two novel functional mutations in the Na+,K+-ATPase alpha2-subunit ATP1A2 gene in patients with familial hemiplegic migraine and associated neurological phenotypes.

    NARCIS (Netherlands)

    Castro, M.J.; Nunes, B.; Vries, B. de; Lemos, C.; Molkot, K.R. van; Heuvel, J.J.M.W. van den; Temudo, T.; Barros, J.; Sequeiros, J.; Frants, R.R.; Koenderink, J.B.; Pereira-Monteiro, J.M.; Maagdenberg, A.M. van den

    2008-01-01

    Mutations in the ATP1A2 gene, encoding the alpha2-subunit of the Na+,K+-ATPase, are associated with familial hemiplegic migraine type 2. The majority of ATP1A2 mutations were reported in patients with hemiplegic migraine without any additional neurological findings. Here, we report on two novel ATP1

  16. Definition of the low molecular weight glutenin subunit gene family members in a set of standard bread wheat (Triticum aestivum L.) varieties

    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 viscoelastic properties of wheat dough. Most of the LMW-GSs are encoded by a multi-gene family located on the short arms of the homoeologous group 1 chromosomes, at...

  17. 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...

  18. 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

  19. Spurious Amplification of a Plasmodium vivax Small-Subunit RNA Gene by Use of Primers Currently Used To Detect P. knowlesi▿

    Science.gov (United States)

    Imwong, Mallika; Tanomsing, Naowarat; Pukrittayakamee, Sasithon; Day, Nicholas P. J.; White, Nicholas J.; Snounou, Georges

    2009-01-01

    The PCR primers commonly used to detect Plasmodium knowlesi infections in humans were found to cross-react stochastically with P. vivax genomic DNA. A nested primer set that targets one of the P. knowlesi small-subunit rRNA genes was validated for specificity and for sensitivity of detection of <10 parasite genomes. PMID:19812279

  20. 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

  1. 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.

  2. ANKS1B Gene Product AIDA-1 Controls Hippocampal Synaptic Transmission by Regulating GluN2B Subunit Localization.

    Science.gov (United States)

    Tindi, Jaafar O; Chávez, Andrés E; Cvejic, Svetlana; Calvo-Ochoa, Erika; Castillo, Pablo E; Jordan, Bryen A

    2015-06-17

    NMDA receptors (NMDARs) are key mediators of glutamatergic transmission and synaptic plasticity, and their dysregulation has been linked to diverse neuropsychiatric and neurodegenerative disorders. While normal NMDAR function requires regulated expression and trafficking of its different subunits, the molecular mechanisms underlying these processes are not fully understood. Here we report that the amyloid precursor protein intracellular domain associated-1 protein (AIDA-1), which associates with NMDARs and is encoded by ANKS1B, a gene recently linked to schizophrenia, regulates synaptic NMDAR subunit composition. Forebrain-specific AIDA-1 conditional knock-out (cKO) mice exhibit reduced GluN2B-mediated and increased GluN2A-mediated synaptic transmission, and biochemical analyses show AIDA-1 cKO mice have low GluN2B and high GluN2A protein levels at isolated hippocampal synaptic junctions compared with controls. These results are corroborated by immunocytochemical and electrophysiological analyses in primary neuronal cultures following acute lentiviral shRNA-mediated knockdown of AIDA-1. Moreover, hippocampal NMDAR-dependent but not metabotropic glutamate receptor-dependent plasticity is impaired in AIDA-1 cKO mice, further supporting a role for AIDA-1 in synaptic NMDAR function. We also demonstrate that AIDA-1 preferentially associates with GluN2B and with the adaptor protein Ca(2+)/calmodulin-dependent serine protein kinase and kinesin KIF17, which regulate the transport of GluN2B-containing NMDARs from the endoplasmic reticulum (ER) to synapses. Consistent with this function, GluN2B accumulates in ER-enriched fractions in AIDA-1 cKO mice. These findings suggest that AIDA-1 regulates NMDAR subunit composition at synapses by facilitating transport of GluN2B from the ER to synapses, which is critical for NMDAR plasticity. Our work provides an explanation for how AIDA-1 dysfunction might contribute to neuropsychiatric conditions, such as schizophrenia. Copyright

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

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    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.

  4. Duplication and Loss of Function of Genes Encoding RNA Polymerase III Subunit C4 Causes Hybrid Incompatibility in Rice

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    Giao Ngoc Nguyen

    2017-08-01

    Full Text Available Reproductive barriers are commonly observed in both animals and plants, in which they maintain species integrity and contribute to speciation. This report shows that a combination of loss-of-function alleles at two duplicated loci, DUPLICATED GAMETOPHYTIC STERILITY 1 (DGS1 on chromosome 4 and DGS2 on chromosome 7, causes pollen sterility in hybrid progeny derived from an interspecific cross between cultivated rice, Oryza sativa, and an Asian annual wild rice, O. nivara. Male gametes carrying the DGS1 allele from O. nivara (DGS1-nivaras and the DGS2 allele from O. sativa (DGS2-T65s were sterile, but female gametes carrying the same genotype were fertile. We isolated the causal gene, which encodes a protein homologous to DNA-dependent RNA polymerase (RNAP III subunit C4 (RPC4. RPC4 facilitates the transcription of 5S rRNAs and tRNAs. The loss-of-function alleles at DGS1-nivaras and DGS2-T65s were caused by weak or nonexpression of RPC4 and an absence of RPC4, respectively. Phylogenetic analysis demonstrated that gene duplication of RPC4 at DGS1 and DGS2 was a recent event that occurred after divergence of the ancestral population of Oryza from other Poaceae or during diversification of AA-genome species.

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

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    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.

  6. 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.

  7. 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.

  8. 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.

  9. Pituitary control of branchial NCC, NKCC and Na(+), K (+)-ATPase α-subunit gene expression in Nile tilapia, Oreochromis niloticus.

    Science.gov (United States)

    Breves, Jason P; Seale, Andre P; Moorman, Benjamin P; Lerner, Darren T; Moriyama, Shunsuke; Hopkins, Kevin D; Grau, E Gordon

    2014-05-01

    This study investigated endocrine control of branchial ionoregulatory function in Nile tilapia (Oreochromis niloticus) by prolactin (Prl188 and Prl177), growth hormone (Gh) and cortisol. Branchial expression of Na(+)/Cl(-) cotransporter (ncc) and Na(+)/K(+)/2Cl(-) cotransporter (nkcc) genes were employed as specific markers for freshwater- and seawater-type ionocytes, respectively. We further investigated whether Prl, Gh and cortisol direct expression of two Na(+), K(+)-ATPase (nka)-α1 subunit genes, denoted nka-α1a and nka-α1b. Tilapia transferred to fresh water following hypophysectomy failed to adequately activate gill ncc expression; ncc expression was subsequently restored by Prl replacement. Prl188 and Prl177 stimulated ncc expression in cultured gill filaments in a concentration-related manner, suggesting that ncc is regulated by Prl in a gill-autonomous fashion. Tilapia transferred to brackish water (23 ‰) following hypophysectomy exhibited a reduced capacity to up-regulate nka-α1b expression. However, Gh and cortisol failed to affect nka-α1b expression in vivo. Similarly, we found no clear effects of Gh or cortisol on nkcc expression both in vivo and in vitro. When considered with patterns previously described in euryhaline Mozambique tilapia (O. mossambicus), the current study suggests that ncc is a conserved target of Prl in tilapiine cichlids. In addition, we revealed contrasting dependencies upon the pituitary to direct nka-α1b expression in hyperosmotic environments between Nile and Mozambique tilapia.

  10. 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.

  11. 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.

  12. 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.

  13. 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.

  14. 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.

  15. 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...

  16. 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.

  17. 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.

  18. 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.

  19. Barber: Sinfonie Nr. 1, Op. 9, Neeme Järvi / Bernhard Uske

    Index Scriptorium Estoniae

    Uske, Bernhard

    1991-01-01

    Uuest heliplaadist "Barber: Sinfonie Nr. 1, Op. 9. Ouvertüre School for Scandal, Op. 5; Beach: Sinfonie e-Moll, Op. 32, "Gaelic". Detroit Symphony Orchestra /Neeme Järvi". Chandes cassette ABTD 1550; CD CHAN 8958 (72 minutes)

  20. 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.

  1. Analysis of the neurofilament heavy subunit (NFH) gene in familial amyotrophic lateral sclerosis

    Energy Technology Data Exchange (ETDEWEB)

    Rooke, K.; Rouleau, G.A. [McGill Univ., Montreal (Canada); Figlewicz, D.A. [Univ. of Rochester Medical Center, NY (United States)

    1994-09-01

    Amyotrophic lateral sclerosis (ALS) is a fatal, adult-onset, degenerative disorder of the motor neurons in the cortex, brainstem and spinal cord. Approximately 10% of ALS cases are familial (FALS) and are inherited as an age-dependent autosomal dominant trait. Mutations in the Cu/Zn superoxide dismutase (SOD-1) gene on chromosome 21 have been found in a subset of cases. However, for the remaining FALS cases, the etiology is unknown. The abnormal accumulation of neurofilaments in the cell body and proximal axon of motor neurons is a characteristic pathological finding in ALS. Furthermore, aberrant neuronal swellings that closely resemble those found in ALS have been reported in transgenic mice overexpressing NFH. The C-terminal region of NFH contains a unique functional domain with multiple repeats of the amino acids (Lys-Ser-Pro) (KSP) and forms the side-arms which appear, at the level of electron microscopy, to cross-link neurofilaments. Recently, deletions in the DSP repeat domain have been identified in five ALS patients diagnosed as sporadic cases of the disease. Based on these findings, we propose to analyze all 4 exons of the NFH gene for variation in FALS. DNA from 110 FALS cases has been amplified by the polymerase chain reaction (PCR) and analyzed by single strand conformation polymorphism (SSCP) analysis. Exon 2, exon 3 and the KSP repeat domain (part of exon 4) appear normal in all our FALS individuals under several different SSCP conditions. The analysis of exon 1 and the remainder of exon 4 has yet to be completed.

  2. Detection of the gene encoding the small subunit of the CO dehydrogenase enzyme in the H{sub 2}-evolving bacterium Rubrivivax gelatinosus CBS

    Energy Technology Data Exchange (ETDEWEB)

    Kish, A.; Levin, D. [Victoria Univ., BC (Canada)]|[Victoria Univ., BC (Canada)

    2001-06-01

    A purple non-sulfur bacterium, Rubrivivax gelatinosus CBS presents great opportunities, on a commercial scale, for the biological hydrogen production. A water-gas shift reaction is catalyzed when the bacterium is cultured in the presence of carbon oxide in the dark. The result is carbon monoxide (and water) being shifted into hydrogen (H{sub 2}) and carbon dioxide in near stoichiometric quantities. The production of hydrogen as a clean alternative fuel could be accomplished by using carbon monoxide generated from gasified waste biomass, using the bacterial water-gas shift reaction for that purpose. The characterization of three key enzymes and the genes encoding them was performed in a closely related purple non-sulfur bacterium called Rhodospirillum rubrum. They were: (1) a carbon monoxide dehydrogenase (CODH), (2) the ferredoxin-like electron-carrier small subunit of the CODH enzyme, and (3) an hydrogen-evolving hydrogenase. A transcriptional unit separate from the genes encoding the CODH and its ferredoxin-like small subunit encode the genes for the hydrogenase. A fragment of the Rhodospirillum rubrum ferredoxin-like subunit gene was amplified through the use of a polymerase chain reaction. Southern blots of restriction endonuclease digested genomic deoxyribonucleic acid (DNA) extracted from Rubrivivax gelatinosus CBS was probed with the fragment of the Rhodospirillum rubrum previously amplified using the polymerase chain reaction. Confirmation of the identification is being confirmed, while the gene is sequenced. 25 refs., 2 figs.

  3. 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.

  4. 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.

  5. Molecular cloning and functional characterization of two novel high molecular weight glutenin subunit genes in Aegilops markgrafii

    Indian Academy of Sciences (India)

    XUYE DU; XIAOCUN ZHANG

    2017-09-01

    The high molecular weight glutenin subunits (HMW-GS) in bread wheat are major determinants of the viscoelastic properties of dough and the end-use quality of wheat flour. Two novel HMW-GSs, 1Cx1.1 and 1Cy9.1, from the diploid speciesAegilops markgrafii (CC) were identified in the present study. The corresponding open-reading frames of the genes of 1Cx1.1 and 1Cy9.1 were isolated and sequenced using allele-specific polymerase chain reaction. Sequence comparison demonstrated that the HMW-GSs from Ae. markgrafii possess a similar primary structure to the homologous proteins in wheat and related species. A tandem tripeptide exists in the central repetitive domain of 1Cx1.1, and this unique structure is very rare in the HMW-GSs of other genomes. To confirm the authenticity of these isolated endogenous HMW-GS, the heterologous proteins produced by removing the signal peptides expressed by E. coli exhibited the same electrophoretic mobility as the native proteins. Subsequently, the singleprotein was purified at a sufficient scale for incorporation into flour to performsodium dodecyl sulphate (SDS) sedimentation testing. Notably, the SDS sedimentation volume was less with the addition of 1Cx1.1 than it was with 1Cy9.1.

  6. Association of the nicotinic receptor α7 subunit gene (CHRNA7 with schizophrenia and visual backward masking

    Directory of Open Access Journals (Sweden)

    George eBakanidze

    2013-10-01

    Full Text Available The nicotinic system is involved in the pathophysiology of schizophrenia. However, very little is known about its genetic basis and how it relates to clinical symptoms and potentially pharmacological intervention. Here, we investigated five single nucleotide polymorphisms (SNPs [rs3826029] [rs2337506] [rs982574] [rs904952] [rs2337980] of the cholinergic nicotinic receptor gene, alpha 7 subunit (CHRNA7 and their association to schizophrenia. We found an association with rs904952 (p=0.009 in a German sample of 224 schizophrenic patients and 224 healthy control subjects. The same trend was shown in an independent Georgian sample of 50 schizophrenic patients, 57 first order unaffected relatives, and 51 healthy controls. In addition, visual backward masking (VBM, a sensitive test for early visual information processing, was assessed in the Georgian sample. In line with prior studies, VBM performance deficits were much more pronounced in schizophrenic patients and their unaffected relatives compared to healthy controls (schizophrenic patients: 156 ms; unaffected relatives: 60 ms; healthy controls: 33 ms. VBM was strongly correlated with SNP rs904952 (H[2]=7.3, p=0.026. Our results further support the notion that changes in the nicotinic system are involved in schizophrenia and open the avenue for pharmacological intervention.

  7. 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

  8. 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.

  9. 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.

  10. 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.

  11. 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.

  12. ASSESSMENT OF GENETIC VARIATION OF PEARL OYSTER, Pinctada maxima, BASED ON THE ANALYSIS OF MITOCHONDRIAL CYTOCHROME OXIDASE SUBUNIT I GENE

    Directory of Open Access Journals (Sweden)

    Achmad Sudradjat

    2009-06-01

    Full Text Available Pearl oyster, Pinctada maxima is one of economical ly important species in aquaculture, particularly in pearl industry. Information on genetic variation of pearl oyster is required in order to be able to make a sound management of it’s natural populations and to utilize it to improve the quality of pearl culture. Five populations from different geographic locations of pearl oyster, Pinctada maxima, (Sumbawa, Bali, Selat Sunda, Belitung, and South Sulawesi were analyzed for genetic variation within a 750-base pair region of the Mitochondrial Cytochrome Oxidase subunit I (MtCOI gene using Restriction Fragment Length Polymorphism (RFLP technique. The analysis of 25 pearl oyster samples, their haplotype diversity ranged from 0.0970 to 0.1939 and the number of haplotype in each population ranged from three to five haplotypes. Clustering of populations based on Nei’s genetic distances and constructed using unweighted pair-group method with Arithmetic mean (UPGMA showed that the populations were clustered into two groups: Belitung, Selat Sunda, Bali and Sumbawa in one group, while South Sulawesi in the second group.

  13. 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.

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

    Directory of Open Access Journals (Sweden)

    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. 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

  16. 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.

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

    Directory of Open Access Journals (Sweden)

    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.

  18. 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

  19. 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.

  20. The bacteriophage P1 hot gene, encoding a homolog of the E. coli DNA polymerase III theta subunit, is expressed during both lysogenic and lytic growth stages.

    Science.gov (United States)

    Chikova, Anna K; Schaaper, Roel M

    2007-11-01

    The bacteriophage P1 hot gene product is a homolog of the theta subunit of E. coli DNA polymerase III. Previous studies with hot cloned on a plasmid have shown that Hot protein can substitute for theta, as evidenced by its stabilizing effect on certain dnaQ mutator mutants carrying an unstable pol III proofreading subunit (epsilon subunit). These results are consistent with Hot, like theta, being a replication protein involved in stabilizing the intrinsically unstable epsilon proofreading function. However, the function of hot for the viral life cycle is less clear. In the present study, we show that the hot gene is not essential. Based on its promoter structure, hot has been previously classified as a "late" phage gene, a property that is not easily reconciled with a presumed replication function. Here, we clarify this issue by demonstrating that P1 hot is actively expressed both during the lysogenic state and in the early stages of a lytic induction, in addition to its expression in the late stage of phage development. The results indicate that P1 hot has a complex expression pattern, compatible with a model in which Hot may affect the host replication machinery to benefit overall phage replication.

  1. 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

  2. Subunit-specific mechanisms and proton sensitivity of NMDA receptor channel block.

    Science.gov (United States)

    Dravid, Shashank M; Erreger, Kevin; Yuan, Hongjie; Nicholson, Katherine; Le, Phuong; Lyuboslavsky, Polina; Almonte, Antoine; Murray, Ernest; Mosely, Cara; Barber, Jeremy; French, Adam; Balster, Robert; Murray, Thomas F; Traynelis, Stephen F

    2007-05-15

    We have compared the potencies of structurally distinct channel blockers at recombinant NR1/NR2A, NR1/NR2B, NR1/NR2C and NR1/NR2D receptors. The IC50 values varied with stereochemistry and subunit composition, suggesting that it may be possible to design subunit-selective channel blockers. For dizocilpine (MK-801), the differential potency of MK-801 stereoisomers determined at recombinant NMDA receptors was confirmed at native receptors in vitro and in vivo. Since the proton sensor is tightly linked both structurally and functionally to channel gating, we examined whether blocking molecules that interact in the channel pore with the gating machinery can differentially sense protonation of the receptor. Blockers capable of remaining trapped in the pore during agonist unbinding showed the strongest dependence on extracellular pH, appearing more potent at acidic pH values that promote channel closure. Determination of pK(a) values for channel blockers suggests that the ionization of ketamine but not of other blockers can influence its pH-dependent potency. Kinetic modelling and single channel studies suggest that the pH-dependent block of NR1/NR2A by (-)MK-801 but not (+)MK-801 reflects an increase in the MK-801 association rate even though protons reduce channel open probability and thus MK-801 access to its binding site. Allosteric modulators that alter pH sensitivity alter the potency of MK-801, supporting the interpretation that the pH sensitivity of MK-801 binding reflects the changes at the proton sensor rather than a secondary effect of pH. These data suggest a tight coupling between the proton sensor and the ion channel gate as well as unique subunit-specific mechanisms of channel block.

  3. 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.

  4. Cloning and gene expression of a cDNA for the chicken follicle-stimulating hormone (FSH)-beta-subunit.

    Science.gov (United States)

    Shen, San-Tai; Yu, John Yuh-Lin

    2002-02-15

    Follicle-stimulating hormone (FSH) is a member of pituitary glycoprotein hormones that are composed of two dissimilar subunits, alpha and beta. Very little information is available regarding the nucleotide and amino acid sequence of FSH-beta in avian species. For better understanding of the phylogenic diversity and evolution of FSH molecule, we have isolated and sequenced the complete complementary DNA (cDNA) encoding chicken FSH-beta precursor molecule by reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA end (RACE) methods. The cloned chicken FSH-beta cDNA consists of 2457-bp nucleotides, including 44-bp nucleotides of the 5'-untranslated region (UTR), 396 bp of the open reading frame, and an extraordinarily long 3'-UTR of 2001-bp nucleotides followed by a poly(A)((16)) tail. It encodes a 131-amino-acid precursor molecule of FSH-beta-subunit with a signal peptide of 20 amino acids followed by a mature protein of 111 amino acids. Twelve cysteine residues, forming six disulfide bonds within beta-subunit and two putative asparagine-linked glycosylation sites, are also conserved in the chicken FSH-beta-subunit. Four proline residues, presumably responsible for changing the backbone direction of protein structure, are conserved in chicken FSH-beta-subunit as well. The nucleotide sequence of chicken FSH-beta cDNA shows high homology with quail FSH-beta cDNA, 97% homology in the open reading frame, and 85% homology in the 3'-UTR. The deduced amino acid sequence of chicken FSH-beta-subunit shows a remarkable similarity to other avian FSH-beta-subunits, 98% homology with quail, and 93% homology with ostrich, whereas a lower similarity (66 to 70%) is noted when compared with mammalian FSH-beta-subunits. By contrast, when comparing with the beta-subunits of chicken luteinizing hormone and thyroid-stimulating hormone, the homologies are as low as 37 and 40%, respectively. FSH-beta mRNA was only expressed in pituitary gland out of various

  5. Treatment with dehydroepiandrosterone sulfate increases NR1 and NR2 subunits of NMDA receptors in rat hippocampus.

    Institute of Scientific and Technical Information of China (English)

    E.DARTOIS; W.PINOTEAU; M.VINCENS

    2004-01-01

    Neurosteroids are present in the central nervous system (CNS) and can be allosteric modulators of neurotransmitter receptors. One of them, the dehydroepiandrosterone sulfate (DHEAS) has been shown to modulate the NMDA receptor, a subtype of glutamate receptor. These NMDA receptors are known to be involved in long-term potentiation and in learning and memory (Tsien 2000). Moreover, DHEAS has been reported

  6. Autoantibodies against the N-Methyl-d-Aspartate Receptor Subunit NR1: Untangling Apparent Inconsistencies for Clinical Practice

    Science.gov (United States)

    Ehrenreich, Hannelore

    2017-01-01

    This viewpoint review provides an integrative picture of seemingly contradictory work published on N-methyl-d-aspartate receptor 1 (NMDAR1) autoantibodies (AB). Based on the present state of knowledge, it gives recommendations for the clinical decision process regarding immunosuppressive treatment. Brain antigen-directed AB in general and NMDAR1-AB in particular belong to a preexisting autoimmune repertoire of mammals including humans. Specific autoimmune reactive B cells may get repeatedly (perhaps transiently) boosted by various potential stimulants (e.g., microbiome, infections, or neoplasms) plus less efficiently suppressed over lifespan (gradual loss of tolerance), likely explaining the increasing seroprevalence upon aging (>20% NMDAR1-AB in 80-year-old humans). Pathophysiological significance emerges (I) when AB-specific plasma cells settle in the brain and produce large amounts of brain antigen-directed AB intrathecally and/or (II) in conditions of compromised blood–brain barrier (BBB), for instance, upon injury, infection, inflammation, or genetic predisposition (APOE4 haplotype), which then allows substantial access of circulating AB to the brain. Regarding NMDAR1-AB, functional effects on neurons in vitro and elicitation of brain symptoms in vivo have been demonstrated for immunoglobulin (Ig) classes, IgM, IgA, and IgG. Under conditions of brain inflammation, intrathecal production and class switch to IgG may provoke high NMDAR1-AB (and other brain antigen-directed AB) levels in cerebrospinal fluid (CSF) and serum, causing the severe syndrome named “anti-NMDAR encephalitis,” which then requires immunosuppressive therapy on top of the causal encephalitis treatment (if available). However, negative CSF NMDAR1-AB results cannot exclude chronic effects of serum NMDAR1-AB on the central nervous system, since the brain acts as “immunoprecipitator,” particularly in situations of compromised BBB. In any case of suspected symptomatic consequences of circulating AB directed against brain antigens, leakiness of the BBB should be evaluated by CSF analysis (albumin quotient as proxy) and magnetic resonance imaging before considering immunosuppression. PMID:28298911

  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. Anti-epileptic effects of neuropeptide Y gene transfection into the rat brain

    Institute of Scientific and Technical Information of China (English)

    Changzheng Dong; Wenqing Zhao; Wenling Li; Peiyuan Lv; Xiufang Dong

    2013-01-01

    Neuropeptide Y gene transfection into normal rat brain tissue can provide gene overexpression, which can attenuate the severity of kainic acid-induced seizures. In this study, a recombinant adeno-associated virus carrying the neuropeptide Y gene was transfected into brain tissue of rats with kainic acid-induced epilepsy through stereotactic methods. Following these transfections, we verified overexpression of the neuropeptide Y gene in the epileptic brain. Electroencephalograms showed that seizure severity was significantly inhibited and seizure latency was significantly prolonged up to 4 weeks after gene transfection. Moreover, quantitative fluorescent PCR and western blot assays revealed that the mRNA and protein expression of the N-methyl-D-aspartate receptor subunits NR1, NR2A, and NR2B was inhibited in the hippocampus of epileptic rats. These findings indicate that neuropeptide Y may inhibit seizures via down-regulation of the functional expression of N-methyl-D-aspartate receptors.

