Biomedical subjects

J Zhou

Publications and source records attributed to J Zhou.

At least 379 records · Page 21Linked to original sources

PubMed1999

Crystal structure and mechanism of histone acetylation of the yeast GCN5 transcriptional coactivator.

The yeast GCN5 (yGCN5) transcriptional coactivator functions as a histone acetyltransferase (HAT) to promote transcriptional activation. Here, we present the high resolution crystal structure of the HAT domain of yGCN5 and probe the functional importance of a conserved glutamate residue. The structure reveals a central protein core associated with AcCoA binding that appears to be structurally conserved among a superfamily of N-acetyltransferases, including yeast histone acetyltransferase 1 and Serratia marcescens aminoglycoside 3-N-acetyltransferase. A pronounced cleft lying above this core, and flanked by N- and C-terminal regions that show no sequence conservation within N-acetyltransferase enzymes, is implicated by cross-species conservation and mutagenesis studies to be a site for histone substrate binding and catalysis. Located at the bottom of this cleft is a conserved glutamate residue (E173) that is in position to play an important catalytic role in histone acetylation. Functional analysis of an E173Q mutant yGCN5 protein implicates glutamate 173 to function as a general base for catalysis. Together, a correlation of the yGCN5 structure with functionally debilitating yGCN5 mutations provides a paradigm for understanding the structure/function relationships of the growing number of transcriptional regulators that function as histone acetyltransferase enzymes.

Acetylation↗

PubMed1999

Identification of a new, unorthodox member of the MAGE gene family.

Several tumor-associated antigen families, such as MAGE, GAGE/PAGE, PRAME, BAGE, and LAGE/NY-ESO-1, exist. These antigens are of particular interest in tumor immunology, because their expression, with exception of testis and fetal tissues, seems to be restricted to tumor cells only. We have identified a novel member of the MAGE gene family, MAGED1. Northern hybridization and RT-PCR demonstrated that the expression level of MAGED1 in different normal adult tissues is comparable to that in testis and fetal liver. Thus, MAGED1 does not possess an expression pattern characteristic of previously identified MAGE family genes, suggesting that the biology of the MAGE-family genes is more complex than previously thought. Chromosome mapping linked MAGED1 to marker AFM119xd6 (DXS1039) on chromosome Xp11.23.

Adult↗

PubMed1999

Identification of a novel transcriptional regulatory element within the promoter region of the keratinocyte growth factor gene that mediates inducibility to cyclic AMP.

Keratinocyte growth factor (KGF) plays a critical role for the normal development and morphogenesis of many different tissues and organs. Furthermore, its expression is induced during wound healing and in various chronic inflammatory diseases. To determine the molecular mechanisms which regulate KGF gene induction at the transcriptional level, we carried out in vitro studies using the human KGF promoter. We have identified a novel regulatory element, TGAGGTCAG, located between -39 and -46 bp (relative to the transcription start site) in the KGF basal promoter region, which binds to inducible transcription factors as determined by electrophoretic mobility shift assay. When cloned in front of a heterologous SV40 promoter this region conferred inducibility to forskolin, a stimulator of adenylate cyclase. In contrast, various mutated forms of this region were either partially or completely impaired in their ability to mediate induction to forskolin. The TGAGGTCAG sequence shared homology to both the cAMP responsive element (CRE) and CCAAT/enhancer binding protein (C/EBP) consensus binding sites. An oligonucleotide comprising a consensus CRE binding site partially competed for the nuclear protein binding to the TGAGGTCAG site. Gel mobility supershift assays indicated that two members of the activating transcription factor (ATF) family of CRE binding proteins, ATF1 and ATF2, were part of the nuclear protein complex bound to this regulatory region. Furthermore, purified recombinant ATF2 was able to directly recognize and bind the TGAGGTCAG sequence. In contrast, no evidence was obtained for C/EBP transcription factors being part of the complex. These results suggest that members of the ATF family are involved in mediating the transcriptional regulation of the KGF gene in response to extracellular stimuli via a novel CRE regulatory element.

Activating Transcription Factor 1↗

PubMed1999

Solution structure of the catalytic domain of GCN5 histone acetyltransferase bound to coenzyme A.

Gene transcription requires the release of inactive DNA from its packaging of histone proteins. Following the discovery of the first transcription-associated histone acetyltransferase, tetrahymena GCN5, it was shown that yeast GCN5 is recruited to the promoter and causes hyper-acetylation of histones and transcriptional activation of target genes, establishing a direct connection between histone acetylation and transcriptional activation. Many other important transcription regulators have been found to have histone acetyltransferase activity, including TAFII230/250, p300/CBP and its associated factor PCAF. Here we present the solution structure of the catalytic domain of tGCN5 (residues 47-210) in complex with coenzyme A. The structure contains two domains; the amino-terminal domain is similar to those of other GCN5-related N-acetyltransferases but the carboxy-terminal domain is not. Coenzyme A binds in a deep hydrophobic pocket between the two domains. Chemical shift changes upon titration with histone H3 peptides indicate a binding site at the domain boundary opposite to the coenzyme A site. The structural data indicate a single-step acetyl-transfer reaction mechanism catalysed by a hydrogen bond to the backbone amide group of leucine 126 and the side-chain carboxyl group of a conserved acidic residue.