  10. 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.

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

    Science.gov (United States)

    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.

  12. 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.

  13. 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.

  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. 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.

  16. 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...

  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. NR1I2 polymorphisms are related to tacrolimus dose-adjusted exposure and BK viremia in adult kidney transplantation

    DEFF Research Database (Denmark)

    Barraclough, Katherine A; Isbel, Nicole M; Lee, Katie J

    2012-01-01

    , median (interquartile range) dose-adjusted exposure to tacrolimus was significantly higher in individuals carrying the NR1I2 8055T variant allele, when compared with exposure in wild-type individuals [20 (14, 22) μg·h/L/mg versus 15 (9, 24) μg·h/L/mg; P =0.0007]. Using multivariable logistic regression......, NR1I2 8055T carrier status was independently predictive of higher dose-adjusted tacrolimus exposure (P=0.0005). Moreover, BK viremia was seen significantly more frequently in NR1I2 8055T allele carriers compared with wild-type individuals (38% vs 18%, P=0.005) and possession of the NR1I2 8055T allele...... imposed significantly higher odds of BK viremia (adjusted odds ratio, 2.76 [95% confidence interval, 1.33-7.73]; P=0.006). No significant difference in geometric mean peak BK viral replication titer was observed between 8055T carriers and noncarriers. No NR1I2 SNP or haplotype was significantly...

  1. 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.

  2. 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.

  3. 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

  4. 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.

  5. Redescriptions of three trachelocercid ciliates (Protista, Ciliophora, Karyorelictea), with notes on their phylogeny based on small subunit rRNA gene sequences.

    Science.gov (United States)

    Yan, Ying; Xu, Yuan; Yi, Zhenzhen; Warren, Alan

    2013-09-01

    Three trachelocercid ciliates, Kovalevaia sulcata (Kovaleva, 1966) Foissner, 1997, Trachelocerca sagitta (Müller, 1786) Ehrenberg, 1840 and Trachelocerca ditis (Wright, 1982) Foissner, 1996, isolated from two coastal habitats at Qingdao, China, were investigated using live observation and silver impregnation methods. Data on their infraciliature and morphology are supplied. The small subunit rRNA (SSU rRNA) genes of K. sulcata and Trachelocerca sagitta were sequenced for the first time. Phylogenetic analyses based on SSU rRNA gene sequence data indicate that both organisms, and the previously sequenced Trachelocerca ditis, are located within the trachelocercid assemblage and that K. sulcata is sister to an unidentified taxon forming a clade that is basal to the core trachelocercids.

  6. 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.

  7. 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.

  8. 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.

  9. Environmental contaminants activate human and polar bear (Ursus maritimus) pregnane X receptors (PXR, NR1I2) differently

    Energy Technology Data Exchange (ETDEWEB)

    Lille-Langøy, Roger, E-mail: Roger.lille-langoy@bio.uib.no [University of Bergen, Department of Biology, P.O. Box 7803, N-5020 Bergen (Norway); Goldstone, Jared V. [Woods Hole Oceanographic Institution, 266 Woods Hole Road, 02543-1050 Woods Hole, MA (United States); Rusten, Marte [University of Bergen, Department of Molecular Biology, P.O. Box 7803, N-5020 Bergen (Norway); Milnes, Matthew R. [Mars Hill University, 100 Athletic Street, Box 6671, Mars Hill, 28754 NC (United States); Male, Rune [University of Bergen, Department of Molecular Biology, P.O. Box 7803, N-5020 Bergen (Norway); Stegeman, John J. [Woods Hole Oceanographic Institution, 266 Woods Hole Road, 02543-1050 Woods Hole, MA (United States); Blumberg, Bruce [University of California, Irvine, 92697 CA (United States); Goksøyr, Anders [University of Bergen, Department of Biology, P.O. Box 7803, N-5020 Bergen (Norway)

    2015-04-01

    Background: Many persistent organic pollutants (POPs) accumulate readily in polar bears because of their position as apex predators in Arctic food webs. The pregnane X receptor (PXR, formally NR1I2, here proposed to be named promiscuous xenobiotic receptor) is a xenobiotic sensor that is directly involved in metabolizing pathways of a wide range of environmental contaminants. Objectives: In the present study, we comparably assess the ability of 51 selected pharmaceuticals, pesticides and emerging contaminants to activate PXRs from polar bears and humans using an in vitro luciferase reporter gene assay. Results: We found that polar bear PXR is activated by a wide range of our test compounds (68%) but has a slightly more narrow ligand specificity than human PXR that was activated by 86% of the 51 test compounds. The majority of the agonists identified (70%) produces a stronger induction of the reporter gene via human PXR than via polar bear PXR, however with some notable and environmentally relevant exceptions. Conclusions: Due to the observed differences in activation of polar bear and human PXRs, exposure of each species to environmental agents is likely to induce biotransformation differently in the two species. Bioinformatics analyses and structural modeling studies suggest that amino acids that are not part of the ligand-binding domain and do not interact with the ligand can modulate receptor activation. - Highlights: • Comparative study of ligand activation of human and polar bear PXRs. • Polar bear PXR is a promiscuous ligand-activated nuclear receptor but less so than human PXR. • Environmental contaminants activate human and polar bear PXRs differently. • Expression and ligand promiscuity indicate that PXR is a xenosensor in polar bears.

  10. 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.

  11. 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.

  12. 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.

  13. 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

  14. 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.

  15. 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.

  16. 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.

  17. 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.

  18. Spinal interleukin-17 promotes thermal hyperalgesia and NMDA NR1 phosphorylation in an inflammatory pain rat model.

    Science.gov (United States)

    Meng, Xianze; Zhang, Yu; Lao, Lixing; Saito, Rikka; Li, Aihui; Bäckman, Cristina M; Berman, Brian M; Ren, Ke; Wei, Pin-Kang; Zhang, Rui-Xin

    2013-02-01

    It is known that interleukin-17 (IL-17) is associated with autoimmune disorders and that peripheral IL-17 plays a role in arthritis and neuropathic pain. The present study investigated the possibility of spinal cell expression of IL-17 during inflammatory pain and possible IL-17 involvement in such pain. Hyperalgesia was induced by injecting complete Freund adjuvant (CFA, 0.08mL, 40μg Mycobacterium tuberculosis) into one hind paw of the rat. Paw withdrawal latency (PWL) was tested before (-48h) and 2 and 24h after CFA injection to assess hyperalgesia. IL-17 antibody (0.2-2μg/rat) was given intrathecally (i.t.) 24h before CFA to block the action of basal IL-17 and 2h before each of 2 PWL tests to block CFA-induced IL-17. I.t. recombinant IL-17 (10-400ng per rat) was administered to naive rats to determine its effects on PWL and phosphorylated NR1 (p-NR1). p-NR1 modulates N-methyl-d-aspartate receptor (NMDAR) activity to facilitate pain. Spinal cords were removed for IL-17 immunostaining, double immunostaining of IL-17/cell markers and IL-17 receptor A (IL-17RA)/NR1, for Western blot testing of IL-17, p-NR1, IL-17RA, and GFAP, for in situ IL-17RA hybridization, and for real time polymerase chain reaction of IL-17RA. The data reveal that IL-17 is up-regulated in activated and nonactivated astrocytes; that IL-17RA is localized in NR1-immunoreactive neurons and up-regulated; and that IL-17 antibody at 2μg/rat significantly increased PWL (P<.05) and decreased p-NR1 and IL-17RA compared to control in CFA- and IL-17-injected rats. The results suggest that spinal IL-17 is produced by astrocytes and enhances p-NR1 to facilitate pain.

  19. Novel mutations in the gene for α-subunit of retinal cone cyclic nucleotide-gated channels in a Japanese patient with congenital achromatopsia.

    Science.gov (United States)

    Kuniyoshi, Kazuki; Muraki-Oda, Sanae; Ueyama, Hisao; Toyoda, Futoshi; Sakuramoto, Hiroyuki; Ogita, Hisakazu; Irifune, Motohiro; Yamamoto, Shuji; Nakao, Akira; Tsunoda, Kazushige; Iwata, Takeshi; Ohji, Masahito; Shimomura, Yoshikazu

    2016-05-01

    To present the characteristics and pathology of a patient with congenital achromatopsia. The patient was a 22-year-old Japanese woman who was 8 years old when she first visited our clinic. Comprehensive ophthalmic examinations including visual acuity measurements, perimetry, optical coherence tomography (OCT), fundus autofluorescence (FAF) imaging, electroretinography (ERG), and color vision tests were performed. Her genomic DNA was used as the template for the amplification of exons of five candidate genes for achromatopsia; CNGA3, CNGB3, GNAT2, PDE6C, and PDE6H, and the amplified products were sequenced. A missense mutation, found in the CNGA3, was studied both electrophysiologically and biochemically. Her phenotype was typical of congenital complete achromatopsia. She was followed for 14 years, and her vision and fundus findings were stable. However, the scotopic ERG b-waves at age 22 were smaller than those at age 8, and her FAF images showed increased autofluorescence in both maculae. Genetic examinations revealed combined heterozygous mutations of c.997_998delGA and p.M424V in the CNGA3 gene. The homomeric channel consisting of the CNGA3 subunit with the p.M424V mutation had a weak cGMP-activated current in patch-clamp recordings. In heterologous expression analyses, the expression at the cell surface of the mutant CNGA3 subunit was about 28 % of the wild type. The two novel mutations found in the CNGA3 gene, c.997_998delGA and p.M424V, can cause complete achromatopsia. The vision of the patient was stationary until the third decade of life although the FAF was altered at the age of 22 years.

  20. 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.

  1. 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. Copyright © 2016 Elsevier Inc. All rights reserved.

  2. 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.

  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. 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.

  5. Integrative proteomic analysis of the NMDA NR1 knockdown mouse model reveals effects on central and peripheral pathways associated with schizophrenia and autism spectrum disorders

    NARCIS (Netherlands)

    H. Wesseling (Hendrik); P.C. Guest (Paul); C.-M. Lee (Chi-Ming); E.H.F. Wong (Erik); H. Rahmoune (Hassan); S. Bahn (Sabine)

    2014-01-01

    textabstractBackground: Over the last decade, the transgenic N-methyl-D-aspartate receptor (NMDAR) NR1-knockdown mouse (NR1neo-/-) has been investigated as a glutamate hypofunction model for schizophrenia. Recent research has now revealed that the model also recapitulates cognitive and negative symp

  6. Aquatic studies at the 100-HR-3 and 100-NR-1 operable units

    Energy Technology Data Exchange (ETDEWEB)

    Cushing, C.E.

    1993-04-01

    Pacific Northwest Laboratory initiated a program to characterize selected aquatic biological populations to determine (1) existing levels of inorganic chemical and radionuclide contamination, and (2) the populations` suitability as indicators of chemical releases during cleanup activities at the US Department of Energy`s Hanford Site. Following work plans for the ground-water operable units, lower trophic levels in the aquatic habitat (periphyton and caddisfly larvae) were evaluated for contaminants at the 100-HR-3 Operable Unit and 100-NR-1 Operable Unit. The results were evaluated to determine the need for further sampling. If the results showed no significant contamination compared to upriver levels, sampling would be discontinued. The periphyton community appears to be suitable for determining contamination levels. Baseline concentrations for stable chromium were established and will be useful for comparing samples collected when contaminant release is expected. Concentrations of {sup 60}Co, {sup 90}Sr, and {sup 137}Cs in periphyton were essentially below detectable limits, which will also make this community useful in detecting potential releases of radionuclides during cleanup activities. Levels for both stable chromium and radionuclides were essentially below detection limits for caddisfly larvae. Thus, these organisms may be used to monitor suspected contaminant releases from cleanup activities; if concentrations exceed detection limits, they may be related to these activities. Two candidate threatened and endangered species of molluscs occur in the Hanford Reach of the Columbia River. These are the shortface lanx (Fisherola nuttalli), which is a Washington State candidate species, and the Columbia pebblesnail (Fluminicola columbiana), which is both a state and federal candidate species. Specimens of the shortface lanx were observed in the vicinity of N Springs (100-NR-1 Operable Unit); they likely occur throughout this area.

  7. Aquatic studies at the 100-HR-3 and 100-NR-1 operable units

    Energy Technology Data Exchange (ETDEWEB)

    Cushing, C.E.

    1993-04-01

    Pacific Northwest Laboratory initiated a program to characterize selected aquatic biological populations to determine (1) existing levels of inorganic chemical and radionuclide contamination, and (2) the populations' suitability as indicators of chemical releases during cleanup activities at the US Department of Energy's Hanford Site. Following work plans for the ground-water operable units, lower trophic levels in the aquatic habitat (periphyton and caddisfly larvae) were evaluated for contaminants at the 100-HR-3 Operable Unit and 100-NR-1 Operable Unit. The results were evaluated to determine the need for further sampling. If the results showed no significant contamination compared to upriver levels, sampling would be discontinued. The periphyton community appears to be suitable for determining contamination levels. Baseline concentrations for stable chromium were established and will be useful for comparing samples collected when contaminant release is expected. Concentrations of [sup 60]Co, [sup 90]Sr, and [sup 137]Cs in periphyton were essentially below detectable limits, which will also make this community useful in detecting potential releases of radionuclides during cleanup activities. Levels for both stable chromium and radionuclides were essentially below detection limits for caddisfly larvae. Thus, these organisms may be used to monitor suspected contaminant releases from cleanup activities; if concentrations exceed detection limits, they may be related to these activities. Two candidate threatened and endangered species of molluscs occur in the Hanford Reach of the Columbia River. These are the shortface lanx (Fisherola nuttalli), which is a Washington State candidate species, and the Columbia pebblesnail (Fluminicola columbiana), which is both a state and federal candidate species. Specimens of the shortface lanx were observed in the vicinity of N Springs (100-NR-1 Operable Unit); they likely occur throughout this area.

  8. 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.

  9. 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.

  10. 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.

  11. Transferring a Gene Expression Cassette Lacking the Vector Backbone Sequences of the 1Ax1 High Molecular Weight Glutenin Subunit into Two Chinese Hexaploid Wheat Genotypes

    Institute of Scientific and Technical Information of China (English)

    SHI Nong-nong; HE Guang-yuan; LI Ke-xiu; WANG Hui-zhong; CHEN Guan-ping; XU Ying

    2007-01-01

    1Ax1 high molecular weight glutenin subunit (HMW-GS) gene expression cassette (GEC) lacking vector backbone sequences together with selectable marker Bar GEC were co-transformed into Chinese hexaploid cultivars Een 1 and Emai 12 to test the feasibility and the efficiency of explant regeneration, transformation frequency and transgene expression comparing with whole vector transformation by the approaches of plasmid extraction and excision, immature embryo isolation, particle co-bombardment, tissue culture, DNA extraction, PCR amplification, southern hybridization, leaf-painting test and SDS-PAGE etc. No significant difference was shown in tissue culture response of the proportion of embryogenic calli, somatic embryogenesis and regeneration frequency between GEC and whole plasmid bombarded embryos, but both regenerated less well than non-bombarded control. Total 56 plantlets that survived PPT selection had insertion of at least the Bar gene, 18 were from the GEC treatment and 38 from the whole plasmid treatment, the escape ratio averaged 0.23. Six independent transplants f230 - f235 with GEC transformation from genotype Emai 12 presented clear PCR amplification bands of Bar and 1Ax1 gene. The transformation and co-transformation frequency were 3.51 and 100% respectively. PCR amplification using a primer-pair specific for ampicillin resistant gene indicated the existence of AmpR gene in whole vectors but the removal in GECs and transplants. Southern blot of total DNA and PCR products from transgenic plants of 1Ax1 GEC confirmed the integration of the transgene 1Ax1 and the absence of the EcoR Ⅰ recognition site at both ends of the 1Ax1 GEC when integrated. SDS-PAGE showed the expression of 1Ax1 GEC and un-expression of whole plasmid. The length of integrated fragment, the proportion of the gene of interest (GOI) and the selectable marker (MG), bombardment pressure and genotypes are vital for the expression of a transformed GEC.

  12. Sibelius. Lemminkäinen-Legenden op. 22 Nr. 1-4 / Christoph Schlüren

    Index Scriptorium Estoniae

    Schlüren, Christoph

    1997-01-01

    Uuest heliplaadist "Sibelius: Lemminkäinen-Legenden op. 22 Nr. 1-4, Nächtlicher Ritt und Sonnenaufgang op. 55, Luonnotar op. 70. Stockholm Philharmoniker / Paavo Järvi. Virgin/EMI CD 545213 2 (WD:70'22")DDD

  13. Medtner: Konzert für Klavier und Orchester Nr. 1 c-Moll op. 33 / Volkmar Fischer

    Index Scriptorium Estoniae

    Fischer, Volkmar

    1992-01-01

    Uuest heliplaadist "Medtner: Konzert für Klavier und Orchester Nr. 1 c-Moll op. 33, Sonaten-Ballade für Klavier Fis-Dur op. 27. London Philharmonic Orchestra / Neeme Järvi". Chandos/Koch CD9039 (WD:54'28")

  14. Sibelius. Lemminkäinen-Legenden op. 22 Nr. 1-4 / Christoph Schlüren

    Index Scriptorium Estoniae

    Schlüren, Christoph

    1997-01-01

    Uuest heliplaadist "Sibelius: Lemminkäinen-Legenden op. 22 Nr. 1-4, Nächtlicher Ritt und Sonnenaufgang op. 55, Luonnotar op. 70. Stockholm Philharmoniker / Paavo Järvi. Virgin/EMI CD 545213 2 (WD:70'22")DDD

  15. 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.

  16. 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.

  17. 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.

  18. Added resolution among ordinal level relationships of tapeworms (Platyhelminthes: Cestoda) with complete small and large subunit nuclear ribosomal RNA genes.

    Science.gov (United States)

    Waeschenbach, Andrea; Webster, Bonnie L; Bray, Rodney A; Littlewood, D T J

    2007-10-01

    The addition of large subunit ribosomal DNA (lsrDNA) to small subunit ribosomal DNA (ssrDNA) has been shown to add resolution to phylogenies at various taxonomic levels for a diversity of phyla. We added nearly complete lsrDNA (4057-4593bp) sequences to ssrDNA (1940-2228bp) for 26 ingroup and 3 outgroup taxa in an attempt to provide an improved ordinal phylogeny for the Cestoda. Ten lsrDNA and seven ssrDNA sequences were generated from new taxa and 13 existing partial lsrDNA sequences were sequenced to completion. The majority of phylogenetic signal in the combined analysis came from lsrDNA (69.6% of parsimonious informative sites, as opposed to 30.4% obtained from ssrDNA), resulting in almost identical topologies for lsrDNA and lsr+ssrDNA (pairwise symmetric distance=6) in model-based analyses. Topology testing found trees based on partial lsrDNA (domains D1-D3)+ssrDNA and complete lsr+ssrDNA to differ significantly; the addition of lsrDNA domains D4-D12 had a significant effect on topology. Overall nodal support was greatest in the combined analysis and weakest for ssrDNA only. Our molecular phylogenies differed significantly from those based on morphology alone. Acetabulate lineages form a monophyletic group, with the Tetraphyllidea being paraphyletic. Support for the combined data was high for the following topology: (Litobothriidea (Lecanicephalidea (Rhinebothrium/Rhodobothrium (Clistobothrium (Pachybothrium(Acanthobothrium Proteocephalidea) (Mesocestoididae, Nippotaeniidea, Cyclophyllidea, Tetrabothriidea)))))); all genus names refer to tetraphyllidean lineages. Although the interrelationships among the four most derived taxa remain uncertain, overall ambiguity of the acetabulate interrelationships was reduced. The Pseudophyllidea were recovered as polyphyletic, with support for a sister-group relationship between Diphyllobothriidae and Haplobothriidea. The monophyly of the Trypanorhyncha was recovered for the first time based on molecular data. The positions

  19. A study of glutamate levels, NR1, NR2A, NR2B receptors and oxidative stress in rat model of Japanese encephalitis.

    Science.gov (United States)

    Chauhan, Prashant Singh; Misra, Usha Kant; Kalita, Jayantee

    2017-03-15

    There is paucity of studies on the role of glutamate excitotoxicity in cell damage in Japanese encephalitis. In this study the glutamate levels and its NMDA receptors, and oxidative stress markers in different brain regions have been evaluated and correlated with neurobehavioral changes at different time points. Twelve day old Wistar rats were inoculated with 3×10(6)pfu/ml intracerebrally. The neurobehavioral effects were evaluated by spontaneous locomotor activity (SLA), grip strength and rota rod test on 10, 33 and 48days post inoculation (dpi). Glutamate level was evaluated by enzyme linked immunosorbent assay, mRNA gene expression of ionotropic glutamate receptors N-methyl d-aspartate (NMDA) receptor 1, 2A and 2B (NR1, NR2A and NR2B) were evaluated by real time PCR. Malondialdehyde (MDA), glutathione (GSH) and glutathione peroxidase (GPx) levels were measured by spectrophotometer in different brain regions of JEV infected rats on 10, 33 and 48dpi. There was significant increase in motor deficit, grip strength and decreased locomotor activity on 10 and 33dpi. Glutamate levels were increased in thalamus, midbrain, frontal cortex, striatum and cerebellum on 10 and 33dpi and were followed by a recovery on 48dpi. Glutamate NMDR receptors NR1, NR2A and NR2B were reduced in thalamus, midbrain, frontal cortex, striatum and cerebellum on 10dpi which was followed by recovery after 33dpi. A significant increase in MDA level in thalamus, midbrain, frontal cortex, striatum and cerebellum was noted on 10 and 33dpi. The antioxidant GSH and GPx were significantly reduced in these brain regions on 10 and 33dpi. Glutamate, MDA, GSH and GPx correlated in different brain regions as the disease progress. Increased Glutamate level may be related to oxidative stress and may be responsible for behavioral alterations in rat model of Japanese encephalitis. Copyright © 2017 Elsevier Inc. All rights reserved.

  20. 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.

  1. Molecular Cloning, Structural Analysis and Tissue Expression of Protein Phosphatase 3 Catalytic Subunit Alpha Isoform (PPP3CA Gene in Tianfu Goat Muscle

    Directory of Open Access Journals (Sweden)

    Lu Wan

    2014-02-01

    Full Text Available Calcineurin, a Ca2+/calmodulin-dependent protein phosphatase, plays a critical role in controlling skeletal muscle fiber type. However, little information is available concerning the expression of calcineurin in goat. Therefore, protein phosphatase 3 catalytic subunit alpha isoform (PPP3CA gene, also called calcineurin Aα, was cloned and its expression characterized in Tianfu goat muscle. Real time quantitative polymerase chain reaction (RT-qPCR analyses revealed that Tianfu goat PPP3CA was detected in cardiac muscle, biceps femoris muscle, abdominal muscle, longissimus dors muscle, and soleus muscle. High expression levels were found in biceps femoris muscle, longissimus muscle and abdominal muscle (p < 0.01, and low expression levels were seen in cardiac muscle and soleus muscle (p > 0.05. In addition, the spatial-temporal mRNA expression levels showed different variation trends in different muscles with the age of the goats. Western blotting further revealed that PPP3CA protein was expressed in the above-mentioned tissues, with the highest level in biceps femoris muscle, and the lowest level in soleus muscle. In this study, we isolated the full-length coding sequence of Tianfu goat PPP3CA gene, analyzed its structure, and investigated its expression in different muscle tissues from different age stages. These results provide a foundation for understanding the function of the PPP3CA gene in goats.

  2. 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.

  3. Molecular cloning, structural analysis and tissue expression of protein phosphatase 3 catalytic subunit alpha isoform (PPP3CA) gene in Tianfu goat muscle.

    Science.gov (United States)

    Wan, Lu; Ma, Jisi; Xu, Gangyi; Wang, Daihua; Wang, Nianlu

    2014-02-07

    Calcineurin, a Ca(2+)/calmodulin-dependent protein phosphatase, plays a critical role in controlling skeletal muscle fiber type. However, little information is available concerning the expression of calcineurin in goat. Therefore, protein phosphatase 3 catalytic subunit alpha isoform (PPP3CA) gene, also called calcineurin Aα, was cloned and its expression characterized in Tianfu goat muscle. Real time quantitative polymerase chain reaction (RT-qPCR) analyses revealed that Tianfu goat PPP3CA was detected in cardiac muscle, biceps femoris muscle, abdominal muscle, longissimus dors muscle, and soleus muscle. High expression levels were found in biceps femoris muscle, longissimus muscle and abdominal muscle (p muscle and soleus muscle (p > 0.05). In addition, the spatial-temporal mRNA expression levels showed different variation trends in different muscles with the age of the goats. Western blotting further revealed that PPP3CA protein was expressed in the above-mentioned tissues, with the highest level in biceps femoris muscle, and the lowest level in soleus muscle. In this study, we isolated the full-length coding sequence of Tianfu goat PPP3CA gene, analyzed its structure, and investigated its expression in different muscle tissues from different age stages. These results provide a foundation for understanding the function of the PPP3CA gene in goats.

  4. 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.

  5. 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.