Acetyltransferases↗

PubMed1999

Immortalized human dorsal root ganglion cells differentiate into neurons with nociceptive properties.

A renewable source of human sensory neurons would greatly facilitate basic research and drug development. We had established previously conditionally immortalized human CNS cell lines that can differentiate into functional neurons (). We report here the development of an immortalized human dorsal root ganglion (DRG) clonal cell line, HD10.6, with a tetracycline-regulatable v-myc oncogene. In the proliferative condition, HD10.6 cells have a doubling time of 1.2 d and exhibit a neuronal precursor morphology. After differentiation of clone HD10.6 for 7 d in the presence of tetracycline, v-myc expression was suppressed, and >50% of the cells exhibited typical neuronal morphology, stained positively for neuronal cytoskeletal markers, and fired action potentials in response to current injection. Furthermore, this cell line was fate-restricted to a neuronal phenotype; even in culture conditions that promote Schwann cell or smooth muscle differentiation of neural crest stem cells, HD10.6 differentiated exclusively into neurons. Moreover, differentiated HD10.6 cells expressed sensory neuron-associated transcription factors and exhibited capsaicin sensitivity. Taken together, these data indicate that we have established an immortalized human DRG cell line that can differentiate into sensory neurons with nociceptive properties. The cell line HD10.6 represents the first example of a human sensory neuronal line and will be valuable for basic research, as well as for the discovery of novel drug targets and clinical candidates.

Action Potentials↗

PubMed1999

DNA damage by copper(II) complexes: coordination-structural dependence of reactivities.

A variety of copper complexes with different structural features have been shown to bind double-helical DNA with binding constants of 10(4)-10(7) M-1 and to promote double-strand DNA damage upon reductant/H2O2 activation. The interaction of the Cu complex with DNA results in hyperchromism and shifts to longer wavelengths of the strongest transitions in the Cu complexes, as well as striking hypochromism or hyperchromism of DNA absorption at 260 nm. In the presence of DMPO as the spin trap, the solution of each copper complex exhibits typical four-line ESR spectra of the hydroxyl radical by adding 2-mercaptoethanol and H2O2 to the solution. Quantitation by 2-deoxy-D-ribose shows that the competence of hydroxyl radical generation by the copper complexes upon reductant and H2O2 activation decreases in order, that is, Cu(HTCD)(2+)-Cu(Im)4Cl2-Cu(IDB) (NO3)2 > Cu(IDB)Cl2 > Cu(IDBt)Cl2. The copper complex-mediated hydroxyl radical, a powerful oxidant that attacks the adjacent DNA, is responsible for the DNA oxidative damage. The lambda DNA damage chemistry illustrates that the competence and selectivity of double-strand lambda DNA damage by the copper complexes are dependent on their geometric structures and types of ligands. The decreasing order of the DNA damage capacity by the present complexes is Cu(Im)4Cl2-Cu(IDB)-(NO3)2 > Cu(HTCD)2+ > Cu(IDBt)Cl2 > Cu(IDB)Cl2.

Animals↗

PubMed1999

Alzheimer's disease-related gene expression in the brain of senescence accelerated mouse.

The levels of Alzheimer's disease (AD)-related genes, including beta-amyloid precursor protein(APP), presenilin-1 (PS-1), PS-2, apoE, tau, c-fos, neural cell adhesion molecular 180 (NCAM-180), TGF-beta 1, IL-1 alpha/beta, IL-6, TNF-alpha/beta, alpha-2-Macroglobulin (alpha 2M), class II major histocompatibility antigen la (MHCII la), bcl-2 alpha, glucocorticoid receptor-alpha (GR alpha) and mineralocorticoid receptor (MR) mRNAs were determined by reverse transcription polymerase chain reaction (RT-PCR) in the hippocampus and cerebral cortex of senescence accelerated mouse (SAM). The levels of TGF-beta 1, IL-1 alpha, TNF-beta, c-fos, NCAM-180, PS-1 and APP mRNAs were normally expressed in SAMP8 compared with age-matched other subline that is resistant (SAMR1). The levels of apoE, GR alpha and MR mRNAs in the hippocampus of SAMP8, especially GR alpha, were evidently lower than those in the hippocampus of SAMR1. While bcl-2 alpha, PS-2 and tau mRNA levels of SAMP8 were significantly higher than those of SAMR1. Inflammatory cytokines (IL-1 beta, IL-6, TNF-alpha), alpha 2M and MHCII la antigen mRNAs were not detected in the brain of SAM. The differences of gene expression in the cerebral cortex were less evident than in the hippocampus. The results indicated that some genes abnormally expressed in the AD brain were also found in the brain of SAMP8, which may contribute to its age-related deterioration of learning and memory. Our results also suggested that functional and pathological changes which occurred in the brain of SAMP8 possessed some different aspects in comparison with the AD in consideration of the differences in gene expression.