  6. Requirement of PSD-95 for dopamine D1 receptor modulating glutamate NR1a/NR2B receptor function

    Institute of Scientific and Technical Information of China (English)

    Wei-hua GU; Shen YANG; Wei-xing SHI; Guo-zhang JIN; Xue-chu ZHEN

    2007-01-01

    Aim: To elucidate the role of scaffold protein postsynaptic density (PSD)-95 in the dopamine D1 receptor (D1R)-modulated NR 1a/NR2B receptor response.Methods: The human embryonic kidney 293 cells expressing D1R (tagged with the enhanced yellow fluorescent protein) and NR1a/NR2B with or without co-expres-sion of PSD-95 were used in the experiments. The Ca2+ influx measured by imaging technique was employed to monitor N-methyl-D-aspartic acid receptors (NMDAR) function.Results: The application of dopamine (DA, 100 μmol/L) did not alter glutamate/glycine (Glu/Gly)-induced NMDAR-mediated Ca2+ influx in cells only expressing the D1R/NR1a/NR2B receptor. However, DA increased Glu/Gly-induced Ca2+ influx in a concentration-dependent manner while the cells were co-expressed with PSD-95. D1.R-stimulated Ca2+ influx was inhibited by a selective DIR antagonist SCH23390. Moreover, pre-incubation with either the protein kinase A (PKA) inhibitor H89, or the protein kinase C (PKC) inhibitor chelerythrine attenuated D1R-enhanced Ca2+ influx induced by the N-methyl-D-aspartie acid (NMDA) agonist. The results clearly indicate that D1R-modulated NR1a/NR2B receptor function depends on PSD-95 and is subjected to the regulation of PKA and PKC.Conclusion: The present study provides the fast evidence that PSD-95 is essential in D iR-regulated NR1a/NR2B receptor function.

  7. 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.

  8. 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.

  9. 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. Copyright © 2016 Elsevier B.V. All rights reserved.

  10. 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...

  11. 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%),

  12. 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

  13. 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.

  14. Phorbol-induced surface expression of NR2A subunit homologues in HEK293 cells

    Institute of Scientific and Technical Information of China (English)

    Chan-ying ZHENG; Xiu-juan YANG; Zhan-yan FU; Jian-hong LUO

    2006-01-01

    Aim: N-methyl-D-aspartate receptors (NMDAR) are heteromeric complexes primarily assembled from NR1 and NR2 subunits. In normal conditions, NR2 sub-units assemble into homodimers in the endoplasmic reticulum (ER). These homodimers remain in the ER until they coassemble with NR1 dimers and are trafficked to the cell surface. However, it still remains unclear whether functional homomeric NMDAR exist in physiological or pathological conditions. Methods: We transfected GFP-NR2A alone into HEK293 cells, treated the cells with PKC activator 12-myristate-13 acetate (PMA), and then detected surface NR2A sub-units with a live cell immunostaining method. We also used a series of NR2A mutants with a partial deletion of its C-terminus to identify the regions that are involved in the PMA-mediated surface expression of NR2A subunits. Results: NR2A subunits were expressed on the cell membrane after incubation with PMA (200 nmol/L,30 min), although no functional NMDA channels were detected after PMA-induced membrane trafficking. Immunostaining with an ER marker also revealed that NR2A subunits were exported from the ER after PMA treatment. Furthermore, the deletion of amino acids between 1149-1347 or 1354-1464 of NR2A inhibited PMA-induced surface expression of NR2A subunits. Conclusion: First, our data suggests that PMA treatment can induce the surface expression of homomeric NR2A subunits. Furthermore, this process is probably mediated by the NR2A C-terminal region between positions 1149 and 1464.

  15. 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.

  16. 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.

  17. A vacuolar-type proton (H+) translocating ATPase alpha subunit encoded by the Hc-vha-6 gene of Haemonchus contortus.

    Science.gov (United States)

    Hu, Min; He, Li; Campbell, Bronwyn E; Zhong, Weiwei; Sternberg, Paul W; Gasser, Robin B

    2010-08-01

    In the present study, a full-length cDNA (designated Hc-vha-6) inferred to encode an alpha subunit of a vacuolar-type proton translocating adenosine triphosphatase (V-ATPase) was isolated from the parasitic nematode Haemonchus contortus, and characterized. The transcript for Hc-vha-6 was detected in all developmental stages and both sexes of H. contortus. Elements, including two TATA box (TATAA), two inverted CAAT box (ATTGG), five E box (CANNTG) and six GATA as well as five inverse GATA (TTATC) transcription factor motifs, were identified in the non-coding region upstream of Hc-vha-6. The open reading frame (ORF) of 2601 nucleotides encoded a protein (Hc-VHA-6) of 866 amino acids and a molecular weight of approximately 98.7 kDa. Comparison with a published protein sequence for a homologue (VPH1P) from yeast showed that Hc-VHA-6 had nine transmembrane domains and the 14 essential amino acid residues associated with enzyme activity, assembly, intracellular and/or membrane targeting. Phylogenetic analyses of selected amino acid sequence data revealed Hc-VHA-6 to be most closely related to VHA-6 of Caenorhabditis elegans. A predictive network analysis inferred that vha-6 interacts with at least seven other genes encoding V-ATPase subunits and a small Rab GTPase. This study provides the first insight into a V-ATPase of parasitic nematodes and a sound basis for future functional genomic work. Copyright 2010 Elsevier Ltd. All rights reserved.

  18. 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.

  19. Partial gene sequences for the A subunit of methyl-coenzyme M reductase (mcrI) as a phylogenetic tool for the family Methanosarcinaceae

    Science.gov (United States)

    Springer, E.; Sachs, M. S.; Woese, C. R.; Boone, D. R.

    1995-01-01

    Representatives of the family Methanosarcinaceae were analyzed phylogenetically by comparing partial sequences of their methyl-coenzyme M reductase (mcrI) genes. A 490-bp fragment from the A subunit of the gene was selected, amplified by the PCR, cloned, and sequenced for each of 25 strains belonging to the Methanosarcinaceae. The sequences obtained were aligned with the corresponding portions of five previously published sequences, and all of the sequences were compared to determine phylogenetic distances by Fitch distance matrix methods. We prepared analogous trees based on 16S rRNA sequences; these trees corresponded closely to the mcrI trees, although the mcrI sequences of pairs of organisms had 3.01 +/- 0.541 times more changes than the respective pairs of 16S rRNA sequences, suggesting that the mcrI fragment evolved about three times more rapidly than the 16S rRNA gene. The qualitative similarity of the mcrI and 16S rRNA trees suggests that transfer of genetic information between dissimilar organisms has not significantly affected these sequences, although we found inconsistencies between some mcrI distances that we measured and and previously published DNA reassociation data. It is unlikely that multiple mcrI isogenes were present in the organisms that we examined, because we found no major discrepancies in multiple determinations of mcrI sequences from the same organism. Our primers for the PCR also match analogous sites in the previously published mcrII sequences, but all of the sequences that we obtained from members of the Methanosarcinaceae were more closely related to mcrI sequences than to mcrII sequences, suggesting that members of the Methanosarcinaceae do not have distinct mcrII genes.

  20. 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

  1. Babesia canis canis, Babesia canis vogeli, Babesia canis rossi: differentiation of the three subspecies by a restriction fragment length polymorphism analysis on amplified small subunit ribosomal RNA genes.

    Science.gov (United States)

    Carret, C; Walas, F; Carcy, B; Grande, N; Précigout, E; Moubri, K; Schetters, T P; Gorenflot, A

    1999-01-01

    The parasites Babesia canis and Babesia gibsoni (phylum Apicomplexa) are responsible for canine babesiosis throughout the world. Babesia canis was previously described as a group of three biologically different subspecies, namely B. canis canis, B. canis vogeli, and B. canis rossi. We report partial sequences of small subunit ribosomal RNA gene (ssu-rDNA) of each subspecies amplified in vitro with primers derived from a semi-conserved region of the ssu-rDNA genes in other Babesia species. The polymerase chain reaction combined with a restriction fragment length polymorphism analysis, using HinfI and TaqI restriction enzymes, confirmed the separation of B. canis into three subspecies. These sequences were compared with previously published sequences of other Babesia species. A phylogenetic approach showed that the three subspecies of B. canis belong to the clade of Babesia species sensu stricto where B. canis canis clusters with B. canis rossi whereas B. canis vogeli might form a monophyletic group with the cluster B. divergens and B. odocoilei. Our results show that the three subspecies of B. canis can readily be differentiated at the molecular level and suggest that they might be considered as true species.

  2. Decreased levels of pNR1 S897 protein in the cortex of neonatal Sprague Dawley rats with hypoxic-ischemic or NMDA-induced brain damage

    Energy Technology Data Exchange (ETDEWEB)

    Hei, Ming-Yan; Tao, Hui-Kang; Tang, Qin; Yu, Bo; Zhao, Ling-Ling [Department of Pediatrics, The Third Xiangya Hospital, Central South University, Changsha, Hunan (China)

    2012-06-22

    Our objective was to investigate the protein level of phosphorylated N-methyl-D-aspartate (NMDA) receptor-1 at serine 897 (pNR1 S897) in both NMDA-induced brain damage and hypoxic-ischemic brain damage (HIBD), and to obtain further evidence that HIBD in the cortex is related to NMDA toxicity due to a change of the pNR1 S897 protein level. At postnatal day 7, male and female Sprague-Dawley rats (13.12 ± 0.34 g) were randomly divided into normal control, phosphate-buffered saline (PBS) cerebral microinjection, HIBD, and NMDA cerebral microinjection groups. Immunofluorescence and Western blot (N = 10 rats per group) were used to examine the protein level of pNR1 S897. Immunofluorescence showed that control and PBS groups exhibited significant neuronal cytoplasmic staining for pNR1 S897 in the cortex. Both HIBD and NMDA-induced brain damage markedly decreased pNR1 S897 staining in the ipsilateral cortex, but not in the contralateral cortex. Western blot analysis showed that at 2 and 24 h after HIBD, the protein level of pNR1 S897 was not affected in the contralateral cortex (P > 0.05), whereas it was reduced in the ipsilateral cortex (P < 0.05). At 2 h after NMDA injection, the protein level of pNR1 S897 in the contralateral cortex was also not affected (P > 0.05). The levels in the ipsilateral cortex were decreased, but the change was not significant (P > 0.05). The similar reduction in the protein level of pNR1 S897 following both HIBD and NMDA-induced brain damage suggests that HIBD is to some extent related to NMDA toxicity possibly through NR1 phosphorylation of serine 897.

  3. 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

  4. 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.

  5. 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.

  6. Bestrahlungsinduziertes kriechen und schwellen des austenitischen werkstoffes NR. 1.4981 zwischen 400 und 500°C (RIPCEX I)

    Science.gov (United States)

    Herschbach, K.; Schneider, W.; Ehrlich, K.

    1981-10-01

    ZusammenfassungFür den Werkstoff Nr. 1.4981 wurde das bestrahlungsinduzierte Volumenschwellen und Kriechen im Temperaturbereich 400 bis 500°C bis zu einer Dosis von max. 63 dpa mittels nichtzerstörender und zerstörender Nachuntersuchungen bestimmt. Dabei zeigte sich eine deutliche Beeinflussung des Volumenschwellens durch eine angelegte Spannung. Das bestrahlungsinduzierte Kriechen wird für den Stahl Nr. 1.4981 durch mindestens zwei Prozesse hervorgerufen, einmal durch den sog. SIPA-Prozess, der auf bevorzugter Absorption von Zwischengitteratomen beruht, zum anderen durch das sog. I-Creep, einem Prozess, der erst nach Einsetzen des Volumenschwellens zum Tragen kommen kann. Für höhere Dosen liefert letzterer Vorgang den dominierenden Beitrag zum Kriechen.

  7. Five phosphonate operon gene products as components of a multi-subunit complex of the carbon-phosphorus lyase pathway

    DEFF Research Database (Denmark)

    Jochimsen, Bjarne; Lolle, Signe; McSorley, Fern R.

    2011-01-01

    by expression in E. coli and purification of Phn-polypeptides. PhnG, PhnH, PhnI, PhnJ, and PhnK copurify as a protein complex by ion-exchange, size-exclusion, and affinity chromatography. The five polypeptides also comigrate in native-PAGE. Cross-linking of the purified protein complex reveals a close proximity...... is suggested to be PhnG4H2I2J2K. Deletion of individual phn genes reveals that a strain harboring plasmid-borne phnGHIJ produces a protein complex consisting of PhnG, PhnH, PhnI, and PhnJ, whereas a strain harboring plasmid-borne phnGIJK produces a protein complex consisting of PhnG and PhnI. We conclude...

  8. 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.

  9. Sibelius. Lemminkäinen-Legenden op. 22 Nr. 1-4 / Christoph Schlüren

    Index Scriptorium Estoniae

    Schlüren, Christoph

    1997-01-01

    Uuest heliplaadist "Sibelius. Lemminkäinen-Legenden op. 22 Nr. 1-4, Pohjolas Tochter op. 49, Nächtlicher Ritt und Sonnenaufgang op. 55; Göteborger Sinfoniker, Neeme Järvi; DG CD 453 426-2 (WD: 70'37") DDD Võrreldud: Opus 22: Segerstam (Ondine 852-2); op. 22 und 55; Paavo Järvi" (Virgin 545 213-2); op. 49: Segerstam (Chandos 8965)

  10. Sibelius. Lemminkäinen-Legenden op. 22 Nr. 1-4 / Christoph Schlüren

    Index Scriptorium Estoniae

    Schlüren, Christoph

    1997-01-01

    Uuest heliplaadist "Sibelius. Lemminkäinen-Legenden op. 22 Nr. 1-4, Pohjolas Tochter op. 49, Nächtlicher Ritt und Sonnenaufgang op. 55; Göteborger Sinfoniker, Neeme Järvi; DG CD 453 426-2 (WD: 70'37") DDD Võrreldud: Opus 22: Segerstam (Ondine 852-2); op. 22 und 55; Paavo Järvi" (Virgin 545 213-2); op. 49: Segerstam (Chandos 8965)

  11. 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.

  12. 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.

  13. 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. Copyright © 2012 Elsevier B.V. All rights reserved.

  14. 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.

  15. 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.

  16. 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.

  17. Further consideration of the phylogeny of some "traditional" heterotrichs (Protista, Ciliophora) of uncertain affinities, based on new sequences of the small subunit rRNA gene.

    Science.gov (United States)

    Miao, Miao; Song, Weibo; Clamp, John C; Al-Rasheid, Khaled A S; Al-Khedhairy, Abdulaziz A; Al-Arifi, Saud

    2009-01-01

    The systematic relationships and taxonomic positions of the traditional heterotrich genera Condylostentor, Climacostomum, Fabrea, Folliculina, Peritromus, and Condylostoma, as well as the licnophorid genus Licnophora, were re-examined using new data from sequences of the gene coding for small subunit ribosomal RNA. Trees constructed using distance-matrix, Bayesian inference, and maximum-parsimony methods all showed the following relationships: (1) the "traditional" heterotrichs consist of several paraphyletic groups, including the current classes Heterotrichea, Armophorea and part of the Spirotrichea; (2) the class Heterotrichea was confirmed as a monophyletic assemblage based on our analyses of 31 taxa, and the genus Peritromus was demonstrated to be a peripheral group; (3) the genus Licnophora occupied an isolated branch on one side of the deepest divergence in the subphylum Intramacronucleata and was closely affiliated with spirotrichs, armophoreans, and clevelandellids; (4) Condylostentor, a recently defined genus with several truly unique morphological features, is more closely related to Condylostoma than to Stentor; (5) Folliculina, Eufolliculina, and Maristentor always clustered together with high bootstrap support; and (6) Climacostomum occupied a paraphyletic position distant from Fabrea, showing a close relationship with Condylostomatidae and Chattonidiidae despite of modest support.

  18. 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.

  19. 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.

  20. 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.

  1. 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.

  2. Two novel functional mutations in the Na+,K+-ATPase alpha2-subunit ATP1A2 gene in patients with familial hemiplegic migraine and associated neurological phenotypes.

    Science.gov (United States)

    Castro, M-J; Nunes, B; de Vries, B; Lemos, C; Vanmolkot, K R J; van den Heuvel, J J M W; Temudo, T; Barros, J; Sequeiros, J; Frants, R R; Koenderink, J B; Pereira-Monteiro, J M; van den Maagdenberg, A M J M

    2008-01-01

    Mutations in the ATP1A2 gene, encoding the alpha2-subunit of the Na+,K+-ATPase, are associated with familial hemiplegic migraine type 2. The majority of ATP1A2 mutations were reported in patients with hemiplegic migraine without any additional neurological findings. Here, we report on two novel ATP1A2 mutations that were identified in two Portuguese probands with hemiplegic migraine and interesting additional clinical features. The proband's of family 1 (with a V362E mutation) had mood alterations, classified as a borderline personality. The proband in family 2 (with a P796S mutation) had mild mental impairment, in addition to hemiplegic migraine; more severe mental retardation was observed in his brother, who also had hemiplegic migraine and carried the same mutation. Cell-survival assays clearly showed abnormal functioning of mutant Na+,K+-ATPase, indicating that both ATP1A2 mutants are disease causing. Additionally, our results suggest a possible causal relationship of the ATP1A2 mutations with the complex clinical phenotypes observed in the probands.

  3. [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.

  4. Schmidt. Sinfonie Nr. 1 E-Dur; Strauss. Vier sinfonische Zwischenspiele aus Intermezzo. Detroit Symphony Orchestra, Neeme Järvi / Helge Grünewald

    Index Scriptorium Estoniae

    Grünewald, Helge

    1996-01-01

    Uuest heliplaadist "Schmidt. Sinfonie Nr. 1 E-Dur; Strauss. Vier sinfonische Zwischenspiele aus Intermezzo. Detroit Symphony Orchestra, Neeme Järvi. Chandos/Koch CD 9357 (WD: 68'20") DDD (WD:114'36")

  5. Schmidt. Sinfonie Nr. 1 E-Dur; Strauss. Vier sinfonische Zwischenspiele aus Intermezzo. Detroit Symphony Orchestra, Neeme Järvi / Helge Grünewald

    Index Scriptorium Estoniae

    Grünewald, Helge

    1996-01-01

    Uuest heliplaadist "Schmidt. Sinfonie Nr. 1 E-Dur; Strauss. Vier sinfonische Zwischenspiele aus Intermezzo. Detroit Symphony Orchestra, Neeme Järvi. Chandos/Koch CD 9357 (WD: 68'20") DDD (WD:114'36")

  6. 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.

  7. Decreased levels of pNR1 S897 protein in the cortex of neonatal Sprague Dawley rats with hypoxic-ischemic or NMDA-induced brain damage

    Directory of Open Access Journals (Sweden)

    Ming-Yan Hei

    2012-10-01

    Full Text Available Our objective was to investigate the protein level of phosphorylated N-methyl-D-aspartate (NMDA receptor-1 at serine 897 (pNR1 S897 in both NMDA-induced brain damage and hypoxic-ischemic brain damage (HIBD, and to obtain further evidence that HIBD in the cortex is related to NMDA toxicity due to a change of the pNR1 S897 protein level. At postnatal day 7, male and female Sprague Dawley rats (13.12 ± 0.34 g were randomly divided into normal control, phosphate-buffered saline (PBS cerebral microinjection, HIBD, and NMDA cerebral microinjection groups. Immunofluorescence and Western blot (N = 10 rats per group were used to examine the protein level of pNR1 S897. Immunofluorescence showed that control and PBS groups exhibited significant neuronal cytoplasmic staining for pNR1 S897 in the cortex. Both HIBD and NMDA-induced brain damage markedly decreased pNR1 S897 staining in the ipsilateral cortex, but not in the contralateral cortex. Western blot analysis showed that at 2 and 24 h after HIBD, the protein level of pNR1 S897 was not affected in the contralateral cortex (P > 0.05, whereas it was reduced in the ipsilateral cortex (P 0.05. The levels in the ipsilateral cortex were decreased, but the change was not significant (P > 0.05. The similar reduction in the protein level of pNR1 S897 following both HIBD and NMDA-induced brain damage suggests that HIBD is to some extent related to NMDA toxicity possibly through NR1 phosphorylation of serine 897.

  8. [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).

  9. Mapping of the {alpha}{sub 4} subunit gene (GABRA4) to human chromosome 4 defines an {alpha}{sub 2}-{alpha}{sub 4}-{beta}{sub 1}-{gamma}{sub 1} gene cluster: Further evidence that modern GABA{sub a} receptor gene clusters are derived from an ancestral cluster

    Energy Technology Data Exchange (ETDEWEB)

    McLean, P.J.; Farb, D.H.; Russek, S.J. [Boston Univ. School of Medicine, MA (United States)] [and others

    1995-04-10

    We demonstrated previously that an {alpha}{sub 1}-{beta}{sub 2}-{gamma}{sub 2} gene cluster of the {gamma}-aminobutyric acid (GABA{sub A}) receptor is located on human chromosome 5q34-q35 and that an ancestral {alpha}-{beta}-{gamma} gene cluster probably spawned clusters on chromosomes 4, 5, and 15. Here, we report that the {alpha}{sub 4} gene (GABRA4) maps to human chromosome 4p14-q12, defining a cluster comprising the {alpha}{sub 2}, {alpha}{sub 4}, {beta}{sub 1}, and {gamma}{sub 1} genes. The existence of an {alpha}{sub 2}-{alpha}{sub 4}-{beta}{sub 1}-{gamma}{sub 2} cluster on chromosome 4 and an {alpha}{sub 1}-{alpha}{sub 6}-{beta}{sub 2}-{gamma}{sub 2} cluster on chromosome 5 provides further evidence that the number of ancestral GABA{sub A} receptor subunit genes has been expanded by duplication within an ancestral gene cluster. Moreover, if duplication of the {alpha} gene occurred before duplication of the ancestral gene cluster, then a heretofore undiscovered subtype of a subunit should be located on human chromosome 15q11-q13 within an {alpha}{sub 5}-{alpha}{sub x}-{beta}{sub 3}-{gamma}{sub 3} gene cluster at the locus for Angelman and Prader-Willi syndromes. 34 refs., 6 figs., 1 tab.

  10. 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.

  11. 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.

  12. NR1H4 analysis in patients with progressive familial intrahepatic cholestasis, drug-induced cholestasis or intrahepatic cholestasis of pregnancy unrelated to ATP8B1, ABCB11 and ABCB4 mutations.

    Science.gov (United States)

    Davit-Spraul, Anne; Gonzales, Emmanuel; Jacquemin, Emmanuel

    2012-12-01

    Farnesoid X receptor (FXR, NR1H4) controls bile acid homeostasis. NR1H4 variants may predispose to intrahepatic cholestasis of pregnancy (ICP). We report on NR1H4 analysis in eight patients with progressive familial intrahepatic cholestasis (PFIC) and in eight women with either ICP and/or drug-induced cholestasis (DIC) in whom no disease causing mutation in ATP8B1, ABCB11 and/or ABCB4 were found. No NR1H4 mutation was found in PFIC patients. In one woman with ICP/DIC, a NR1H4 heterozygous variant (c.-1G>T) was found. This suggests that a NR1H4 mutation is not or rarely involved in hepatocellular cholestasis of unknown cause. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

  13. Extensive neuroadaptive changes in cortical gene-transcript expressions of the glutamate system in response to repeated intermittent MDMA administration in adolescent rats

    Directory of Open Access Journals (Sweden)

    Malki Rana

    2008-04-01

    Full Text Available Abstract Background Many studies have focused on the implication of the serotonin and dopamine systems in neuroadaptive responses to the recreational drug 3,4-methylenedioxy-metamphetamine (MDMA. Less attention has been given to the major excitatory neurotransmitter glutamate known to be implicated in schizophrenia and drug addiction. The aim of the present study was to investigate the effect of repeated intermittent MDMA administration upon gene-transcript expression of the glutamate transporters (EAAT1, EAAT2-1, EAAT2-2, the glutamate receptor subunits of AMPA (GluR1, GluR2, GluR3, the glutamate receptor subunits of NMDA (NR1, NR2A and NR2B, as well as metabotropic glutamate receptors (mGluR1, mGluR2, mGluR3, mGluR5 in six different brain regions. Adolescent male Sprague Dawley rats received MDMA at the doses of 3 × 1 and 3 × 5 mg/kg/day, or 3× vehicle 3 hours apart, every 7th day for 4 weeks. The gene-transcript levels were assessed using real-time PCR validated with a range of housekeeping genes. Results The findings showed pronounced enhancements in gene-transcript expression of GluR2, mGluR1, mGluR5, NR1, NR2A, NR2B, EAAT1, and EAAT2-2 in the cortex at bregma +1.6. In the caudate putamen, mRNA levels of GluR3, NR2A, and NR2B receptor subunits were significantly increased. In contrast, the gene-transcript expression of GluR1 was reduced in the hippocampus. In the hypothalamus, there was a significant increase of GluR1, GluR3, mGluR1, and mGluR3 gene-transcript expressions. Conclusion Repeated intermittent MDMA administration induces neuroadaptive changes in gene-transcript expressions of glutamatergic NMDA and AMPA receptor subunits, metabotropic receptors and transporters in regions of the brain regulating reward-related associative learning, cognition, and memory and neuro-endocrine functions.

  14. 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.

  15. Molecular cloning of the cDNA encoding follicle-stimulating hormone beta subunit of the Chinese soft-shell turtle Pelodiscus sinensis, and its gene expression.

    Science.gov (United States)

    Chien, Jung-Tsun; Shen, San-Tai; Lin, Yao-Sung; Yu, John Yuh-Lin

    2005-04-01

    Follicle-stimulating hormone (FSH) is a member of the pituitary glycoprotein hormone family. These hormones are composed of two dissimilar subunits, alpha and beta. Very little information is available regarding the nucleotide and amino acid sequence of FSHbeta in reptilian species. For better understanding of the phylogenetic diversity and evolution of FSH molecule, we have isolated and sequenced the complementary DNA (cDNA) encoding the Chinese soft-shell turtle (Pelodiscus sinensis, Family of Trionychidae) FSHbeta precursor molecule by reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA end (RACE) methods. The cloned Chinese soft-shell turtle FSHbeta cDNA consists of 602-bp nucleotides, including 34-bp nucleotides of the 5'-untranslated region (UTR), 396-bp of the open reading frame, and 3'-UTR of 206-bp nucleotides. It encodes a 131-amino acid precursor molecule of FSHbeta subunit with a signal peptide of 20 amino acids followed by a mature protein of 111 amino acids. Twelve cysteine residues, forming six disulfide bonds within beta-subunit and two putative asparagine-linked glycosylation sites, are also conserved in the Chinese soft-shell turtle FSHbeta subunit. The deduced amino acid sequence of the Chinese soft-shell turtle FSHbeta shares identities of 97% with Reeves's turtle (Family of Bataguridae), 83-89% with birds, 61-70% with mammals, 63-66% with amphibians and 40-58% with fish. By contrast, when comparing the FSHbeta with the beta-subunits of the Chinese soft-shell turtle luteinizing hormone and thyroid stimulating hormone, the homologies are as low as 38 and 39%, respectively. A phylogenetic tree including reptilian species of FSHbeta subunits, is presented for the first time. Out of various tissues examined, FSHbeta mRNA was only expressed in the pituitary gland and can be up-regulated by gonadotropin-releasing hormone in pituitary tissue culture as estimated by fluorescence real-time PCR analysis.

  16. 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

  17. 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.

  18. Effects of sevoflurane on NR1 mRNA of NMDA receptor in rat hippocampal slices during oxygen and glucose deprivation%七氟醚对缺氧无糖损伤大鼠海马 NR1亚基mRNA表达的影响

    Institute of Scientific and Technical Information of China (English)

    张宏金; 王志萍; 于常州

    2005-01-01

    目的探讨七氟醚对缺氧无糖(oxygen and glucose deprivation,OGD)损伤大鼠海马NMDA受体NR1亚基mRNA表达的影响.方法大鼠海马脑片随机分为3组(n=3),用RT-PCR方法检测对照组、缺氧组及七氟醚组大鼠离体海马脑片OGD损伤14 min恢复氧糖供应孵育l、2、4 h后NMDA受体NR1亚基mRNA的表达.结果恢复氧糖供应1、2 h后NR1亚基mRNA的表达3组无明显差异,而缺氧组4 h后NR1亚基mRNA的表达增高,七氟醚组恢复氧糖供应后4 h NR1亚基mRNA的表达明显降低.结论七氟醚可通过下调OGD损伤引起的NR1亚基mRNA的表达发挥脑保护作用.

  19. NMDA-NR1 and AMPA-GluR4 receptor subunit immunoreactivities in the absence epileptic WAG/Rij rat

    NARCIS (Netherlands)

    Bovenkamp-Janssen, M.C. van de; Kloet, J.C. van der; Luijtelaar, G. van; Roubos, E.W.

    2006-01-01

    From an age of 2-3 months onwards, the WAG/Rij rat, a genetic model for absence epilepsy, develops spike-wave discharges (SWD). SWD start in the peri-oral somatosensory cortex (POsc), whereas the rostral reticular thalamic nucleus (rRTN) contributes to synchronizing the thalamo-cortical

  20. 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.

  1. 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

  2. Transcriptome profile analysis of young floral buds of fertile and sterile plants from the self-pollinated offspring of the hybrid between novel restorer line NR1 and Nsa CMS line in Brassica napus

    Directory of Open Access Journals (Sweden)

    Yan Xiaohong

    2013-01-01

    Full Text Available Abstract Background The fertile and sterile plants were derived from the self-pollinated offspring of the F1 hybrid between the novel restorer line NR1 and the Nsa CMS line in Brassica napus. To elucidate gene expression and regulation caused by the A and C subgenomes of B. napus, as well as the alien chromosome and cytoplasm from Sinapis arvensis during the development of young floral buds, we performed a genome-wide high-throughput transcriptomic sequencing for young floral buds of sterile and fertile plants. Results In this study, equal amounts of total RNAs taken from young floral buds of sterile and fertile plants were sequenced using the Illumina/Solexa platform. After filtered out low quality data, a total of 2,760,574 and 2,714,441 clean tags were remained in the two libraries, from which 242,163 (Ste and 253,507 (Fer distinct tags were obtained. All distinct sequencing tags were annotated using all possible CATG+17-nt sequences of the genome and transcriptome of Brassica rapa and those of Brassica oleracea as the reference sequences, respectively. In total, 3231 genes of B. rapa and 3371 genes of B. oleracea were detected with significant differential expression levels. GO and pathway-based analyses were performed to determine and further to understand the biological functions of those differentially expressed genes (DEGs. In addition, there were 1089 specially expressed unknown tags in Fer, which were neither mapped to B. oleracea nor to B. rapa, and these unique tags were presumed to arise basically from the added alien chromosome of S. arvensis. Fifteen genes were randomly selected and their expression levels were confirmed by quantitative RT-PCR, and fourteen of them showed consistent expression patterns with the digital gene expression (DGE data. Conclusions A number of genes were differentially expressed between the young floral buds of sterile and fertile plants. Some of these genes may be candidates for future research on CMS in

  3. 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. Copyright 2009 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  4. PPAR alpha-dependent induction of the energy homeostasis-regulating nuclear receptor NR1i3 (CAR) in rat hepatocytes: Potential role in starvation adaptation

    NARCIS (Netherlands)

    Wieneke, N.; Hirsch-Ernst, K.I.; Kuna, M.; Kersten, A.H.; Pueschel, G.P.

    2007-01-01

    A tight hormonal control of energy homeostasis is of pivotal relevance for animals. Recent evidence suggests an involvement of the nuclear receptor NR1i3 (CAR). Fasting induces CAR by largely unknown mechanisms and CAR-deficient mice are defective in fasting adaptation. In rat hepatocytes CAR was

  5. Sodium channel β subunits: emerging targets in channelopathies.

    Science.gov (United States)

    O'Malley, Heather A; Isom, Lori L

    2015-01-01

    Voltage-gated sodium channels (VGSCs) are responsible for the initiation and propagation of action potentials in excitable cells. VGSCs in mammalian brain are heterotrimeric complexes of α and β subunits. Although β subunits were originally termed auxiliary, we now know that they are multifunctional signaling molecules that play roles in both excitable and nonexcitable cell types and with or without the pore-forming α subunit present. β subunits function in VGSC and potassium channel modulation, cell adhesion, and gene regulation, with particularly important roles in brain development. Mutations in the genes encoding β subunits are linked to a number of diseases, including epilepsy, sudden death syndromes like SUDEP and SIDS, and cardiac arrhythmia. Although VGSC β subunit-specific drugs have not yet been developed, this protein family is an emerging therapeutic target.

  6. 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.

  7. Mutations in mitochondrial-encoded cytochrome c oxidase subunits I, II, and III genes detected in Alzheimer's disease using single-strand conformation polymorphism.

    Science.gov (United States)

    Hamblet, Natasha S; Ragland, Brian; Ali, Mervat; Conyers, Barbara; Castora, Frank J

    2006-02-01

    A "mitochondrial hypothesis" of late onset Alzheimer's disease (AD) has been proposed. Biochemical studies indicate that there is a significant decrease in cytochrome oxidase (CO) activity as well as perturbed CO I and CO III mRNA levels in platelets and brain tissue from Alzheimer's patients. Using the electrophoretic mutation detection technique SSCP and DNA sequencing, we have identified 20 point mutations in the mitochondrial-encoded CO subunits (CO I, II, and III) in AD and age-matched control brain samples. Eight of the mutations are new variants of the mitochondrial genome. The efficiency of SSCP in detecting mutations in the CO subunits was estimated to be 80% when compared to dideoxy sequencing. One of the mutations (at position 9,861) results in a phenylalanine-->leucine substitution at a highly conserved residue in CO III. CO activity was reduced by an average of 35% in all AD brains compared to age-matched control samples, which agrees with previous reports. CO activity in one of the AD brain samples carrying the 9,861 mutation decreased by 80% relative to control brain samples, suggesting that the phenotypic expression of this mutation may result in reduced CO activity and compromised mitochondrial function.

  8. 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.

  9. Impact of rs361072 in the phosphoinositide 3-kinase p110beta gene on whole-body glucose metabolism and subunit protein expression in skeletal muscle

    DEFF Research Database (Denmark)

    Ribel-Madsen, Rasmus; Poulsen, Pernille; Holmkvist, Johan

    2010-01-01

    . The aim was to investigate the influence of rs361072 on in vivo glucose metabolism, skeletal muscle PI3K subunit protein levels, and type 2 diabetes. RESEARCH DESIGN AND METHODS: The functional role of rs361072 was studied in 196 Danish healthy adult twins. Peripheral and hepatic insulin sensitivity...... was assessed by a euglycemic-hyperinsulinemic clamp. Basal and insulin-stimulated biopsies were taken from the vastus lateralis muscle, and tissue p110beta and p85alpha proteins were measured by Western blotting. The genetic association with type 2 diabetes and quantitative metabolic traits was investigated...... in 9,316 Danes with glucose tolerance ranging from normal to overt type 2 diabetes. RESULTS: While hepatic insulin resistance was similar in the fasting state, carriers of the minor G allele had lower hepatic glucose output (per-allele effect: -16%, P(add) = 0.004) during high physiological insulin...

  10. 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.

  11. 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.

  12. 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.

  13. Analysis of NR3A receptor subunits in human native NMDA receptors

    DEFF Research Database (Denmark)

    Nilsson, Anna; Eriksson, Maria; Muly, E Chris

    2007-01-01

    NR3A, representing the third class of NMDA receptor subunits, was first studied in rats, demonstrating ubiquitous expression in the developing central nervous system (CNS), but in the adult mainly expressed in spinal cord and some forebrain nuclei. Subsequent studies showed that rodent and non-human...... primate NR3A expression differs. We have studied the distribution of NR3A in the human CNS and show a widespread distribution of NR3A protein in adult human brain. NR3A mRNA and protein were found in all regions of the cerebral cortex, and also in the subcortical forebrain, midbrain and hindbrain. Only...... very low levels of NR3A mRNA and protein could be detected in homogenized adult human spinal cord, and in situ hybridization showed that expression was limited to ventral motoneurons. We found that NR3A is associated with NR1, NR2A and NR2B in adult human CNS, suggesting the existence of native NR1-NR2...

  14. 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.

  15. [A novel gene (Aa-accA ) encoding acetyl-CoA carboxyltransferase alpha-subunit of Alkalimonas amylolytica N10 enhances salt and alkali tolerance of Escherichia coli and tobacco BY-2 cells].

    Science.gov (United States)

    Xian, Mingjie; Zhai, Lei; Zhong, Naiqin; Ma, Yiwei; Xue, Yanfen; Ma, Yanhe

    2013-08-04

    Acetyl-CoA carboxylase (ACC) catalyzes the first step of fatty acid synthesis. In most bacteria, ACC is composed of four subunits encoded by accA, accB, accC, and accD. Of them, accA encodes acetyl-CoA carboxyltransferase alpha-subunit. Our prior work on proteomics of Alkalimonas amylolytica N10 showed that the expression of the Aa-accA has a remarkable response to salt and alkali stress. This research aimed to find out the Aa-accA gene contributing to salt and alkali tolerance. The Aa-accA was amplified by PCR from A. amylolytica N10 and expressed in E. coli K12 host. The effects of Aa-accA expression on the growth of transgenic strains were examined under different NaCl concentration and pH conditions. Transgenic tobacco BY-2 cells harboring Aa-accA were also generated via Agrobacterium-mediated transformation. The viability of BY-2 cells was determined with FDA staining method after salt and alkali shock. The Aa-accA gene product has 318 amino acids and is homologous to the carboxyl transferase domain of acyl-CoA carboxylases. It showed 76% identity with AccA (acetyl-CoA carboxylase carboxyltransferase subunit alpha) from E. coli. Compared to the wild-type strains, transgenic E. coli K12 strain containing Aa-accA showed remarkable growth superiority when grown in increased NaCl concentrations and pH levels. The final cell density of the transgenic strains was 2.6 and 3.5 times higher than that of the control type when they were cultivated in LB medium containing 6% (W/V) NaCl and at pH 9, respectively. Complementary expression of Aa-accA in an accA-depletion E. coli can recover the tolerance of K12 delta accA to salt and alkali stresses to some extent. Similar to the transgenic E. coli, transgenic tobacco BY-2 cells showed higher percentages of viability compared to the wild BY-2 cells under the salt or alkali stress condition. We found that Aa-accA from A. amylolytica N10 overexpression enhances the tolerance of both transgenic E. coli and tobacco BY-2 cells to

  16. 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.

  17. Amebic colitis in an antigenically and serologically negative patient: usefulness of a small-subunit ribosomal RNA gene-based polymerase chain reaction in diagnosis.

    Science.gov (United States)

    Solaymani-Mohammadi, Shahram; Coyle, Christina M; Factor, Stephen M; Petri, William A

    2008-11-01

    Specific identification of Entamoeba histolytica in clinical specimens is an essential confirmatory diagnostic step in the management of amebiasis. Here, we report an unusual case of amebic colitis in a 20-year-old female immigrant from South China. The patient had experienced diarrhea, crampy abdominal pain, and fever for approximately 3 weeks prior to admission to hospital and had treated herself at home with metronidazole. On admission, stool microscopy and serology for E. histolytica were negative. Because the clinical findings raised the suspicion of Clostridium difficile fulminant colitis, she underwent a subtotal colectomy. Histopathology revealed flask-shaped ulcers characteristic of amebic colitis. Consequently, E. histolytica DNA was detected by a sensitive small-subunit rRNA polymerase chain reaction (PCR) from feces, and the patient was successfully treated for amebiasis with metronidazole. This case exemplifies the relative insensitivity of serologic tests for the diagnosis of intestinal amebiasis and the difficulties encountered in detecting the parasite antigen in a patient partially treated with metronidazole. We conclude that when the possibility of invasive intestinal amebiasis is suspected, detecting the parasite DNA directly in the stool sample by PCR using E. histolytica-specific primers may be an alternative, noninvasive, and reliable tool for the specific diagnosis of the disease.

  18. 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.

  19. 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.

  20. 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.

  1. Hydroxysafflor yellow A improves learning and memory in a rat model of vascular dementia by increasing VEGF and NR1 in the hippocampus.

    Science.gov (United States)

    Zhang, Nan; Xing, Mengya; Wang, Yiyi; Liang, Hao; Yang, Zhuo; Shi, Fudong; Cheng, Yan

    2014-06-01

    Hydroxysafflor yellow A (HSYA) has angiogenesis-regulating and neuro-protective effects, but its effects on vascular dementia (VaD) are unknown. In this study, 30 adult Sprague-Dawley rats were randomly allocated to five groups: normal, sham-operation, VaD alone (bilateral carotid artery occlusion), VaD plus saline (control), and VaD plus HSYA. One week after operation, the HSYA group received one daily tail-vein injection of 0.6 mg/100 g HSYA for two weeks. Five weeks after operation, the spatial memory of all five groups was evaluated by the water maze task, and synaptic plasticity in the hippocampus was assessed by the long-term potentiation (LTP) method. Vascular endothelial growth factor (VEGF) and N-methyl-Daspartic acid receptor 1 (NR1) expression in the hippocampus was detected via Western blot. We found that, compared with the group with VaD alone, the group with HSYA had a reduced escape latency in the water maze (P CA3-CA1 synapses in the hippocampus was enhanced (P < 0.05). Western blot in the late-phase VaD group showed slight up-regulation of VEGF and downregulation of NR1 in the hippocampus, while HSYA significantly up-regulated both VEGF and NR1. These results suggested that HSYA promotes angiogenesis and increases synaptic plasticity, thus improving spatial learning and memory in the rat model of VaD.

  2. Paclitaxel-induced hyperalgesia modulates negative affective component of pain and NR1 receptor expression in the frontal cortex in rats.

    Science.gov (United States)

    Noda, Kazuko; Akita, Hisanao; Ogata, Masanori; Saji, Makoto

    2014-03-01

    Paclitaxel, one of the chemotherapeutic agents clinically used to treat several types of cancer, produces side effects such as peripheral neuropathy, sensory abnormalities, and hyperalgesia. Since hyperalgesia remains after cessation of paclitaxel therapy and becomes chronic, we hypothesize that alteration in memory and the cognitive process of pain underlies hyperalgesia. To test this hypothesis, we examined whether drug-induced hyperalgesia alters the affective component of pain and the NMDA-NR1 and mGluR1 receptors as a mediator for signal transmission and memory of pain. Mechanical sensitivity was measured by von Frey filament test after intraperitoneal injection of paclitaxel in rats. Paclitaxel-induced hyperalgesia was confirmed over almost the entire 14-day period of observation after the treatment. The effect of paclitaxel-induced hyperalgesia on the affective component of pain was assessed using pain-induced place aversion. The formalin-induced conditioned place aversion was completely abolished in the paclitaxel-treated rats. Immunoblot analysis of NR1 and mGluR1 protein levels in various brain regions was performed after paclitaxel treatment. Treatment reduced only the NR1 expression within the frontal cortex. These results suggest that the hypofunction of memory processes with the reduced NMDA receptors in the frontal cortex might be involved in the expression of abnormal emotional behaviors accompanied by hyperalgesia.

  3. 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.

  4. The LIM domain protein FHL2 interacts with the NR5A family of nuclear receptors and CREB to activate the inhibin-α subunit gene in ovarian granulosa cells.

    Science.gov (United States)

    Matulis, Christina K; Mayo, Kelly E

    2012-08-01

    Nuclear receptor transcriptional activity is enhanced by interaction with coactivators. The highly related nuclear receptor 5A (NR5A) subfamily members liver receptor homolog 1 and steroidogenic factor 1 bind to and activate several of the same genes, many of which are important for reproductive function. To better understand transcriptional activation by these nuclear receptors, we sought to identify interacting proteins that might function as coactivators. The LIM domain protein four and a half LIM domain 2 (FHL2) was identified as interacting with the NR5A receptors in a yeast two-hybrid screen of a human ovary cDNA library. FHL2, and the closely related FHL1, are both expressed in the rodent ovary and in granulosa cells. Small interfering RNA-mediated knockdown of FHL1 and FHL2 in primary mouse granulosa cells reduced expression of the NR5A target genes encoding inhibin-α and P450scc. In vitro assays confirmed the interaction between the FHL and NR5A proteins and revealed that a single LIM domain of FHL2 is sufficient for this interaction, whereas determinants in both the ligand binding domain and DNA binding domain of NR5A proteins are important. FHL2 enhances the ability of both liver receptor homolog 1 and steroidogenic factor 1 to activate the inhibin-α subunit gene promoter in granulosa cells and thus functions as a transcriptional coactivator. FHL2 also interacts with cAMP response element-binding protein and substantially augments activation of inhibin gene expression by the combination of NR5A receptors and forskolin, suggesting that FHL2 may facilitate integration of these two signals. Collectively these results identify FHL2 as a novel coactivator of NR5A nuclear receptors in ovarian granulosa cells and suggest its involvement in regulating target genes important for mammalian reproduction.

  5. Reconsideration of systematic relationships within the order Euplotida (Protista, Ciliophora) using new sequences of the gene coding for small-subunit rRNA and testing the use of combined data sets to construct phylogenies of the Diophrys-complex.

    Science.gov (United States)

    Yi, Zhenzhen; Song, Weibo; Clamp, John C; Chen, Zigui; Gao, Shan; Zhang, Qianqian

    2009-03-01

    Comprehensive molecular analyses of phylogenetic relationships within euplotid ciliates are relatively rare, and the relationships among some families remain questionable. We performed phylogenetic analyses of the order Euplotida based on new sequences of the gene coding for small-subunit RNA (SSrRNA) from a variety of taxa across the entire order as well as sequences from some of these taxa of other genes (ITS1-5.8S-ITS2 region and histone H4) that have not been included in previous analyses. Phylogenetic trees based on SSrRNA gene sequences constructed with four different methods had a consistent branching pattern that included the following features: (1) the "typical" euplotids comprised a paraphyletic assemblage composed of two divergent clades (family Uronychiidae and families Euplotidae-Certesiidae-Aspidiscidae-Gastrocirrhidae), (2) in the family Uronychiidae, the genera Uronychia and Paradiophrys formed a clearly outlined, well-supported clade that seemed to be rather divergent from Diophrys and Diophryopsis, suggesting that the Diophrys-complex may have had a longer and more separate evolutionary history than previously supposed, (3) inclusion of 12 new SSrRNA sequences in analyses of Euplotidae revealed two new clades of species within the family and cast additional doubt on the present classification of genera within the family, and (4) the intraspecific divergence among five species of Aspidisca was far greater than those of closely related genera. The ITS1-5.8S-ITS2 coding regions and partial histone H4 genes of six morphospecies in the Diophrys-complex were sequenced along with their SSrRNA genes and used to compare phylogenies constructed from single data sets to those constructed from combined sets. Results indicated that combined analyses could be used to construct more reliable, less ambiguous phylogenies of complex groups like the order Euplotida, because they provide a greater amount and diversity of information.

  6. 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.

  7. 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.

  8. 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.

  9. Prebiotic feeding elevates central brain derived neurotrophic factor, N-methyl-D-aspartate receptor subunits and D-serine.

    Science.gov (United States)

    Savignac, Helene M; Corona, Giulia; Mills, Henrietta; Chen, Li; Spencer, Jeremy P E; Tzortzis, George; Burnet, Philip W J

    2013-12-01

    The influence of the gut microbiota on brain chemistry has been convincingly demonstrated in rodents. In the absence of gut bacteria, the central expression of brain derived neurotropic factor, (BDNF), and N-methyl-d-aspartate receptor (NMDAR) subunits are reduced, whereas, oral probiotics increase brain BDNF, and impart significant anxiolytic effects. We tested whether prebiotic compounds, which increase intrinsic enteric microbiota, also affected brain BDNF and NMDARs. In addition, we examined whether plasma from prebiotic treated rats released BDNF from human SH-SY5Y neuroblastoma cells, to provide an initial indication of mechanism of action. Rats were gavaged with fructo-oligosaccharides (FOS), galacto-oligosaccharides (GOS) or water for five weeks, prior to measurements of brain BDNF, NMDAR subunits and amino acids associated with glutamate neurotransmission (glutamate, glutamine, and serine and alanine enantiomers). Prebiotics increased hippocampal BDNF and NR1 subunit expression relative to controls. The intake of GOS also increased hippocampal NR2A subunits, and frontal cortex NR1 and d-serine. Prebiotics did not alter glutamate, glutamine, l-serine, l-alanine or d-alanine concentrations in the brain, though GOSfeeding raised plasma d-alanine. Elevated levels of plasma peptide YY (PYY) after GOS intake was observed. Plasma from GOS rats increased the release of BDNF from SH-SY5Y cells, but not in the presence of PYY antisera. The addition of synthetic PYY to SH-SY5Y cell cultures, also elevated BDNF secretion. We conclude that prebiotic-mediated proliferation of gut microbiota in rats, like probiotics, increases brain BDNF expression, possibly through the involvement of gut hormones. The effect of GOS on components of central NMDAR signalling was greater than FOS, and may reflect the proliferative potency of GOS on microbiota. Our data therefore, provide a sound basis to further investigate the utility of prebiotics in the maintenance of brain health and

  10. 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

  11. 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.

  12. 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....

  13. 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

  14. Diversity of heterotrimeric G-protein γ subunits in plants

    Directory of Open Access Journals (Sweden)

    Trusov Yuri

    2012-10-01

    Full Text Available Abstract Background Heterotrimeric G-proteins, consisting of three subunits Gα, Gβ and Gγ are present in most eukaryotes and mediate signaling in numerous biological processes. In plants, Gγ subunits were shown to provide functional selectivity to G-proteins. Three unconventional Gγ subunits were recently reported in Arabidopsis, rice and soybean but no structural analysis has been reported so far. Their relationship with conventional Gγ subunits and taxonomical distribution has not been yet demonstrated. Results After an extensive similarity search through plant genomes, transcriptomes and proteomes we assembled over 200 non-redundant proteins related to the known Gγ subunits. Structural analysis of these sequences revealed that most of them lack the obligatory C-terminal prenylation motif (CaaX. According to their C-terminal structures we classified the plant Gγ subunits into three distinct types. Type A consists of Gγ subunits with a putative prenylation motif. Type B subunits lack a prenylation motif and do not have any cysteine residues in the C-terminal region, while type C subunits contain an extended C-terminal domain highly enriched with cysteines. Comparative analysis of C-terminal domains of the proteins, intron-exon arrangement of the corresponding genes and phylogenetic studies suggested a common origin of all plant Gγ subunits. Conclusion Phylogenetic analyses suggest that types C and B most probably originated independently from type A ancestors. We speculate on a potential mechanism used by those Gγ subunits lacking isoprenylation motifs to anchor the Gβγ dimer to the plasma membrane and propose a new flexible nomenclature for plant Gγ subunits. Finally, in the light of our new classification, we give a word of caution about the interpretation of Gγ research in Arabidopsis and its generalization to other plant species.

  15. The site specific demethylation in the 5'-regulatory area of NMDA receptor 2B subunit gene associated with CIE-induced up-regulation of transcription.

    Directory of Open Access Journals (Sweden)

    Mei Qiang

    Full Text Available BACKGROUND: The NMDA receptor represents a particularly important site of ethanol action in the CNS. We recently reported that NMDA receptor 2B (NR2B gene expression was persistently up-regulated following chronic intermittent ethanol (CIE treatment. Increasing evidence that epigenetic mechanisms are involved in dynamic and long-lasting regulation of gene expression in multiple neuroadaptive processes prompted us to investigate the role of DNA methylation in mediating CIE-induced up-regulation of NR2B gene transcription. To dissect the changes of DNA methylation in the NR2B gene, we have screened a large number of CpG sites within its 5'-regulatory area following CIE treatment. METHODS: Primary cortical cultured neurons were subjected to ethanol treatment in a CIE paradigm. Bisulfite conversion followed by pyrosequencing was used for quantitative measurement and analysis of CpG methylation status within the 5'-regulatory area of the NR2B gene; chromatin immunoprecipitation (ChIP assay was used to examine DNA levels associated with methylation and transcription factor binding. Electrophoretic mobility shift assay (EMSA and in vitro DNA methylation assays were performed to determine the direct impact of DNA methylation on the interaction between DNA and transcription factor and promoter activity. RESULTS: Analysis of individual CpG methylation sites within the NR2B 5'regulatory area revealed three regions with clusters of site-specific CpG demethylation following CIE treatment and withdrawal. This was confirmed by ChIP showing similar decreases of methylated DNA in the same regions. The CIE-induced demethylation is characterized by being located near certain transcription factor binding sequences, AP-1 and CRE, and occurred during treatment as well as after ethanol withdrawal. Furthermore, the increase in vitro of methylated DNA decreased transcription factor binding activity and promoter activity. An additional ChIP assay indicated that the CIE

  16. 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.

  17. Gene structure and mutations of glutaryl-coenzyme A dehydrogenase: Impaired association of enzyme subunits that is due to an A421V substitution causes glutaric acidemia type I in the Amish

    Energy Technology Data Exchange (ETDEWEB)

    Biery, B.J.; Stein, D.E.; Goodman, S.I. [Univ. of Colorado School of Medicine, Denver, CO (United States)] [and others

    1996-11-01

    The structure of the human glutaryl coenzyme A dehydrogenase (GCD) gene was determined to contain 11 exons and to span {approximately}7 kb. Fibroblast DNA from 64 unrelated glutaric academia type I (GA1) patients was screened for mutations by PCR amplification and analysis of SSCP. Fragments with altered electrophoretic mobility were subcloned and sequenced to detect mutations that caused GA1. This report describes the structure of the GCD gene, as well as point mutations and polymorphisms found in 7 of its 11 exons. Several mutations were found in more than one patient, but no one prevalent mutation was detected in the general population. As expected from pedigree analysis, a single mutant allele causes GA1 in the Old Order Amish of Lancaster County, Pennsylvania. Several mutations have been expressed in Escherichia coli, and all produce diminished enzyme activity. Reduced activity in GCD encoded by the A421V mutation in the Amish may be due to impaired association of enzyme subunits. 13 refs., 5 figs., 3 tabs.

  18. The human [gamma]-aminobutyric acid receptor subunit [beta]3 and [alpha]5 gene cluster in chromosome 15q11-q13 is rich in highly polymorphic (CA)[sub n] repeats

    Energy Technology Data Exchange (ETDEWEB)

    Glatt, K.; Lalande, M. (Howard Hughes Medical Institute, Boston, MA (United States)); Sinnett, D. (Harvard Medical School, Boston, MA (United States))

    1994-01-01

    The [gamma]-aminobutyric acid (GABA[sub A]) receptor [beta]33 (GABRB3) and [alpha]5 (GABRA5) subunit genes have been localized to the Angelman and Prader-Willi syndrome region of chromosome 15q11-q13. GABRB3, which encompasses 250 kb, is located 100 kb proximal of GABRA5, with the two genes arranged in head-to-head transcriptional orientation. In screening 135 kb of cloned DNA within a 260-kb interval extending from within GABRB3 to the 5[prime] end of GABRA5, 10 new (CA), repeats have been identified. Five of these have been analyzed in detail and found to be highly polymorphic, with the polymorphism information content (PIC) ranging from 0.7 to 0.85 and with heterozygosities of 67 to 94%. In the clones from GABRB3/GABRA5 region, therefore, the frequency of (CA)[sub n] with PICs [ge] 0.7 is 1 per 27 kb. Previous estimates of the density of (CA)[sub n] with PICs [ge] 0.7 in the human genome have been approximately 10-fold lower. The GABRB3/GABRA5 region appears, therefore, to be enriched for highly informative (CA)[sub n]. This set of closely spaced, short tandem repeat polymorphisms will be useful in the molecular analyses of Prader-Willi and Angelman syndromes and in high-resolution studies of genetic recombination within this region. 21 refs., 2 figs., 1 tab.

  19. 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.

  20. Association of sequence variants in CKM (creatine kinase, muscle) and COX4I2 (cytochrome c oxidase, subunit 4, isoform 2) genes with racing performance in Thoroughbred horses.

    Science.gov (United States)

    Gu, J; MacHugh, D E; McGivney, B A; Park, S D E; Katz, L M; Hill, E W

    2010-11-01

    The wild progenitors of the domestic horse were subject to natural selection for speed and stamina for millennia. Uniquely, this process has been augmented in Thoroughbreds, which have undergone at least 3 centuries of intense artificial selection for athletic phenotypes. While the phenotypic adaptations to exercise are well described, only a small number of the underlying genetic variants contributing to these phenotypes have been reported. A panel of candidate performance-related genes was examined for DNA sequence variation in Thoroughbreds and the association with racecourse performance investigated. Eighteen candidate genes were chosen for their putative roles in exercise. Re-sequencing in Thoroughbred samples was successful for primer sets in 13 of these genes. SNPs identified in this study and from the EquCab2.0 SNP database were genotyped in 2 sets of Thoroughbred samples (n = 150 and 148) and a series of population-based case-control investigations were performed by separating the samples into discrete cohorts on the basis of retrospective racecourse performance. Twenty novel SNPs were detected in 3 genes: ACTN3, CKM and COX4I2. Genotype frequency distributions for 3 SNPs in CKM and COX4I2 were significantly (P race. These associations were not validated when an additional (n = 130) independent set of samples was genotyped, but when analyses included all samples (n = 278) the significance of association at COX4I2 g.22684390C > T was confirmed (P horse industries, it is vital that rigour is applied to studies generating these data and that adequate and appropriate sample sets, particularly for independent replication, are used. © 2010 EVJ Ltd.

  1. 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.

  2. Heterozygous deletion of the LRFN2 gene is associated with working memory deficits.

    Science.gov (United States)

    Thevenon, Julien; Souchay, Céline; Seabold, Gail K; Dygai-Cochet, Inna; Callier, Patrick; Gay, Sébastien; Corbin, Lucie; Duplomb, Laurence; Thauvin-Robinet, Christel; Masurel-Paulet, Alice; El Chehadeh, Salima; Avila, Magali; Minot, Delphine; Guedj, Eric; Chancenotte, Sophie; Bonnet, Marlène; Lehalle, Daphne; Wang, Ya-Xian; Kuentz, Paul; Huet, Frédéric; Mosca-Boidron, Anne-Laure; Marle, Nathalie; Petralia, Ronald S; Faivre, Laurence

    2016-06-01

    Learning disabilities (LDs) are a clinically and genetically heterogeneous group of diseases. Array-CGH and high-throughput sequencing have dramatically expanded the number of genes implicated in isolated intellectual disabilities and LDs, highlighting the implication of neuron-specific post-mitotic transcription factors and synaptic proteins as candidate genes. We report a unique family diagnosed with autosomal dominant learning disability and a 6p21 microdeletion segregating in three patients. The 870 kb microdeletion encompassed the brain-expressed gene LRFN2, which encodes for a synaptic cell adhesion molecule. Neuropsychological assessment identified selective working memory deficits, with borderline intellectual functioning. Further investigations identified a defect in executive function, and auditory-verbal processes. These data were consistent with brain MRI and FDG-PET functional brain imaging, which, when compared with controls, revealed abnormal brain volume and hypometabolism of gray matter structures implicated in working memory. We performed electron microscopy immunogold labeling demonstrating the localization of LRFN2 at synapses of cerebellar and hippocampal rat neurons, often associated with the NR1 subunit of N-methyl-D-aspartate receptors (NMDARs). Altogether, the combined approaches imply a role for LRFN2 in LD, specifically for working memory processes and executive function. In conclusion, the identification of familial cases of clinically homogeneous endophenotypes of LD might help in both the management of patients and genetic counseling for families.

  3. Changes in expression of NR-1 and GluR-2 in the trigeminal motor nucleus of rats after psychological stress%心理应激后三叉神经运动核神经元两种谷氨酸受体亚单位表达变化的实验研究

    Institute of Scientific and Technical Information of China (English)

    逯宜; 胡波; 朱永进

    2012-01-01

    目的 观察心理应激对大鼠三叉神经运动核(trigeminal motor nucleus,Mo5)内神经元谷氨酸受体亚单位N-甲基-D-天门冬氨酸受体-1型(N-methyl-D-aspartate receptor-1,NR-1)和α-氨基-3-羧基-5-甲基-4-异恶唑丙酸受体-2型(glutamic acid receptor-2,GluR-2)表达的影响,探讨心理应激在夜磨牙症中可能的作用机制.方法 选取健康雄性SD大鼠80只,按体质量分层后根据随机数字表随机分为心理应激组和空白对照组(各40只),采用荧光免疫组化和反转录聚合酶链反应检测心理应激1、4、8和12 d大鼠Mo5内NR-1和GluR-2的表达差异.结果 8和12d时心理应激组大鼠Mo5内GluR-2免疫荧光吸光度值(0.152 ±0.056、0.125±0.076)显著低于空白对照组(0.352±0.104、0.342 ±0.100)(P<0.05),其mRNA水平(0.566±0.101、0.384±0.097)也显著低于空白对照组(0.913±0.095、0.892 ±0.099) (P <0.05);而NR-1的免疫荧光吸光度值及mRNA水平可见增高趋势,但与空白对照组差异无统计学意义(P>0.05).结论 心理应激可导致大鼠Mo5内GluR-2表达发生改变,这一改变可能通过支配颌面部肌肉的运动神经元兴奋性的变化,进而影响口颌功能.%Objective To explore the underlying neurobiological mechanism of psychological stress involved in bruxism,and to investigate the changes of N-methyl-D-aspartate receptor-1 (NR-1) and glutamic acid receptor-2 (GluR-2) subunits in the trigeminal motor nucleus (Mo5) of rats after psychological stress.Methods Healthy male SD rats were selected and randomly divided into 2 groups according to body weight:psychological stress (PS) group and control (Con) group.The expression of NR-1 and GIuR-2 subunits in Mo5 were detected by reverse transcription polymerase chain reaction (RT-PCR) and immunofluorescence assay at 1,4,8 and 12 day during PS.Results At 8 and 12 day of PS,the average optical density of GluR-2 immunofluorescence in the Mo5 of PS rats (0.152 ±0.056,0.125 ±0.076) were

  4. 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

  5. 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.

  6. 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.

  7. Nordslesvigeren er nr. 1

    DEFF Research Database (Denmark)

    Christensen, Steffen Lind

    2015-01-01

    Historien om de nordslesvigske krigsdeltagere under 1. Verdenskrig er traditionelt blevet behandlet i et nationalt perspektiv. Her benævnes soldaterne oftest som ’danske’. Denne artikel undersøger, hvordan nordslesvigske soldater på Østfronten selv udtrykte deres identitet i krigssituationen....... Udgangspunktet er krigsdeltagernes beskrivelser af og syn på dels de fællesskaber, de var en del af ved fronten, og dels de grupperinger, de anså for deres modsætninger. Hovedpointen er, at det mest italesatte tilhørsforhold blandt disse soldater var det regionale bånd til Nordslesvig....

  8. Enhanced thyroid hormone breakdown in hepatocytes by mutual induction of the constitutive androstane receptor (CAR, NR1I3) and arylhydrocarbon receptor by benzo[a]pyrene and phenobarbital.

    Science.gov (United States)

    Schraplau, Anne; Schewe, Bettina; Neuschäfer-Rube, Frank; Ringel, Sebastian; Neuber, Corinna; Kleuser, Burkhard; Püschel, Gerhard P

    2015-02-03

    Xenobiotics may interfere with the hypothalamic-pituitary-thyroid endocrine axis by inducing enzymes that inactivate thyroid hormones and thereby reduce the metabolic rate. This induction results from an activation of xeno-sensing nuclear receptors. The current study shows that benzo[a]pyrene, a frequent contaminant of processed food and activator of the arylhydrocarbon receptor (AhR) activated the promoter and induced the transcription of the nuclear receptor constitutive androstane receptor (CAR, NR1I3) in rat hepatocytes. Likewise, phenobarbital induced the AhR transcription. This mutual induction of the nuclear receptors enhanced the phenobarbital-dependent induction of the prototypic CAR target gene Cyp2b1 as well as the AhR-dependent induction of UDP-glucuronosyltransferases. In both cases, the induction by the combination of both xenobiotics was more than the sum of the induction by either substance alone. By inducing the AhR, phenobarbital enhanced the benzo[a]pyrene-dependent reduction of thyroid hormone half-life and the benzo[a]pyrene-dependent increase in the rate of thyroid hormone glucuronide formation in hepatocyte cultures. CAR ligands might thus augment the endocrine disrupting potential of AhR activators by an induction of the AhR. Copyright © 2014. Published by Elsevier Ireland Ltd.

  9. Loss of the flagellum happened only once in the fungal lineage: phylogenetic structure of Kingdom Fungi inferred from RNA polymerase II subunit genes

    Directory of Open Access Journals (Sweden)

    Hodson Matthew C

    2006-09-01

    Full Text Available Abstract Background At present, there is not a widely accepted consensus view regarding the phylogenetic structure of kingdom Fungi although two major phyla, Ascomycota and Basidiomycota, are clearly delineated. Regarding the lower fungi, Zygomycota and Chytridiomycota, a variety of proposals have been advanced. Microsporidia may or may not be fungi; the Glomales (vesicular-arbuscular mycorrhizal fungi may or may not constitute a fifth fungal phylum, and the loss of the flagellum may have occurred either once or multiple times during fungal evolution. All of these issues are capable of being resolved by a molecular phylogenetic analysis which achieves strong statistical support for major branches. To date, no fungal phylogeny based upon molecular characters has satisfied this criterion. Results Using the translated amino acid sequences of the RPB1 and RPB2 genes, we have inferred a fungal phylogeny that consists largely of well-supported monophyletic phyla. Our major results, each with significant statistical support, are: (1 Microsporidia are sister to kingdom Fungi and are not members of Zygomycota; that is, Microsporidia and fungi originated from a common ancestor. (2 Chytridiomycota, the only fungal phylum having a developmental stage with a flagellum, is paraphyletic and is the basal lineage. (3 Zygomycota is monophyletic based upon sampling of Trichomycetes, Zygomycetes, and Glomales. (4 Zygomycota, Basidiomycota, and Ascomycota form a monophyletic group separate from Chytridiomycota. (5 Basidiomycota and Ascomycota are monophyletic sister groups. Conclusion In general, this paper highlights the evolutionary position and significance of the lower fungi (Zygomycota and Chytridiomycota. Our results suggest that loss of the flagellum happened only once during early stages of fungal evolution; consequently, the majority of fungi, unlike plants and animals, are nonflagellated. The phylogeny we infer from gene sequences is the first one that is

  10. Loss of the flagellum happened only once in the fungal lineage: phylogenetic structure of kingdom Fungi inferred from RNA polymerase II subunit genes.

    Science.gov (United States)

    Liu, Yajuan J; Hodson, Matthew C; Hall, Benjamin D

    2006-09-29

    At present, there is not a widely accepted consensus view regarding the phylogenetic structure of kingdom Fungi although two major phyla, Ascomycota and Basidiomycota, are clearly delineated. Regarding the lower fungi, Zygomycota and Chytridiomycota, a variety of proposals have been advanced. Microsporidia may or may not be fungi; the Glomales (vesicular-arbuscular mycorrhizal fungi) may or may not constitute a fifth fungal phylum, and the loss of the flagellum may have occurred either once or multiple times during fungal evolution. All of these issues are capable of being resolved by a molecular phylogenetic analysis which achieves strong statistical support for major branches. To date, no fungal phylogeny based upon molecular characters has satisfied this criterion. Using the translated amino acid sequences of the RPB1 and RPB2 genes, we have inferred a fungal phylogeny that consists largely of well-supported monophyletic phyla. Our major results, each with significant statistical support, are: (1) Microsporidia are sister to kingdom Fungi and are not members of Zygomycota; that is, Microsporidia and fungi originated from a common ancestor. (2) Chytridiomycota, the only fungal phylum having a developmental stage with a flagellum, is paraphyletic and is the basal lineage. (3) Zygomycota is monophyletic based upon sampling of Trichomycetes, Zygomycetes, and Glomales. (4) Zygomycota, Basidiomycota, and Ascomycota form a monophyletic group separate from Chytridiomycota. (5) Basidiomycota and Ascomycota are monophyletic sister groups. In general, this paper highlights the evolutionary position and significance of the lower fungi (Zygomycota and Chytridiomycota). Our results suggest that loss of the flagellum happened only once during early stages of fungal evolution; consequently, the majority of fungi, unlike plants and animals, are nonflagellated. The phylogeny we infer from gene sequences is the first one that is congruent with the widely accepted morphology

  11. 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.

  12. 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.

  13. 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.

  14. 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.

  15. 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.

  16. Description of Eurystomatella sinica n. gen., n. sp., with establishment of a new family Eurystomatellidae n. fam. (Protista, Ciliophora, Scuticociliatia) and analyses of its phylogeny inferred from sequences of the small-subunit rRNA gene.

    Science.gov (United States)

    Miao, Miao; Wang, Yangang; Song, Weibo; Clamp, John C; Al-Rasheid, Khaled A S

    2010-02-01

    Recently, an undescribed marine ciliate was isolated from China. Investigation of its morphology and infraciliature revealed it as an undescribed species representing a new genus, Eurystomatella n. gen., the type of the new family Eurystomatellidae n. fam. The new family is defined by close-set, apically positioned oral membranelles and a dominant buccal field that is surrounded by an almost completely circular paroral membrane. The new genus is defined by having a small oral membranelle 1 (M1), bipartite M2 and well-developed M3, a body surface faintly sculptured with a silverline system in a quadrangular, reticulate pattern and a cytostome located at the anterior third of a large buccal field. The type species of the new genus, Eurystomatella sinica n. sp., is a morphologically unique form that is defined mainly by the combination of a conspicuously flattened body, several caudal cilia, extremely long cilia associated with the buccal apparatus and a contractile vacuole located subcaudally. According to phylogenetic analyses of small-subunit (SSU) rRNA gene sequences, Eurystomatella clusters with the genus Cyclidium, as a sister group to the family Pleuronematidae. The great divergence in both buccal and somatic ciliature between Eurystomatella and all other known scuticociliates supports the establishment of a new family for Eurystomatella.

  17. 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.

  18. 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.

  19. 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.

  20. 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.

  1. Caryotricha minuta (Xu et al., 2008) nov. comb., a unique marine ciliate (Protista, Ciliophora, Spirotrichea), with phylogenetic analysis of the ambiguous genus Caryotricha inferred from the small-subunit rRNA gene sequence.

    Science.gov (United States)

    Miao, Miao; Shao, Chen; Jiang, Jiamei; Li, Liqiong; Stoeck, Thorsten; Song, Weibo

    2009-02-01

    A population of Kiitricha minuta Xu et al., 2008, a small kiitrichid ciliate, was isolated from a brackish water sample in Jiaozhou Bay, Qingdao, northern China. After comparison of its morphology and infraciliature, it is believed that this morphotype should be assigned to the genus Caryotricha; hence, a new combination is suggested, Caryotricha minuta (Xu et al., 2008) nov. comb. The small-subunit (SSU) rRNA gene sequence was determined in order to elucidate the phylogenetic position of this poorly known, ambiguous genus. The organism can be clearly separated from its congener, Caryotricha convexa Kahl, 1932, by the extremely shortened ventral cirral rows in the posterior ends. Based on the data available, an improved diagnosis is given for the genus: marine Kiitrichidae with prominent buccal field; two highly developed undulating membranes; non-grouped, uniform cirral rows on both ventral and dorsal sides; enlarged transverse cirri present, which are the only differentiated cirri; marginal cirri not present; one short migratory row located posterior to buccal field; structure of dorsal kineties generally in Kiitricha pattern. The sequence of the SSU rRNA gene of C. minuta differs by 13 % from that of Kiitricha marina. Molecular phylogenetic analyses (Bayesian inference, least squares, neighbour joining, maximum parsimony) indicate that Caryotricha, together with Kiitricha, diverges at a deep level from all other spirotrichs. Its branching position is between Phacodiniidia and Licnophoridia. The results strongly support the distinct separation of the Kiitricha-Caryotricha clade, which always branches basal to the Stichotrichia-Hypotrichia-Oligotrichia-Choreotrichia assemblage. These results also confirm the previous hypothesis that the Kiitricha-Caryotricha group, long assumed to be a close relation to the euplotids, represents a taxon at subclass level within the spirotrichs.

  2. 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.

  3. 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

  4. Comparison of sequencing the D2 region of the large subunit ribosomal RNA gene (MicroSEQ®) versus the internal transcribed spacer (ITS) regions using two public databases for identification of common and uncommon clinically relevant fungal species.

    Science.gov (United States)

    Arbefeville, S; Harris, A; Ferrieri, P

    2017-09-01

    Fungal infections cause considerable morbidity and mortality in immunocompromised patients. Rapid and accurate identification of fungi is essential to guide accurately targeted antifungal therapy. With the advent of molecular methods, clinical laboratories can use new technologies to supplement traditional phenotypic identification of fungi. The aims of the study were to evaluate the sole commercially available MicroSEQ® D2 LSU rDNA Fungal Identification Kit compared to the in-house developed internal transcribed spacer (ITS) regions assay in identifying moulds, using two well-known online public databases to analyze sequenced data. 85 common and uncommon clinically relevant fungi isolated from clinical specimens were sequenced for the D2 region of the large subunit (LSU) of ribosomal RNA (rRNA) gene with the MicroSEQ® Kit and the ITS regions with the in house developed assay. The generated sequenced data were analyzed with the online GenBank and MycoBank public databases. The D2 region of the LSU rRNA gene identified 89.4% or 92.9% of the 85 isolates to the genus level and the full ITS region (f-ITS) 96.5% or 100%, using GenBank or MycoBank, respectively, when compared to the consensus ID. When comparing species-level designations to the consensus ID, D2 region of the LSU rRNA gene aligned with 44.7% (38/85) or 52.9% (45/85) of these isolates in GenBank or MycoBank, respectively. By comparison, f-ITS possessed greater specificity, followed by ITS1, then ITS2 regions using GenBank or MycoBank. Using GenBank or MycoBank, D2 region of the LSU rRNA gene outperformed phenotypic based ID at the genus level. Comparing rates of ID between D2 region of the LSU rRNA gene and the ITS regions in GenBank or MycoBank at the species level against the consensus ID, f-ITS and ITS2 exceeded performance of the D2 region of the LSU rRNA gene, but ITS1 had similar performance to the D2 region of the LSU rRNA gene using MycoBank. Our results indicated that the MicroSEQ® D2 LSU r

  5. PKA regulatory subunit expression in tooth development.

    Science.gov (United States)

    de Sousa, Sílvia Ferreira; Kawasaki, Katsushige; Kawasaki, Maiko; Volponi, Ana Angelova; Gomez, Ricardo Santiago; Gomes, Carolina Cavaliéri; Sharpe, Paul T; Ohazama, Atsushi

    2014-05-01

    Protein kinase A (PKA) plays critical roles in many biological processes including cell proliferation, cell differentiation, cellular metabolism and gene regulation. Mutation in PKA regulatory subunit, PRKAR1A has previously been identified in odontogenic myxomas, but it is unclear whether PKA is involved in tooth development. The aim of the present study was to assess the expression of alpha isoforms of PKA regulatory subunit (Prkar1a and Prkar2a) in mouse and human odontogenesis by in situ hybridization. PRKAR1A and PRKAR2A mRNA transcription was further confirmed in a human deciduous germ by qRT-PCR. Mouse Prkar1a and human PRKAR2A exhibited a dynamic spatio-temporal expression in tooth development, whereas neither human PRKAR1A nor mouse Prkar2a showed their expression in odontogenesis. These isoforms thus showed different expression pattern between human and mouse tooth germs. Copyright © 2014 Elsevier B.V. All rights reserved.

  6. Regulation of BK channels by auxiliary γ subunits

    Directory of Open Access Journals (Sweden)

    Jiyuan eZhang

    2014-10-01

    Full Text Available The large-conductance, calcium- and voltage-activated potassium (BK channel has the largest single-channel conductance among potassium channels and can be activated by both membrane depolarization and increases in intracellular calcium concentration. BK channels consist of pore-forming, voltage- and calcium-sensing α subunits, either alone or in association with regulatory subunits. BK channels are widely expressed in various tissues and cells including both excitable and non-excitable cells and display diverse biophysical and pharmacological characteristics. This diversity can be explained in part by posttranslational modifications and alternative splicing of the α subunit, which is encoded by a single gene, KCNMA1, as well as by tissue-specific β subunit modulation. Recently, a leucine-rich repeat-containing membrane protein, LRRC26, was found to interact with BK channels and cause an unprecedented large negative shift (~-140 mV in the voltage dependence of the BK channel activation. LRRC26 allows BK channels to open even at near-physiological calcium concentration and membrane voltage in non-excitable cells. Three LRRC26-related proteins, LRRC52, LRRC55, and LRRC38, were subsequently identified as BK channel modulators. These LRRC proteins are structurally and functionally distinct from the BK channel β subunits and were designated as γ subunits. The discovery of the γ subunits adds a new dimension to BK channel regulation and improves our understanding of the physiological functions of BK channels in various tissues and cell types. Unlike BK channel β subunits, which have been intensively investigated both mechanistically and physiologically, our understanding of the γ subunits is very limited at this stage. This article reviews the structure, modulatory mechanisms, physiological relevance, and potential therapeutic implications of γ subunits as they are currently understood.

  7. Long-term changes in cyclosporine pharmacokinetics after renal transplantation in children: evidence for saturable presystemic metabolism and effect of NR1I2 polymorphism.

    Science.gov (United States)

    Fanta, Samuel; Jönsson, Siv; Karlsson, Mats O; Niemi, Mikko; Holmberg, Christer; Hoppu, Kalle; Backman, Janne T

    2010-05-01

    To improve cyclosporine dose individualization, the authors carried out a comprehensive analysis of the effects of clinical and genetic factors on cyclosporine pharmacokinetics in 176 children before and up to 16 years after renal transplantation. Pretransplantation test doses of cyclosporine were given intravenously and orally, followed by blood sampling for 24 hours. After transplantation, cyclosporine was quantified at trough, 2 hours postdose, or with dose-interval curves. A 3-compartment population pharmacokinetic model was used to describe the data. Cyclosporine oral bioavailability increased more than 1.5-fold in the first month after transplantation, returning thereafter gradually to its initial value in 1 to 1.5 years. Moreover, older children receiving cyclosporine twice daily as the gelatin capsule microemulsion formulation had an about 1.25 to 1.3 times higher bioavailability than did the younger children receiving the liquid formulation thrice daily. In 91 children with genetic data after transplantation, patients carrying the NR1I2 g.-25385C-g.-24381A-g.-205_-200GAGAAG-g.7635G-g.8055C haplotype had about one-tenth lower bioavailability, per allele, than did noncarriers (P = .039). The significance of the NR1I2 genotype warrants further study. In conclusion, by accounting for the effects of developmental factors (body weight), time after transplantation, and cyclosporine dosing frequency/formulation, it may be possible to improve individualization of cyclosporine dosing in children.

  8. 夏枯草RuBPCase大亚基基因的克隆与表达%Molecular Cloning and Expression Analysis of A Large Subunit Gene of RuBPCase ( PvrbcL ) from Prunella vulgaris

    Institute of Scientific and Technical Information of China (English)

    许锋; 蒋丽阳; 曹腾; 宁迎晶; 程水源

    2012-01-01

    In order to clone the large subunit gene (rbcL) of Rubisco-1,5-bisphosphate carboxylase/ oxylase (RuBPCase), one of the important enzyme involved in photosynthesis and photorespiration, from P. vulgaris, and so as to provide basic data for studying on the structure and function of RuBPCase and discussing the photosynthesis mechanism on the molecular level of P. vulgaris, a pair of specific primers was designed. An rbcL gene fragment, named PvrbcL., was cloned from P. vulgaris using genomic DNA as template. The results showed that the length of PvrbcL sequence was 837 bp, encoding a 278-amino-acid protein. The GenBank accession No. of PvrbcL gene was JN692563. The BLAST results showed that the homology of the PvrbcL nucleotide sequence with rbcL genes from Prunella grandiflora , Salvia pachyphylla , Nicotiana tabacum , Glycine max , Oryza sativa , Phaseolus vulgaris , Vitis vinifer, and Brassica napus was 86%~100%, and the homology of amino acid sequences was 92//o~ 100%. Northern blot analysis revealed that PvrbcL gene expressed in leaves and stems, and the highest level was detected in leaves. Developmental expression patter indicated that the expression level of PvrbcL gene was increased along with the growth of the leaves. The isolation and expression of PvrbcL gene could provide the basis for further studying the structure and function of RuBPCase in P. vulgaris.%为从夏枯草中克隆参与光合作用和光呼吸代谢的关键酶RuBPcase大亚基基因rbcL,为今后研究夏枯草RuBPcase的结构和功能以及从分子水平探讨夏枯草的光合作用机理提供必要的基础数据,以夏枯草基因组DNA为模板,采用PCR法对夏枯草RuBPCase大亚基基因(PvrbcL)的克隆与表达进行研究.结果表明:PCR法扩增出夏枯草RuBPcase大亚基基因片段PvrbcL,基因长837bp,编码278个氨基酸,GenBank登录号为JN692563;经BLAST序列比较,夏枯草PvrbcL基因核苷酸序列与大花夏枯草、鼠尾草、烟草、大豆、水稻

  9. Early expression of GABA(A) receptor delta subunit in the neonatal rat hippocampus.

    Science.gov (United States)

    Didelon, F; Mladinic', M; Cherubini, E; Bradbury, A

    2000-12-01

    The cDNA library screening strategy was used to identify the genes encoding for GABA(A) receptor subunits in the rat hippocampus during development. With this technique, genes encoding eleven GABA(A) receptor subunits were identified. The alpha5 subunit was by far the most highly expressed, followed by the gamma2, alpha2 and alpha4 subunits respectively. The expression of the beta2, alpha1, gamma1, beta1 and beta3 subunits was moderate, although that of the alpha3 and delta subunits was weak. In situ hybridization experiments, using digoxigenin-labeled cRNA probes, confirmed that the delta subunit was expressed in the neonatal as well as in the adult hippocampus, and is likely to form functional receptors in association with other subunits of the GABA(A) receptor. When the more sensitive RT-PCR approach was used, the gamma3 subunit was also detected, suggesting that this subunit is present in the hippocampus during development but at low levels of expression. The insertion of the delta subunit into functional GABA(A) receptors may enhance the efficacy of GABA in the immediate postnatal period when this amino acid is still exerting a depolarizing and excitatory action.

  10. Cloning, Expression and Sequence Analysis of the Subunit Gene Atp9 Unit in Trametes Gallica%粗毛栓菌atp9基因的克隆、表达及序列分析

    Institute of Scientific and Technical Information of China (English)

    雍彬; 司文杰; 陶宗娅; 严伟

    2011-01-01

    ATP合酶是生物体内能量代谢的关键酶,参与多种氧化磷酸化和光合磷酸化反应.atp9基因是ATP合酶的重要组成部分,其编码了ATP合酶A亚基上第9亚单位,与呼吸作用和光合作用密切相关.本研究利用atP9基因在进化过程中高度保守的特点,据已知近缘真菌基因序列,设计并合成了一对引物,以粗毛栓菌mRNA反转录得到的cDNA第一链为模板,PCR扩增得到atp9基因完整序列,并连接于原核表达载体pET32a(+)上.测序与序列分析表明:该克隆片段全长222 bp,共编码73个氨基酸,翻译的蛋白质分子量是7.35 kDa.转化大肠杆菌后经IPTG诱导,可高效表达外源融合蛋白,分子量大小与预测结果一致.经过同源比对和进化树分析,该克隆基因编码的氨基酸序列与可可丛枝病菌(Crinipellis perniciosa)和瓣环栓菌(Trametes cingulata strain ATCC 26747)中相对应的氨基酸序列相似度最高.本实验为未来进一步研究粗毛栓菌atp9基因和其蛋白功能,阐明其调控和作用机制奠定了基础.%ATP synthase is a key enzyme of energy metabolism, which was involved in a variety of oxidative phosphorylation and photophosphorylation in vivo. The atp9 gene encoding the ninth part of ATP synthase A sub-unit is an important component of ATP synthase. It is closely related with respiration and photosynthesis. For its highly conserved sequence in evolution, one pair of primers were designed and synthesized according to the known gene sequences from closely related fungi. The first strand of Trametes gallica Cdna was amplified by reverse transcription and then the complete atp9 gene sequences were obtained by PCR. And then the product of PCR was ligated to the prokaryotic expression vector pET32a ( + ) and transformed into E. Coll for the expression of fusion protein. The sequenced and bioinformatics analysis showed that: the complete length of atp9 gene was 222 bp, and the peptide it encoded had 73 amino acids with

  11. Morphology and small-subunit rRNA gene sequences of two novel marine ciliates, Metanophrys orientalis spec. nov. and Uronemella sinensis spec. nov. (Protista, Ciliophora, Scuticociliatia), with an improved diagnosis of the genus Uronemella.

    Science.gov (United States)

    Pan, Xuming; Zhu, Mingzhuang; Ma, Honggang; Al-Rasheid, Khaled A S; Hu, Xiaozhong

    2013-09-01

    The morphology and infraciliature of two novel marine scuticociliates, Metanophrys orientalis spec. nov. and Uronemella sinensis spec. nov., collected from sandy beaches at Qingdao, China, were investigated using live observation and protargol-staining methods. Metanophrys orientalis spec. nov. is distinguished by the following characteristics: marine habitat and a slender to elongate oval body with pointed anterior end and rounded caudal end, in vivo about 25-50 µm long; buccal field about a quarter to a third of body length; nine or ten somatic kineties with dikinetids approximately in anterior half of body, monokinetids in posterior half; membranelles 1 and 2 almost equal in length and composed of two and three longitudinal rows of kinetids respectively; paroral membrane with zigzag structure extending anteriorly to middle portion of membranelle 2; contractile vacuole pore located at posterior end of somatic kinety 1. The genus Uronemella is redefined as follows: marine form with an elongate-elliptical or inverted pear-shaped body; apical plate conspicuous; buccal field about two-thirds of body length, cytostome subequatorially located; oral apparatus Uronema-like; somatic kineties comprising a mixture of dikinetids and monokinetids. Uronemella sinensis spec. nov. is recognized by having an elongate-elliptical body with truncated apical frontal plate, size in vivo about 25-35 × 15-20 µm, nine or ten somatic kineties, membranelle 1 consisting of two or three basal bodies, contractile vacuole pore at posterior end of somatic kinety 1. This study also compared the small-subunit rRNA gene sequences of these two species with other closely related species to show the sequence divergence, which ranged from 3.53 to 9.60%. Phylogenetic analyses support the contention that the genus Uronemella is monophyletic, while Metanophrys is non-monophyletic.

  12. 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.

    Science.gov (United States)

    Zelinka, Tomás; Timmers, Henri J L M; Kozupa, Anna; Chen, Clara C; Carrasquillo, Jorge A; Reynolds, James C; Ling, Alexander; Eisenhofer, Graeme; Lazúrová, Ivica; Adams, Karen T; Whatley, Millie A; Widimsky, Jirí; Pacak, Karel

    2008-03-01

    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%), 39% with liver metastases (SDHB +/-: 27% vs 47%), and 32% with lung metastases (SDHB +/-: 37% vs 29%). The most common sites of bone involvement were thoracic spine (80%; SDHB+/-: 83% vs 77%), lumbar spine (78%; SDHB +/-: 78% vs 75%), and pelvic and sacral bones (78%; SDHB +/-: 91% vs 65%, P=0.04). Subjects with SDHB mutation also showed significantly higher involvement of long bones (SDHB +/-: 78% vs 30%, P=0.007) than those without the mutation. The best overall sensitivity in detecting bone metastases demonstrated positron emission tomography (PET) with 6-[(18)F]-fluorodopamine ([(18)F]-FDA; 90%), followed by bone scintigraphy (82%), computed tomography or magnetic resonance imaging (CT/MRI; 78%), 2-[(18)F]-fluoro-2-deoxy-d-glucose ([(18)F]-FDG) PET (76%), and scintigraphy with [(123/131)I]-metaiodobenzylguanidine (71%). In subjects with SDHB mutation, imaging modalities with best sensitivities for detecting bone metastases were CT/MRI (96%), bone scintigraphy (95%), and [(18)F]-FDG PET (92%). In subjects without SDHB mutations, the modality with the best sensitivity for bone metastases was [(18)F]-FDA PET (100%). In conclusion, bone scintigraphy should be used in the staging of patients with malignant pheochromocytoma and paraganglioma, particularly in patients with SDHB mutations. As for PET imaging, [(18)F]-FDG PET is highly recommended in SDHB mutation patients, whereas [(18)F]-FDA PET is recommended in patients without the mutation.

  13. Molecular identification of sibling species of Sclerodermus (Hymenoptera: Bethylidae that parasitize buprestid and cerambycid beetles by using partial sequences of mitochondrial DNA cytochrome oxidase subunit 1 and 28S ribosomal RNA gene.

    Directory of Open Access Journals (Sweden)

    Yuan Jiang

    Full Text Available The species belonging to Sclerodermus (Hymenoptera: Bethylidae are currently the most important insect natural enemies of wood borer pests, mainly buprestid and cerambycid beetles, in China. However, some sibling species of this genus are very difficult to distinguish because of their similar morphological features. To address this issue, we conducted phylogenetic and genetic analyses of cytochrome oxidase subunit I (COI and 28S RNA gene sequences from eight species of Sclerodermus reared from different wood borer pests. The eight sibling species were as follows: S. guani Xiao et Wu, S. sichuanensis Xiao, S. pupariae Yang et Yao, and Sclerodermus spp. (Nos. 1-5. A 594-bp fragment of COI and 750-bp fragment of 28S were subsequently sequenced. For COI, the G-C content was found to be low in all the species, averaging to about 30.0%. Sequence divergences (Kimura-2-parameter distances between congeneric species averaged to 4.5%, and intraspecific divergences averaged to about 0.09%. Further, the maximum sequence divergences between congeneric species and Sclerodermus sp. (No. 5 averaged to about 16.5%. All 136 samples analyzed were included in six reciprocally monophyletic clades in the COI neighbor-joining (NJ tree. The NJ tree inferred from the 28S rRNA sequence yielded almost identical results, but the samples from S. guani, S. sichuanensis, S. pupariae, and Sclerodermus spp. (Nos. 1-4 clustered together and only Sclerodermus sp. (No. 5 clustered separately. Our findings indicate that the standard barcode region of COI can be efficiently used to distinguish morphologically similar Sclerodermus species. Further, we speculate that Sclerodermus sp. (No. 5 might be a new species of Sclerodermus.

  14. Phylogenetic relationships in the mushroom genus Coprinus and dark-spored allies based on sequence data from the nuclear gene coding for the large ribosomal subunit RNA: divergent domains, outgroups, and monophyly.

    Science.gov (United States)

    Hopple, J S; Vilgalys, R

    1999-10-01

    Phylogenetic relationships were investigated in the mushroom genus Coprinus based on sequence data from the nuclear encoded large-subunit rDNA gene. Forty-seven species of Coprinus and 19 additional species from the families Coprinaceae, Strophariaceae, Bolbitiaceae, Agaricaceae, Podaxaceae, and Montagneaceae were studied. A total of 1360 sites was sequenced across seven divergent domains and intervening sequences. A total of 302 phylogenetically informative characters was found. Ninety-eight percent of the average divergence between taxa was located within the divergent domains, with domains D2 and D8 being most divergent and domains D7 and D10 the least divergent. An empirical test of phylogenetic signal among divergent domains also showed that domains D2 and D3 had the lowest levels of homoplasy. Two equally most parsimonious trees were resolved using Wagner parsimony. A character-state weighted analysis produced 12 equally most parsimonious trees similar to those generated by Wagner parsimony. Phylogenetic analyses employing topological constraints suggest that none of the major taxonomic systems proposed for subgeneric classification is able to completely reflect phylogenetic relationships in Coprinus. A strict consensus integration of the two Wagner trees demonstrates the problematic nature of choosing outgroups within dark-spored mushrooms. The genus Coprinus is found to be polyphyletic and is separated into three distinct clades. Most Coprinus taxa belong to the first two clades, which together form a larger monophyletic group with Lacrymaria and Psathyrella in basal positions. A third clade contains members of Coprinus section Comati as well as the genus Leucocoprinus, Podaxis pistillaris, Montagnea arenaria, and Agaricus pocillator. This third clade is separated from the other species of Coprinus by members of the families Strophariaceae and Bolbitiaceae and the genus Panaeolus.

  15. NADH dehydrogenase subunit 1 and cytochrome c oxidase subunit I sequences compared for members of the genus Taenia (Cestoda).

    Science.gov (United States)

    Gasser, R B; Zhu, X; McManus, D P

    1999-12-01

    Nine members of the genus Taenia (Taenia taeniaeformis, Taenia hydatigena, Taenia pisiformis, Taenia ovis, Taenia multiceps, Taenia serialis, Taenia saginata, Taenia solium and the Asian Taenia) were characterised by their mitochondrial NADH dehydrogenase subunit 1 gene sequences and their genetic relationships were compared with those derived from the cytochrome c oxidase subunit 1 sequence data. The extent of inter-taxon sequence difference in NADH dehydrogenase subunit 1 (approximately 5.9-30.8%) was usually greater than in cytochrome c oxidase subunit 1 (approximately 2.5-18%). Although topology of the phenograms derived from NADH dehydrogenase subunit 1 and cytochrome c oxidase subunit 1 sequence data differed, there was concordance in that T. multiceps, T. serialis (of canids), T. saginata and the Asian Taenia (of humans) were genetically most similar, and those four members were genetically more similar to T. ovis and T. solium than they were to T. hydatigena and T. pisiformis (of canids) or T. taeniaeformis (of cats). The NADH dehydrogenase subunit 1 sequence data may prove useful in studies of the systematics and population genetic structure of the Taeniidae.

  16. A Study on Campylobacter jejuni and Campylobacter coli through Commercial Broiler Production Chains in Thailand: Antimicrobial Resistance, the Characterization of DNA Gyrase Subunit A Mutation, and Genetic Diversity by Flagellin A Gene Restriction Fragment Length Polymorphism.

    Science.gov (United States)

    Thomrongsuwannakij, Thotsapol; Blackall, Patrick J; Chansiripornchai, Niwat

    2017-06-01

    Contaminated poultry meat is regarded as the main source of human campylobacteriosis. During September 2014 and February 2015, breeder flocks, hatcheries, and broiler farms from two chicken production chains were investigated chronologically. Five commercial breeder flocks (Breeder Flocks 1-5), two hatcheries (Hatcheries A and B), and five broiler flocks (Broiler Flocks 1-5) were sampled in this study. Campylobacter colonization of both breeder and broiler flocks was determined from cloacal swabs and environmental samples (pan feeders, footwear, darkling beetles, flies, feed, and water). The eggs from the breeder flocks were followed to hatcheries. At the hatcheries, early embryonic deaths, egg trays, eggshells, hatchers, and water were investigated. Cloacal swabs were taken from broilers at Days 1, 14, and 28 (all broiler flocks), and either 35 (Broiler Flocks 1 and 2) or 43 (Broiler Flocks 3-5). Thirty-six Campylobacter jejuni and 94 Campylobacter coli isolates collected through two broiler production chains were tested by twofold agar dilution for their susceptibility to antimicrobial agents. Most Campylobacter isolates were multidrug resistant (MDR), defined as being resistant to three or more antimicrobial classes ( C. jejuni : 100%; C. coli : 98.9%), and exhibited high resistance to enrofloxacin ( C. jejuni : 100%; C. coli : 98.9%). The vast majority of C. coli were resistant to tetracycline (97.9%), trimethoprim-sulfamethoxazole (81.9%), and doxycycline (79.8%), but only 55.6%, 36.1%, and 50% of C. jejuni isolates revealed resistance to these antimicrobial agents, respectively. A selected subset of 24 C. jejuni and 24 C. coli were characterized for their mutations in the quinolone resistance determining region of the DNA gyrase subunit A gene by nucleotide sequence analysis. The Thr-86-Ile substitution (ACA-ATA in C. jejuni or ACT-ATT in C. coli ) was found in all isolates. Moreover, a total of 130 Campylobacter isolates were typed with the use of polymerase

  17. NR2 subunits and NMDA receptors on lamina II inhibitory and excitatory interneurons of the mouse dorsal horn

    Directory of Open Access Journals (Sweden)

    MacDermott Amy B

    2010-05-01

    Full Text Available Abstract Background NMDA receptors expressed by spinal cord neurons in the superficial dorsal horn are involved in the development of chronic pain associated with inflammation and nerve injury. The superficial dorsal horn has a complex and still poorly understood circuitry that is mainly populated by inhibitory and excitatory interneurons. Little is known about how NMDA receptor subunit composition, and therefore pharmacology and voltage dependence, varies with neuronal cell type. NMDA receptors are typically composed of two NR1 subunits and two of four NR2 subunits, NR2A-2D. We took advantage of the differences in Mg2+ sensitivity of the NMDA receptor subtypes together with subtype preferring antagonists to identify the NR2 subunit composition of NMDA receptors expressed on lamina II inhibitory and excitatory interneurons. To distinguish between excitatory and inhibitory interneurons, we used transgenic mice expressing enhanced green fluorescent protein driven by the GAD67 promoter. Results Analysis of conductance ratio and selective antagonists showed that lamina II GABAergic interneurons express both the NR2A/B containing Mg2+ sensitive receptors and the NR2C/D containing NMDA receptors with less Mg2+ sensitivity. In contrast, excitatory lamina II interneurons express primarily NR2A/B containing receptors. Despite this clear difference in NMDA receptor subunit expression in the two neuronal populations, focally stimulated synaptic input is mediated exclusively by NR2A and 2B containing receptors in both neuronal populations. Conclusions Stronger expression of NMDA receptors with NR2C/D subunits by inhibitory interneurons compared to excitatory interneurons may provide a mechanism to selectively increase activity of inhibitory neurons during intense excitatory drive that can provide inhibitory feedback.

  18. Voltage-gated calcium channel subunits from platyhelminths: Potential role in praziquantel action✩

    Science.gov (United States)

    Jeziorski, Michael C.; Greenberg, Robert M.

    2013-01-01

    Voltage-gated calcium (Ca2+) channels provide the pathway for Ca2+ influxes that underlie Ca2+-dependent responses in muscles, nerves and other excitable cells. They are also targets of a wide variety of drugs and toxins. Ca2+ channels are multisubunit protein complexes consisting of a pore-forming α1 subunit and other modulatory subunits, including the β subunit. Here, we review the structure and function of schistosome Ca2+ channel subunits, with particular emphasis on variant Ca2+ channel β subunits (Cavβvar) found in these parasites. In particular, we examine the role these β subunits may play in the action of praziquantel, the current drug of choice against schistosomiasis. We also present evidence that Cavβvar homologs are found in other praziquantel-sensitive platyhelminths such as the pork tapeworm, Taenia solium, and that these variant β subunits may thus represent a platyhelminth-specific gene family. PMID:16545816

  19. Eduard Tubina "Kogutud teoste" köidetest I/I (sümfooniad nr. 1 ja 2) ning I/VII (orkestrisüidid) / Timo Virtanen ; (tõlkinud Merike Vaitmaa)

    Index Scriptorium Estoniae

    Virtanen, Timo, 1956-

    2013-01-01

    Arvustus: Lauri Sirp, Toomas Trass (toim.). Kogutud teosed. I seeria, I köide : Sümfoonia nr. 1 ; Sümfoonia nr. 2 : Legendaarne / Eduard Tubin. Stockholm : Rahvusvaheline Eduard Tubina Ühing ; Gehrmans Musikförlag, 2012

  20. Stenhammer, Wilhelm. sinfonien Nr. 1 F-Dur und Nr. 2 g-Moll op. 34, Excelsior! op. 13, Serenade F-Dur op. 31. Sinfonieorchester Göteborg, Neeme Järvi / Andreas K. W. Meyer

    Index Scriptorium Estoniae

    Meyer, Andreas K. W.

    1995-01-01

    Uuest heliplaadist "Stenhammer, Wilhelm. sinfonien Nr. 1 F-Dur und Nr. 2 g-Moll op. 34, Excelsior! op. 13, Serenade F-Dur op. 31. Sinfonieorchester Göteborg, Neeme Järvi". DG 2 CD 445857-2 (WD: 138'37")

  1. Stenhammer, Wilhelm. sinfonien Nr. 1 F-Dur und Nr. 2 g-Moll op. 34, Excelsior! op. 13, Serenade F-Dur op. 31. Sinfonieorchester Göteborg, Neeme Järvi / Andreas K. W. Meyer

    Index Scriptorium Estoniae

    Meyer, Andreas K. W.

    1995-01-01

    Uuest heliplaadist "Stenhammer, Wilhelm. sinfonien Nr. 1 F-Dur und Nr. 2 g-Moll op. 34, Excelsior! op. 13, Serenade F-Dur op. 31. Sinfonieorchester Göteborg, Neeme Järvi". DG 2 CD 445857-2 (WD: 138'37")

  2. Eduard Tubina "Kogutud teoste" köidetest I/I (sümfooniad nr. 1 ja 2) ning I/VII (orkestrisüidid) / Timo Virtanen ; (tõlkinud Merike Vaitmaa)

    Index Scriptorium Estoniae

    Virtanen, Timo, 1956-

    2013-01-01

    Arvustus: Lauri Sirp, Toomas Trass (toim.). Kogutud teosed. I seeria, I köide : Sümfoonia nr. 1 ; Sümfoonia nr. 2 : Legendaarne / Eduard Tubin. Stockholm : Rahvusvaheline Eduard Tubina Ühing ; Gehrmans Musikförlag, 2012

  3. Subunit structure of the acetylcholine receptor from Electrophorus electricus.

    Science.gov (United States)

    Conti-Tronconi, B M; Hunkapiller, M W; Lindstrom, J M; Raftery, M A

    1982-11-01

    The amino-terminal amino acid sequences of the four major peptides (Mr 41,000, 50,000, 55,000, and 62,000) present in purified preparations of Electrophorus electricus nicotinic acetylcholine receptor (AcChoR) have been determined for 24 cycles by automated sequence analysis procedures yielding four unique polypeptide sequences. The sequences showed a high degree of similarity, having identical residues in a number of positions ranging between 37% and 50% for specific pairs of subunits. Comparison of the sequences obtained with those of the subunits of similar molecular weight from Torpedo californica AcChoR revealed an even higher degree of homology (from 46% to 71%) for these two highly diverged species. Simultaneous sequence analysis of the amino termini present in native, purified Electrophorus AcChoR showed that these four related sequences were the only ones present and that they occur in a ratio of 2:1:1:1, with the smallest subunit ("alpha 1") being present in two copies. Genealogical analysis suggests that the subunits of both Torpedo and Electrophorus AcChoRs derive from a common ancestral gene, the divergence having occurred early in the evolution of the receptor. This shared ancestry and the very early divergence of the four subunits, as well as the highly conserved structure of the AcChoR complex along animal evolution, suggest that each of the subunits evolved to perform discrete crucial roles in the physiological function of the AcChoR.

  4. Expression of accessory colonization factor subunit A (ACFA) of Vibrio cholerae and ACFA fused to cholera toxin B subunit in transgenic tomato (Solanum lycopersicum).

    Science.gov (United States)

    Sharma, Manoj Kumar; Jani, Dewal; Thungapathra, M; Gautam, J K; Meena, L S; Singh, Yogendra; Ghosh, Amit; Tyagi, Akhilesh Kumar; Sharma, Arun Kumar

    2008-05-20

    In earlier study from our group, cholera toxin B subunit had been expressed in tomato for developing a plant-based vaccine against cholera. In the present investigation, gene for accessory colonization factor (acf) subunit A, earlier reported to be essential for efficient colonization in the intestine, has been expressed in Escherichia coli as well as tomato plants. Gene encoding for a chimeric protein having a fusion of cholera toxin B subunit and accessory colonization factor A was also expressed in tomato to generate more potent combinatorial antigen. CaMV35S promoter with a duplicated enhancer sequence was used for expression of these genes in tomato. Integration of transgenes into tomato genome was confirmed by PCR and Southern hybridization. Expression of the genes was confirmed at transcript and protein levels. Accessory colonization factor A and cholera toxin B subunit fused to this protein accumulated up to 0.25% and 0.08% of total soluble protein, respectively, in the fruits of transgenic plants. Whereas protein purified from E. coli, in combination with cholera toxin B subunit can be used for development of conventional subunit vaccine, tomato fruits expressing these proteins can be used together with tomato plants expressing cholera toxin B subunit for development of oral vaccine against cholera.

  5. Global gene expression profiling of brown to white adipose tissue transformation in sheep reveals novel transcriptional components linked to adipose remodeling

    DEFF Research Database (Denmark)

    Basse, Astrid L.; Dixen, Karen; Yadav, Rachita;

    2015-01-01

    NR1H3, MYC, KLF4, ESR1, RELA and BCL6, which were linked to the overall changes in gene expression during the adipose tissue remodeling. Finally, the perirenal adipose tissue expressed both brown and brite/beige adipocyte marker genes at birth, the expression of which changed substantially over time...

  6. 编码酶复合体Ⅰ亚单位的线粒体基因新突变导致的MELAS综合征%Novel mutations in the mitochondrial DNA encoded complex Ⅰ subunit genes associated with MELAS

    Institute of Scientific and Technical Information of China (English)

    赵丹华; 王朝霞; 李务荣; 洪道俊; 郑日亮; 孙永安; 张巍; 袁云

    2011-01-01

    目的:报道4例由编码酶复合体Ⅰ中NADH脱氢酶(ND)亚单位的线粒体基因(mtDNA)突变所导致的线粒体脑肌病患者,分析其临床及骨骼肌病理改变特点.方法:4例患者的发病年龄在6 ~21岁之间,病程在7~ 20年.其中1例为MELAS、3例为MELAS/Leigh叠加综合征.对4例患者进行肌肉活检和mtDNA全长测序检查.结果:骨骼肌病理检查发现1例同时存在破碎红纤维(RRFs)及SDH深染的血管(SSVs),2例仅有SSVs,另1例未见异常.4例患者均携带mtDNA编码的ND基因突变,分别为位于ND3编码区的T10191C(p.S45P)、ND4编码区的A11470C(p.K237N)、ND5编码区的T13046C(p.M237T)点突变以及累及ND5和ND6编码区的单一大片段缺失( 13025-13033:14417-14425),后3种突变均为新发现的致病性突变.结论:ND基因突变是导致部分MELAS或MELAS/Leigh叠加综合征患者的分子病理学基础,这些患者的骨骼肌病理检查常缺乏典型的线粒体脑肌病的病理改变,如RRFs.%Objective To report the clinical and myopathological features of 4 patients with mitochondrial encepha-lomyopathy associated with mutations in mitochondrial DNA (mtDNA) encoded NADH dehydrogenase (ND) subunit genes of complex I. Methods The onset age of 4 patients ranged from 6 to 21 years,with a clinical course from 7 to 20 years. A-mong them, 1 case was consistent with MELAS and 3 cases with MELAS/Leigh overlap syndrome. Muscle biopsy and whole sequencing of mtDNA were performed on these patients. Results Skeletal muscle biopsy disclosed both ragged-red fibers ( RRFs) and strongly succinate dehydrogenase-reactive vessels (SSVs) in one case,only SSVs without RRFs in two cases, and no abnormality in one. Whole sequencing of mtDNA revealed T10191C( p. S45P) in ND3 , A11470C( p. K237N) in ND4,T13046C( p. M237T) in ND5 and a single large-scale deletion ranging from 13025-13033 to 14417-14425 encompassing ND5 and ND6 in these patients respectively. Among them,Al 1470C,T13046C and the

  7. 姜黄素对糖尿病神经病理性痛大鼠脊髓背角NR2B与NR1活性的影响%Effects of curcumin on activity of NR2B and NR1 in spinal dorsal horn in a rat model of diabetic neuropathic pain

    Institute of Scientific and Technical Information of China (English)

    李佳佳; 马益梅; 连庆泉; 李军; 曹红

    2014-01-01

    Objective To evaluate the effects of curcumin on the activity of NR2B and NR1 in the spinal dorsal horn in a rat model of diabetic neuropathic pain (DNP).Methods Diabetes mellitus was induced by high-sucrose and high-fat diet and intraperitoneal streptozotocin 35 mg/kg,then confirmed by fasting blood glucose level ≥ 16.7 mmol/L 3 days later in male Sprague-Dawley rats.DNP was confirmed by the mechanical paw withdrawal threshold (MWT) and thermal paw withdrawal latency (TWL) measured on day 14 after streptozotocin administration < 80% of the baseline value.The rats were then randomly divided into 3 groups (n =27 each) using a random number table:DNP,DNP+ curcumin group (group DCur)and DNP + solvent control group (group DSC).Curcumin 100 mg· kg-1 · d-1 and corn oil 4 ml· kg-1 · d-1 were injected intraperitoneally for 14 consecutive days starting from 14 days after administration of streptozotocin in DCur and DSC groups,respectively.Another 27 normal male Sprague-Dawley rats served as control group (group C) and were fed with normal forage.At 3,7 and 14 days after curcumin injection,MWT and TWL were measured and the lumbar segments (L4-6) of the spinal cord were removed.The expression of pTyr1472-NR2B and pSer896-NR1 in the spinal dorsal horn was determined by immunohistochemistry and Western blot.Results Compared with group C,MWT was significantly decreased,TWL was shortened,and the expression of pTyr1472-NR2B was up-regulated at each time point in group DNP.Compared with group DNP,MWT was significantly increased,and TWL was prolonged at 7 days after curcumin injection,and the expression of pTyr1472-NR2B was down-regulated at 3 days after curcumin injection in group DCur.There was no significant difference in each parameter between DNP and DSC groups,and in the expression of pSer896-NR1 between the four groups.Conclusion The mechanism by which curcumin mitigates neuropathic pain in type 2 diabetic rats may be related to inhibition of up-regulation of p

  8. Dravet综合征的临床特点分析及SCN1A基因新突变%Clinical Analysis of Dravet Syndrome and Novel Gene Mutation of Voltage-Gated Sodium Channel α1-Su-bunit

    Institute of Scientific and Technical Information of China (English)

    王新华; 周水珍

    2011-01-01

    Objective To study the clinical characteristics of Dravet syndrome and to screen the voltage -gated sodium channel αl -subunit( SCNI A ) of a newly diagnosed child, hoping to find the gene mutation. Methods The clinical information of 3 Dravet syndrome children were collected,the blood sample of a new diagnostic child was provided. DNA was extracted from peripheral blood leukocytes using relax gene blood DNA system. The total 26 exons of SCN1A were amplified by polymerase chain reaction( PCR), and the PCR products were screened by Denaturing high performance liquid chromatography, then the abnormal fragments were sequenced by Sanger method in order to find the mutations of SCN 1A gene. Results 1. The common manifestations of 3 Dravet syndrome cases: onset during the first year of life; in all children, the seizures were associated with febrile seizures and they changed to afebrile seizures after 1 year; the forms of seizures included clonus,myoclonus and atypical absence;the seizures were difficult to control with anti -epileptic drugs; all children presented some degree of psychomotor development delay; there were sharp - slow waves, spike - slow waves and multi spike - slow waves in EEG of diapause. 2. A missense mutation of SCNI A gene (c. 2867T > G, M956R) was found in the Dravet syndrome child, which had not been reported up to Nov.2010. Conclusions Dravet syndrome is an epileptic encephalopathy with a bad prognosis,and it needs to be differentiate it frome febrile seizures. The missense mutation of SCNIA gene supports the relationship of SCNIA mutation and Dravet syndrom.%目的 分析Dravet综合征的临床特点,并对新诊断患儿进行SCN1A基因筛查,寻找基因突变.方法收集3例Dravet综合征患儿临床资料,留取例1患儿血样标本,提取外周血白细胞基因组DNA,对SCN1A全部外显子进行PCR扩增,通过变性高效液相色谱法对PCR产物进行突变片段筛查,对于变性高效液相色谱法筛查有异常的片段

  9. Problems connected with the use of oligonucleotide probes with a high degree of degeneracy. Identification of mRNA and of cDNA clones corresponding to the gene of the. cap alpha. -subunit of Na/sup +/, K/sup +/-ATPase

    Energy Technology Data Exchange (ETDEWEB)

    Petrukhin, K.E.; Grishin, A.V.; Arsenyan, S.G.; Broude, N.E.; Grinkevich, V.A.; Filippova, L.Yu.; Severtsova, I.V.; Modyanov, N.N.

    1986-10-01

    To identify and search for nucleotide sequences containing the structural part of the gene of the ..cap alpha..-subunit of Na/sup +/, K/sup +/-ATPase, 17-membered oligonucleotide probes corresponding to the peptide Lys-Asp-Ala-Phe-Gln-Asn have been synthesized. It has been shown that, with a 64-fold degeneracyd, the 17-membered probe is suitable only for the identification of a specific sequence in mRNA. To search for clones containing cDNA fragments, preliminary fractionation of the probes with the aid of HPLC or the resynthesis of groups of oligonucleotides with a lower degeneracy is necessary.

  10. Accessory subunits are integral for assembly and function of human mitochondrial complex I.

    Science.gov (United States)

    Stroud, David A; Surgenor, Elliot E; Formosa, Luke E; Reljic, Boris; Frazier, Ann E; Dibley, Marris G; Osellame, Laura D; Stait, Tegan; Beilharz, Traude H; Thorburn, David R; Salim, Agus; Ryan, Michael T

    2016-10-06

    Complex I (NADH:ubiquinone oxidoreductase) is the first enzyme of the mitochondrial respiratory chain and is composed of 45 subunits in humans, making it one of the largest known multi-subunit membrane protein complexes. Complex I exists in supercomplex forms with respiratory chain complexes III and IV, which are together required for the generation of a transmembrane proton gradient used for the synthesis of ATP. Complex I is also a major source of damaging reactive oxygen species and its dysfunction is associated with mitochondrial disease, Parkinson's disease and ageing. Bacterial and human complex I share 14 core subunits that are essential for enzymatic function; however, the role and necessity of the remaining 31 human accessory subunits is unclear. The incorporation of accessory subunits into the complex increases the cellular energetic cost and has necessitated the involvement of numerous assembly factors for complex I biogenesis. Here we use gene editing to generate human knockout cell lines for each accessory subunit. We show that 25 subunits are strictly required for assembly of a functional complex and 1 subunit is essential for cell viability. Quantitative proteomic analysis of cell lines revealed that loss of each subunit affects the stability of other subunits residing in the same structural module. Analysis of proteomic changes after the loss of specific modules revealed that ATP5SL and DMAC1 are required for assembly of the distal portion of the complex I membrane arm. Our results demonstrate the broad importance of accessory subunits in the structure and function of human complex I. Coupling gene-editing technology with proteomics represents a powerful tool for dissecting large multi-subunit complexes and enables the study of complex dysfunction at a cellular level.

  11. Using yeast two-hybrid system to detect interactions of ATP synthase subunits from Spinacia oleracea

    Institute of Scientific and Technical Information of China (English)

    石晓冰; 魏家绵; 沈允钢

    2000-01-01

    Subunit interactions among the chloroplast ATP synthase subunits were studied using the yeast two-hybrid system. Various pairwise combinations of genes encoding a, p, y, 8 and e subunits of Spinach ATP synthase fused to the binding domain or activation domain of GAL4 DNA were introduced into yeast and then expression of a reporter gene encoding p-galactosidase was detected. Of all the combinations, that of y and e subunit genes showed the highest level of reporter gene expression, while those of a and p, a and e, p and e and p and 8 induced stable and significant reporter gene expression. The combination of 8 and e as well as that of 8 and y induced weak and unstable reporter gene expression. However, combinations of a and y, p and y and a and 8 did not induce reporter gene expression. These results suggested that specific and strong interactions between y and e, a and p, a and e, p and e and p and 8 subunits, and weak and transient interactions between 8 and e and 8 and y subunits occurred in the yeast

  12. Gene fusions with human carbonic anhydrase II for efficient expression and rapid single-step recovery of recombinant proteins: expression of the Escherichia coli F1-ATPase epsilon subunit.

    Science.gov (United States)

    Van Heeke, G; Shaw, R; Schnizer, R; Couton, J M; Schuster, S M; Wagner, F W

    1993-08-01

    A new expression vector was constructed which allows the overproduction in Escherichia coli of tripartite proteins consisting of human carbonic anhydrase isozyme II (hCAII), a peptide linker containing an enterokinase cleavage site, and a target protein of interest. Carbonic anhydrase is soluble and stable in E. coli and serves as a highly specific purification tag in the recovery of the fusion protein by a single affinity chromatography step. The enterokinase cleavage site was engineered into the construct to allow accurate and efficient release of the target protein. To demonstrate the practical value of this vector, the E. coli F1-ATPase epsilon subunit was expressed as a fusion with hCAII. After a single purification step, biologically active recombinant E. coli F1-ATPase epsilon subunit was recovered following proteolytic removal of the hCAII moiety.

  13. Associations of ABCB1, NFKB1, CYP3A, and NR1I2 polymorphisms with cyclosporine trough concentrations in Chinese renal transplant recipients.

    Science.gov (United States)

    Zhang, Yu; Li, Jia-li; Fu, Qian; Wang, Xue-ding; Liu, Long-shan; Wang, Chang-xi; Xie, Wen; Chen, Zhuo-jia; Shu, Wen-ying; Huang, Min

    2013-04-01

    Cyclosporine requires close therapeutic drug monitoring because of its narrow therapeutic index and marked inter-individual pharmacokinetic variation. In this study, we investigated the associations of CYP3A4, CYP3A5, ABCB1, NFKB1, and NR1I2 polymorphisms with cyclosporine concentrations in Chinese renal transplant recipients in the early period after renal transplantation. A total of 101 renal transplant recipients receiving cyclosporine were genotyped for CYP3A4(*)1G, CYP3A5(*)3, ABCB1 C1236T, G2677T/A, C3435T, NFKB1 -94 ins/del ATTG, and NR1I2 polymorphisms. Cyclosporine whole blood levels were measured by a fluorescence polarization immunoassay. Trough concentrations of cyclosporine were determined for days 7-18 following transplantation. The dose-adjusted trough concentration (C0) of cyclosporine in ABCB1 2677 TT carriers was significantly higher than that in GG carriers together with GT carriers [90.4±24.5 vs 67.8±26.8 (ng/mL)/(mg/kg), P=0.001]. ABCB1 3435 TT carriers had a significantly higher dose-adjusted C0 of cyclosporine than CC carriers together with CT carriers [92.0±24.0 vs 68.4±26.5 (ng/mL)/(mg/kg), P=0.002]. Carriers of the ABCB1 1236TT-2677TT-3435TT haplotype had a considerably higher CsA C0/D than carriers of other genotypes [97.2±21.8 vs 68.7±26.9 (ng/mL)/(mg/kg), P=0.001]. Among non-carriers of the ABCB1 2677 TT and 3435 TT genotypes, patients with the NFKB1 -94 ATTG ins/ins genotype had a significantly higher dose-adjusted C0 than those with the -94 ATTG del/del genotype [75.9±32.9 vs 55.1±15.1 (ng/mL)/(mg/kg), P=0.026]. These results illustrate that the ABCB1 and NFKB1 genotypes are closely correlated with cyclosporine trough concentrations, suggesting that these SNPs are useful for determining the appropriate dose of cyclosporine.

  14. AChR deficiency due to epsilon-subunit mutations : two common mutations in the Netherlands

    NARCIS (Netherlands)

    Faber, Catharina G.; Molenaar, Peter C.; Vles, Johannes S. H.; Bonifati, Domenic M.; Verschuuren, Jan J. G. M.; van Doorn, Pieter A.; Kuks, Jan B. M.; Wokke, John H. J.; Beeson, David; De Baets, Marc

    2009-01-01

    Congenital myasthenic syndromes are a clinically and genetically heterogeneous group of hereditary disorders affecting neuromuscular transmission. We have identified mutations within the acetylcholine receptor (AChR) epsilon-subunit gene underlying congenital myasthenic syndromes in nine patients

  15. AChR deficiency due to ε-subunit mutations: Two common mutations in the Netherlands

    NARCIS (Netherlands)

    C.G. Faber (Carin); P.C. Molenaar (Peter); J.S.H. Vles (Johannes); D.M. Bonifati (Domenic); J.J. Verschuuren (Jan); P.A. van Doorn (Pieter); J.B.M. Kuks (Jan); J.H.J. Wokke (John); D. Beeson (David); M.H. de Baets (Marc)

    2009-01-01

    textabstractCongenital myasthenic syndromes are a clinically and genetically heterogeneous group of hereditary disorders affecting neuromuscular transmission. We have identified mutations within the acetylcholine receptor (AChR) ε-subunit gene underlying congenital myasthenic syndromes in nine

  16. Multiple roles of Rev3, the catalytic subunit of polzeta in maintaining genome stability in vertebrates

    NARCIS (Netherlands)

    E. Sonoda (Eiichiro); S. Takeda (Shiunichi); T. Okada (Takashi); G.Y. Zhao (Guang); S. Tateishi (Satoshi); K. Araki (Kasumi); M. Yamaizumi (Masaru); T. Yagi (Takashi); N.S. Verkaik (Nicole); D.C. van Gent (Dik); M. Takata (Minoru)

    2003-01-01

    textabstractTranslesion DNA synthesis (TLS) and homologous DNA recombination (HR) are two major postreplicational repair (PRR) pathways. The REV3 gene of Saccharomyces cerevisiae encodes the catalytic subunit of DNA polymerase zeta, which is involved in mutagenic TLS. To

  17. AChR deficiency due to ε-subunit mutations: Two common mutations in the Netherlands

    NARCIS (Netherlands)

    C.G. Faber (Carin); P.C. Molenaar (Peter); J.S.H. Vles (Johannes); D.M. Bonifati (Domenic); J.J. Verschuuren (Jan); P.A. van Doorn (Pieter); J.B.M. Kuks (Jan); J.H.J. Wokke (John); D. Beeson (David); M.H. de Baets (Marc)

    2009-01-01

    textabstractCongenital myasthenic syndromes are a clinically and genetically heterogeneous group of hereditary disorders affecting neuromuscular transmission. We have identified mutations within the acetylcholine receptor (AChR) ε-subunit gene underlying congenital myasthenic syndromes in nine patie

  18. AChR deficiency due to epsilon-subunit mutations : two common mutations in the Netherlands

    NARCIS (Netherlands)

    Faber, Catharina G.; Molenaar, Peter C.; Vles, Johannes S. H.; Bonifati, Domenic M.; Verschuuren, Jan J. G. M.; van Doorn, Pieter A.; Kuks, Jan B. M.; Wokke, John H. J.; Beeson, David; De Baets, Marc

    2009-01-01

    Congenital myasthenic syndromes are a clinically and genetically heterogeneous group of hereditary disorders affecting neuromuscular transmission. We have identified mutations within the acetylcholine receptor (AChR) epsilon-subunit gene underlying congenital myasthenic syndromes in nine patients (s

  19. AChR deficiency due to epsilon-subunit mutations : two common mutations in the Netherlands

    NARCIS (Netherlands)

    Faber, Catharina G.; Molenaar, Peter C.; Vles, Johannes S. H.; Bonifati, Domenic M.; Verschuuren, Jan J. G. M.; van Doorn, Pieter A.; Kuks, Jan B. M.; Wokke, John H. J.; Beeson, David; De Baets, Marc

    2009-01-01

    Congenital myasthenic syndromes are a clinically and genetically heterogeneous group of hereditary disorders affecting neuromuscular transmission. We have identified mutations within the acetylcholine receptor (AChR) epsilon-subunit gene underlying congenital myasthenic syndromes in nine patients (s

  20. Protein kinase A catalytic subunit isoform PRKACA; History, function and physiology.

    Science.gov (United States)

    Turnham, Rigney E; Scott, John D

    2016-02-15

    Our appreciation of the scope and influence of second messenger signaling has its origins in pioneering work on the cAMP-dependent protein kinase. Also called protein kinase A (PKA), this holoenzyme exists as a tetramer comprised of a regulatory (R) subunit dimer and two catalytic (C) subunits. Upon binding of two molecules of the second messenger cAMP to each R subunit, a conformational change in the PKA holoenzyme occurs to release the C subunits. These active kinases phosphorylate downstream targets to propagate cAMP responsive cell signaling events. This article focuses on the discovery, structure, cellular location and physiological effects of the catalytic subunit alpha of protein kinase A (encoded by the gene PRKACA). We also explore the potential role of this essential gene as a molecular mediator of certain disease states. Copyright © 2015 Elsevier B.V. All rights reserved.

  1. MspA nanopores from subunit dimers.

    Directory of Open Access Journals (Sweden)

    Mikhail Pavlenok

    Full Text Available Mycobacterium smegmatis porin A (MspA forms an octameric channel and represents the founding member of a new family of pore proteins. Control of subunit stoichiometry is important to tailor MspA for nanotechnological applications. In this study, two MspA monomers were connected by linkers ranging from 17 to 62 amino acids in length. The oligomeric pore proteins were purified from M. smegmatis and were shown to form functional channels in lipid bilayer experiments. These results indicated that the peptide linkers did not prohibit correct folding and localization of MspA. However, expression levels were reduced by 10-fold compared to wild-type MspA. MspA is ideal for nanopore sequencing due to its unique pore geometry and its robustness. To assess the usefulness of MspA made from dimeric subunits for DNA sequencing, we linked two M1-MspA monomers, whose constriction zones were modified to enable DNA translocation. Lipid bilayer experiments demonstrated that this construct also formed functional channels. Voltage gating of MspA pores made from M1 monomers and M1-M1 dimers was identical indicating similar structural and dynamic channel properties. Glucose uptake in M. smegmatis cells lacking porins was restored by expressing the dimeric mspA M1 gene indicating correct folding and localization of M1-M1 pores in their native membrane. Single-stranded DNA hairpins produced identical ionic current blockades in pores made from monomers and subunit dimers demonstrating that M1-M1 pores are suitable for DNA sequencing. This study provides the proof of principle that production of single-chain MspA pores in M. smegmatis is feasible and paves the way for generating MspA pores with altered stoichiometries. Subunit dimers enable better control of the chemical and physical properties of the constriction zone of MspA. This approach will be valuable both in understanding transport across the outer membrane in mycobacteria and in tailoring MspA for nanopore

  2. Sigma-1 Receptor Antagonist BD1047 Reduces Mechanical Allodynia in a Rat Model of Bone Cancer Pain through the Inhibition of Spinal NR1 Phosphorylation and Microglia Activation.

    Science.gov (United States)

    Zhu, Shanshan; Wang, Chenchen; Han, Yuan; Song, Chao; Hu, Xueming; Liu, Yannan

    2015-01-01

    Previous studies have demonstrated that sigma-1 receptor plays important roles in the induction phase of rodent neuropathic pain; however, whether it is involved in bone cancer pain (BCP) and the underlying mechanisms remain elusive. The aim of this study was to examine the potential role of the spinal sigma-1 receptor in the development of bone cancer pain. Walker 256 mammary gland carcinoma cells were implanted into the intramedullary space of the right tibia of Sprague-Dawley rats to induce ongoing bone cancer-related pain behaviors; our findings indicated that, on days 7, 10, 14, and 21 after operation, the expression of sigma-1 receptor in the spinal cord was higher in BCP rats compared to the sham rats. Furthermore, intrathecal injection of 120 nmol of sigma-1 receptor antagonist BD1047 on days 5, 6, and 7 after operation attenuated mechanical allodynia as well as the associated induction of c-Fos and activation of microglial cells, NR1, and the subsequent Ca(2+)-dependent signals of BCP rats. These results suggest that sigma-1 receptor is involved in the development of bone cancer pain and that targeting sigma-1 receptor may be a new strategy for the treatment of bone cancer pain.

  3. The Niwot Ridge Subalpine Forest US-NR1 AmeriFlux site - Part 1: Data acquisition and site record-keeping

    Science.gov (United States)

    Burns, Sean P.; Maclean, Gordon D.; Blanken, Peter D.; Oncley, Steven P.; Semmer, Steven R.; Monson, Russell K.

    2016-09-01

    The Niwot Ridge Subalpine Forest AmeriFlux site (US-NR1) has been measuring eddy-covariance ecosystem fluxes of carbon dioxide, heat, and water vapor since 1 November 1998. Throughout this 17-year period there have been changes to the instrumentation and improvements to the data acquisition system. Here, in Part 1 of this three-part series of papers, we describe the hardware and software used for data-collection and metadata documentation. We made changes to the data acquisition system that aimed to reduce the system complexity, increase redundancy, and be as independent as possible from any network outages. Changes to facilitate these improvements were (1) switching to a PC/104-based computer running the National Center for Atmospheric Research (NCAR) In-Situ Data Acquisition Software (NIDAS) that saves the high-frequency data locally and over the network, and (2) time-tagging individual 10 Hz serial data samples using network time protocol (NTP) coupled to a GPS-based clock, providing a network-independent, accurate time base. Since making these improvements almost 2 years ago, the successful capture of high-rate data has been better than 99.98 %. We also provide philosophical concepts that shaped our design of the data system and are applicable to many different types of environmental data collection.

  4. Metagenomics of Water Column Microbes Near Brine Pool NR1 and adjacent regions of the Northern Gulf of Mexico Collected in Fall 2009

    Science.gov (United States)

    Wood, A. M.; Goodwin, K. D.; Brami, D.; Schwartz, A.; Toledo, G.

    2012-12-01

    High-throughput sequencing was applied to eight water column samples collected from the Gulf of Mexico in 2009 in regions SW and west of the 2010 Macondo oil spill. Samples were collected by Niskin-equipped CTD (~200 and ~650 m depths) at two locations, including a site over a methane brine pool (Brine Pool NR1). In addition, seawater was collected ~3m lateral of the pool (649m depth) via Niskin bottle equipped on the Johnson-Sea-Link submersible. Unassembled reads were submitted to the Synthetic Genomics bioinformatics pipeline for taxonomic analysis. The distribution of Bacteria (56-73%), Archae (7-16%), Eukaryotes (12-23%), and unclassified sequences (6-10%) were similar for all samples. However, certain taxonomic classifications were relatively more abundant in deeper samples, and differences were noted for samples collected by submersible. For example, Methylophaga was classified as 38% of the order Thiotrichales for the Niskin/submersible sample compared to 0% in the 200m-depth samples and 3-11% in the 650m samples. Methylophaga is a genus of indigenous methylotrophs reported to respond during the Deepwater Horizon event of 2010. In contrast, sequence abundance for Oceanospirillales, also reported to respond during the event, was similar for all samples (6-9% of the gamma-proteobacteria).

  5. Identification of the fifth subunit of Saccharomyces cerevisiae replication factor C.

    Science.gov (United States)

    Gary, S L; Burgers, M J

    1995-01-01

    Yeast replication factor C (RF-C) is a multipolypeptide complex required for chromosomal DNA replication. Previously this complex was known to consist of at least four subunits. We here report the identification of a fifth RF-C subunit from Saccharomyces cerevisiae, encoded by the RFC5 (YBR0810) gene. This subunit exhibits highest homology to the 38 kDa subunit (38%) of human RF-C (activator 1). Like the other four RFC genes, the RFC5 gene is essential for yeast viability, indicating an essential function for each subunit. RFC5 mRNA is expressed at steady-state levels throughout the mitotic cell cycle. Upon overexpression in Escherichia coli Rfc5p has an apparent molecular mass of 41 kDa. Overproduction of RF-C activity in yeast is dependent on overexpression of the RFC5 gene together with overexpression of the RFC1-4 genes, indicating that the RFC5 gene product forms an integral subunit of this replication factor. Images PMID:8559655

  6. Control of gene expression by the retinoic acid-related orphan receptor alpha in HepG2 human hepatoma cells.

    Directory of Open Access Journals (Sweden)

    Caroline Chauvet

    Full Text Available Retinoic acid-related Orphan Receptor alpha (RORα; NR1F1 is a widely distributed nuclear receptor involved in several (pathophysiological functions including lipid metabolism, inflammation, angiogenesis, and circadian rhythm. To better understand the role of this nuclear receptor in liver, we aimed at displaying genes controlled by RORα in liver cells by generating HepG2 human hepatoma cells stably over-expressing RORα. Genes whose expression was altered in these cells versus control cells were displayed using micro-arrays followed by qRT-PCR analysis. Expression of these genes was also altered in cells in which RORα was transiently over-expressed after adenoviral infection. A number of the genes found were involved in known pathways controlled by RORα, for instance LPA, NR1D2 and ADIPOQ in lipid metabolism, ADIPOQ and PLG in inflammation, PLG in fibrinolysis and NR1D2 and NR1D1 in circadian rhythm. This study also revealed that genes such as G6PC, involved in glucose homeostasis, and AGRP, involved in the control of body weight, are also controlled by RORα. Lastly, SPARC, involved in cell growth and adhesion, and associated with liver carcinogenesis, was up-regulated by RORα. SPARC was found to be a new putative RORα target gene since it possesses, in its promoter, a functional RORE as evidenced by EMSAs and transfection experiments. Most of the other genes that we found regulated by RORα also contained putative ROREs in their regulatory regions. Chromatin immunoprecipitation (ChIP confirmed that the ROREs present in the SPARC, PLG, G6PC, NR1D2 and AGRP genes were occupied by RORα in HepG2 cells. Therefore these genes must now be considered as direct RORα targets. Our results open new routes on the roles of RORα in glucose metabolism and carcinogenesis within cells of hepatic origin.

  7. Changes Of GR and hippocampal NR1 and effect of midazolam-Ketamine in mice after severe TBI%小鼠严重颅脑撞击伤早期咪唑安定-氯胺酮对肝脏GR与海马NR1变化的影响

    Institute of Scientific and Technical Information of China (English)

    屈强; 史忠; 粟永萍

    2010-01-01

    目的 研究小鼠闭合性严重颅脑损伤(TBI)后肝脏糖皮质激素受体(GR)、海马N-甲基-D-天门冬氨酸受体(NR)蛋白功能亚单位1(NR1)蛋白水平变化及其与血清TNF-α、IL-1β变化的关系,以及应用咪唑安定-氯胺酮干预后的变化.方法 利用BIM-Ⅲ型小型多功能动物撞击机对小鼠清醒致伤后将其随机分为5组,即假致伤组(J组)、致伤对照组(N组)、致伤后氯胺酮治疗组(K组)、致伤后咪唑安定治疗组(M组)、致伤后复合用药治疗组(F组).于致伤后30 min及2、8、24、48、72 h采用Western blot免疫印迹法检测大脑皮质和肝脏GR蛋白水平变化,用酶联免疫吸附法(ELISA)检测各组外周血清中TNF-α、IL-1β含量.结果 肝脏GR蛋白表达在致伤2 h开始降低,8 h呈现恢复趋势,72 h仍未完全正常;海马NR1蛋白表达在致伤后2 h开始明显降低,24 h基本恢复,72 h明显增加;应用咪唑安定-氯胺酮干预后可明显降低GR、NR1蛋白表达的这种变化趋势;外周血中TNF-α、IL-1β含量致伤后明显升高,均具有两个峰值特征,应用咪唑安定-氯胺酮干预后可明显降低二者的升高.结论 严重TBI后存在糖皮质激素抵抗,NR激活可能是其中一个重要原因.咪唑安定、氯胺酮能明显抑制HPA轴的兴奋性,改善糖皮质激素抵抗,调控应激反应.其作用机制除直接抑制炎性细胞因子释放外,还可能有中枢性的作用机制,其中之一可能与调节NR1蛋白表达有关.

  8. NMDA receptor subunit expression and PAR2 receptor activation in colospinal afferent neurons (CANs during inflammation induced visceral hypersensitivity

    Directory of Open Access Journals (Sweden)

    Caudle Robert M

    2009-09-01

    Full Text Available Abstract Background Visceral hypersensitivity is a clinical observation made when diagnosing patients with functional bowel disorders. The cause of visceral hypersensitivity is unknown but is thought to be attributed to inflammation. Previously we demonstrated that a unique set of enteric neurons, colospinal afferent neurons (CANs, co-localize with the NR1 and NR2D subunits of the NMDA receptor as well as with the PAR2 receptor. The aim of this study was to determine if NMDA and PAR2 receptors expressed on CANs contribute to visceral hypersensitivity following inflammation. Recently, work has suggested that dorsal root ganglion (DRG neurons expressing the transient receptor potential vanilloid-1 (TRPV1 receptor mediate inflammation induced visceral hypersensitivity. Therefore, in order to study CAN involvement in visceral hypersensitivity, DRG neurons expressing the TRPV1 receptor were lesioned with resiniferatoxin (RTX prior to inflammation and behavioural testing. Results CANs do not express the TRPV1 receptor; therefore, they survive following RTX injection. RTX treatment resulted in a significant decrease in TRPV1 expressing neurons in the colon and immunohistochemical analysis revealed no change in peptide or receptor expression in CANs following RTX lesioning as compared to control data. Behavioral studies determined that both inflamed non-RTX and RTX animals showed a decrease in balloon pressure threshold as compared to controls. Immunohistochemical analysis demonstrated that the NR1 cassettes, N1 and C1, of the NMDA receptor on CANs were up-regulated following inflammation. Furthermore, inflammation resulted in the activation of the PAR2 receptors expressed on CANs. Conclusion Our data show that inflammation causes an up-regulation of the NMDA receptor and the activation of the PAR2 receptor expressed on CANs. These changes are associated with a decrease in balloon pressure in response to colorectal distension in non-RTX and RTX lesioned

  9. The phosphorylation pattern of bovine heart complex I subunits

    DEFF Research Database (Denmark)

    Palmisano, Giuseppe; Sardanelli, Anna Maria; Signorile, Anna;

    2007-01-01

    The phosphoproteome of bovine heart complex I of the respiratory chain has been analysed with a procedure based on nondenaturing gel electrophoretic separation of complex I from small quantities of mitochondria samples, in-gel digestion, in combination with phosphopeptide enrichment by titanium...... dioxide and MS. The results, complemented by analyses of purified samples of complex I, showed phosphorylation of five subunits of the complex, 42 kDa (human gene NDUFA10), ESSS, B14.5a (human gene NDUFA7), B14.5b (human gene NDUFC2) and B16.6 (GRIM-19). MS also revealed the presence of phosphorylated...

  10. Lurasidone and fluoxetine reduce novelty-induced hypophagia and NMDA receptor subunit and PSD-95 expression in mouse brain.

    Science.gov (United States)

    Stan, Tiberiu Loredan; Sousa, Vasco Cabral; Zhang, Xiaoqun; Ono, Michiko; Svenningsson, Per

    2015-10-01

    Lurasidone, a novel second-generation antipsychotic agent, exerts antidepressant actions in patients suffering from bipolar type I disorder. Lurasidone acts as a high affinity antagonist at multiple monoamine receptors, particularly 5-HT2A, 5-HT7, D2 and α2 receptors, and as a partial agonist at 5-HT1A receptors. Accumulating evidence indicates therapeutic actions by monoaminergic antidepressants are mediated via alterations of glutamate receptor-mediated neurotransmission. Here, we used mice and investigated the effects of chronic oral administration of vehicle, lurasidone (3 or 10mg/kg) or fluoxetine (20mg/kg) in the novelty induced hypophagia test, a behavioral test sensitive to chronic antidepressant treatment. We subsequently performed biochemical analyses on NMDA receptor subunits and associated proteins. Both lurasidone and fluoxetine reduced the latency to feed in the novelty-induced hypophagia test. Western blotting experiments showed that both lurasidone and fluoxetine decreased the total levels of NR1, NR2A and NR2B subunits of NMDA receptors and PSD-95 (PostSynaptic Density-95) in hippocampus and prefrontal cortex. Taken together, these data indicate that antidepressant/anxiolytic-like effects of lurasidone, as well as fluoxetine, could involve reduced NMDA receptor-mediated signal transduction, particularly in pathways regulated by PSD-95, in hippocampus and prefrontal cortex.

  11. Effect of developmental lead exposure on synaptic plasticity and N—methyl—D—aspartate receptor subunit in rat hippocampus

    Institute of Scientific and Technical Information of China (English)

    RuanDY; SuiL

    2002-01-01

    Chronic lead(Pb) exposure is known to be associated with learning and memory,and cognitive dysfunction in children.Previous studies have demonstrated that Pb exposure may impair neuronal process underlying synaptic plasticity via a direct interaction with N-methyl-D-aspartate (NMDA) receptors(NMDARs).The studies described here were carried out to investigate effect of developmental Pb exposure on long-term potentiation(LTP),long-tern depression(LTD) and NMDAs subunits in rat hippocampus.The results are listed as follows:(1)low-level Pb exposture can impair the induction and maintenance of LTP in vivo and in vitro;(2)the Pb-induced impairment of LTD magnitude was an age-related decline in area CA1 of rat hippocampus;(3)chronic Pb exposure affected two components,voltage-gated calcium channel-dependent LTD and NMDARs-dependent LTD,of LTD induction in area CA1 of rat hippocampus;(4)different effects of developmental Pb exposure on NMDA receptor NR1,NR2A,NR2B,NR2C,NR2D and NR3A subunits in area CA1,CA2,CA3 and CA4 of rat hippocampus were observed;(5)the α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors enriched in area CA1,CA3 and dentate gyrus and kainite receptors enriched in area CA1 and dentate gyrus of rat hippocampus were impaired by Pb exposure.

  12. Localization of P42 and F(1)-ATPase α-subunit homolog of the gliding machinery in Mycoplasma mobile revealed by newly developed gene manipulation and fluorescent protein tagging.

    Science.gov (United States)

    Tulum, Isil; Yabe, Masaru; Uenoyama, Atsuko; Miyata, Makoto

    2014-05-01

    Mycoplasma mobile has a unique mechanism that enables it to glide on solid surfaces faster than any other gliding mycoplasma. To elucidate the gliding mechanism, we developed a transformation system for M. mobile based on a transposon derived from Tn4001. Modification of the electroporation conditions, outgrowth time, and colony formation from the standard method for Mycoplasma species enabled successful transformation. A fluorescent-protein tagging technique was developed using the enhanced yellow fluorescent protein (EYFP) and applied to two proteins that have been suggested to be involved in the gliding mechanism: P42 (MMOB1050), which is transcribed as continuous mRNA with other proteins essential for gliding, and a homolog of the F1-ATPase α-subunit (MMOB1660). Analysis of the amino acid sequence of P42 by PSI-BLAST suggested that P42 evolved from a common ancestor with FtsZ, the bacterial tubulin homologue. The roles of P42 and the F(1)-ATPase subunit homolog are discussed as part of our proposed gliding mechanism.

  13. Localization of P42 and F1-ATPase α-Subunit Homolog of the Gliding Machinery in Mycoplasma mobile Revealed by Newly Developed Gene Manipulation and Fluorescent Protein Tagging

    Science.gov (United States)

    Tulum, Isil; Yabe, Masaru; Uenoyama, Atsuko

    2014-01-01

    Mycoplasma mobile has a unique mechanism that enables it to glide on solid surfaces faster than any other gliding mycoplasma. To elucidate the gliding mechanism, we developed a transformation system for M. mobile based on a transposon derived from Tn4001. Modification of the electroporation conditions, outgrowth time, and colony formation from the standard method for Mycoplasma species enabled successful transformation. A fluorescent-protein tagging technique was developed using the enhanced yellow fluorescent protein (EYFP) and applied to two proteins that have been suggested to be involved in the gliding mechanism: P42 (MMOB1050), which is transcribed as continuous mRNA with other proteins essential for gliding, and a homolog of the F1-ATPase α-subunit (MMOB1660). Analysis of the amino acid sequence of P42 by PSI-BLAST suggested that P42 evolved from a common ancestor with FtsZ, the bacterial tubulin homologue. The roles of P42 and the F1-ATPase subunit homolog are discussed as part of our proposed gliding mechanism. PMID:24509320

  14. Structural characterization of recombinant crustacyanin subunits from the lobster Homarus americanus.

    Science.gov (United States)

    Ferrari, Michele; Folli, Claudia; Pincolini, Elisa; McClintock, Timothy S; Rössle, Manfred; Berni, Rodolfo; Cianci, Michele

    2012-08-01

    Crustacean crustacyanin proteins are linked to the production and modification of carapace colour, with direct implications for fitness and survival. Here, the structural and functional properties of the two recombinant crustacyanin subunits H(1) and H(2) from the American lobster Homarus americanus are reported. The two subunits are structurally highly similar to the corresponding natural apo crustacyanin CRTC and CRTA subunits from the European lobster H. gammarus. Reconstitution studies of the recombinant crustacyanin proteins H(1) and H(2) with astaxanthin reproduced the bathochromic shift of 85-95 nm typical of the natural crustacyanin subunits from H. gammarus in complex with astaxanthin. Moreover, correlations between the presence of crustacyanin genes in crustacean species and the resulting carapace colours with the spectral properties of the subunits in complex with astaxanthin confirmed this genotype-phenotype linkage.

  15. Stoichiometry of δ subunit containing GABAA receptors

    Science.gov (United States)

    Patel, B; Mortensen, M; Smart, T G

    2014-01-01

    Background and Purpose Although the stoichiometry of the major synaptic αβγ subunit-containing GABAA receptors has consensus support for 2α:2β:1γ, a clear view of the stoichiometry of extrasynaptic receptors containing δ subunits has remained elusive. Here we examine the subunit stoichiometry of recombinant α4β3δ receptors using a reporter mutation and a functional electrophysiological approach. Experimental Approach Using site-directed mutagenesis, we inserted a highly characterized 9′ serine to leucine mutation into the second transmembrane (M2) region of α4, β3 and δ subunits that increases receptor sensitivity to GABA. Whole-cell, GABA-activated currents were recorded from HEK-293 cells co-expressing different combinations of wild-type (WT) and/or mutant α4(L297S), β3(L284S) and δ(L288S) subunits. Key Results Recombinant receptors containing one or more mutant subunits showed increased GABA sensitivity relative to WT receptors by approximately fourfold, independent of the subunit class (α, β or δ) carrying the mutation. GABA dose–response curves of cells co-expressing WT subunits with their respective L9′S mutants exhibited multiple components, with the number of discernible components enabling a subunit stoichiometry of 2α, 2β and 1δ to be deduced for α4β3δ receptors. Varying the cDNA transfection ratio by 10-fold had no significant effect on the number of incorporated δ subunits. Conclusions and Implications Subunit stoichiometry is an important determinant of GABAA receptor function and pharmacology, and δ subunit-containing receptors are important mediators of tonic inhibition in several brain regions. Here we demonstrate a preferred subunit stoichiometry for α4β3δ receptors of 2α, 2β and 1δ. PMID:24206220

  16. 云南保山和普洱地区带绦虫线粒体DNA基因编码核糖体RNA小亚基基因序列分析%Analysis of the mitochondrial DNA-gene encoding ribosomal RNA small subunit gene sequence of Taenia cestode from Baoshan and Puer areas in Yunnan Province

    Institute of Scientific and Technical Information of China (English)

    刘爱波; 杨毅梅

    2011-01-01

    Objective To identify Taenia cestodes specimens collected from Baoshan and Puer regions of Yunnan Province by analyzing mitochondrial DNA gene encoding ribosomal RNA small subunit (mtDNA-12S rRNA) gene sequence. Methods The adult Taenia cestode samples were collected from Baoshan and Puer regions of Yunnan Province. The genomic DNA was extracted and mtDNA-12S rRAN gene was amplified by polymerase chain reaction (PCR), then sequenced.Combined with the known mtDNA-12S rRNA gene sequence of Taenia solium, Taenia saginata,Taenia asiatica in GenBank, homology tree and phylogenetic tree were constructed by DNA MAN software. Results Taenia cestode homology tree and phylogenetic tree showed that gene sequences of BS1, BS2, BS4 and BS5 were most close to YZ with identity of 99% and those of BS3, BS6, BST,PE1 and PE2 were most close to ND with identity of 99%. Conclusions Taenia saginata and Taenia asiatica can be found in Baoshan area, while Taenia saginata can be found in Puer area. The gene sequence of mtDNA-12S rRNA can be used for clarifying the three types of Taenia cestode.%目的 利用线粒体DNA基因编码核糖体RNA小亚基(mtDNA-12S rRNA)基因序列分析对采自云南保山、普洱地区的带绦虫标本进行鉴定.方法 选取保山(7条,BS1-BS7)、普洱(2条,PE1~PE2)带绦虫成虫节片,抽提基因组DNA,PCR扩增mtDNA-12S rRNA基因序列,并测序;结合GenBank中已知的猪带绦虫(ZD)、牛带绦虫(ND)、亚洲带绦虫(YZ)mtDNA-12S rRNA基因序列,经DNA MAN软件处理后构建同源树状图与系统发育树状图.结果 带绦虫同源树与系统发育树状图显示,BS1、BS2、BS4、BS5与YZ的同源性最近(99%).BS3、BS6、BS7、PE1、PE2与ND的同源性最近(99%).结论 云南保山存在牛带绦虫与亚洲带绦虫,普洱存在牛带绦虫,mtDNA-12S rRNA基因序列可用于三种带绦虫的分类研究.

  17. Isoform-specific Regulation of Sodium pump α-subunit Gene Expression i n Aortic Smooth Muscle Cell of 1-kidney-1-clip Hypertensive Rats%“一肾一夹”高血压大鼠动脉平滑肌细胞钠泵α亚单位的基因表达

    Institute of Scientific and Technical Information of China (English)

    原卫清; 王颢; 吕卓人

    2001-01-01

    Objective To investigate the gene expression of sodium pump α-subunit in aortic smooth muscle of 1-kidney-1-clip (1k1c) hypertensive rats.  Methods 1k1c hypertensive rats were prepared by partially ligating the left renal artery and removing the right kidney. 4 weeks later, all the rats were killed and sodium pump α1- , α2-, and α3-subunit in aortic smooth muscles were detected with re