Aging↗

PubMed1999

Nucleotides 1506-1625 of bovine papillomavirus type 1 genome can enhance DNA packaging by L1/L2 capsids.

We have previously described a DNA-packaging assay using bovine papillomavirus type 1 (BPV-1) virus-like particles (VLPs) and have identified a region of the BPV genome that assists in packaging. In this study, we identify a specific BPV sequence involved in DNA packaging by BPV-1 VLPs. In the initial screening of BPV-1 genomic sequences essential for DNA packaging, we observed that a plasmid with deletions between nucleotides (nt) 948 and 2113 failed to be packaged into BPV-1 VLPs. However, plasmids containing nt 948 to 2113 were efficiently packaged, suggesting that this 1.2-kb fragment contains a packaging enhancement sequence (PES). Further mapping of the BPV-1 genome showed that this packaging sequence lies between nt 1506 and 1625. Furthermore, this packaging sequence is also recognized by HPV6b VLPs, suggesting that a common packaging mechanism may be used by the two papillomavirus types. Given the phylogenetic difference between these two viral types, it is likely that other papillomavirus types may also use the same packaging mechanism. Identification of the PES has allowed a minimal viral genome sequence to be used in the packaging assay, improving the usefulness of the assay in studying the process of papillomavirus DNA encapsidation.

Animals↗

PubMed1999

Brain-derived neurotrophic factor promotes survival and chemoprotection of human neuroblastoma cells.

Brain-derived neurotrophic factor (BDNF) promotes neuronal survival and protection against neuronal damage. We addressed whether BDNF might promote survival and chemoprotection in neuroblastoma (NB) using a drug-sensitive human NB cell line. All-trans-retinoic acid (ATRA) induces a striking phenotypic differentiation of NB1643 cells, and exogenous BDNF treatment promotes survival of these differentiated cells. ATRA induces TRKB expression, and exogenous BDNF stimulates both autophosphorylation of TRKB and induction of the immediate early gene, FOS, in these cells. BDNF mRNA is expressed in NB1643 cells. Because the time course of TRKB induction closely parallels phenotypic differentiation of these cells, it seems probable that ATRA induces differentiation of NB1643 cells by establishing an autocrine loop involving BDNF and TRKB. Exogenous BDNF treatment resulted in a further increase in neurite outgrowth, which again suggests that an autocrine loop is involved in differentiation of NB1643 cells in response to ATRA. We then tested whether BDNF might afford drug resistance in NB and found that BDNF does indeed protect in this NB model against cisplatin, a DNA-damaging agent actually used in the treatment of NB.

Antineoplastic Agents↗

PubMed1999

Structure of human ornithine decarboxylase antizyme 2 gene.

Ornithine decarboxylase antizyme 1, a negative regulator of polyamine levels, acts by destabilizing the first enzyme in the polyamine biosynthetic pathway (ODC) and by inhibiting uptake of polyamines. Recently, a second mammalian antizyme was discovered. Here, we report the genomic organization and chromosomal position of the new human gene, HuAZ2, and make a comparison with its antizyme 1 homologue. Fluorescence in-situ hybridization (FISH) localized the sequences from a bacterial artificial chromosomal (BAC) clone containing HuAZ2 at band 22 of the long arm of chromosome 15. This gene spans at least 15kb and contains five exons. No CAAT or TATA sequences could be identified within 800bp upstream of the translation initiation codon, but the 5'-flanking region contains several putative binding sites for other transcription factors. Transcription initiation of this gene appears to be heterogeneous.

5' Untranslated Regions↗

PubMed1999

Preparation and Characterization of Nanoscopic Organic Semiconductor of Oxovanadium Phthalocyanine.

Nanoscopic particles of oxovanadium phthalocyanine (VOPc, phase-II) with different particle sizes are prepared by aggregation of VOPc molecules in the presence of surfactants for the first time. A size effect in the region of nanoscale was observed in the UV-visible spectra. The photoconductivity of the nanoscopic VOPc is much higher than that of bulk VOPc. Copyright 1999 Academic Press.

Journal Article↗

PubMed1999

Ile-177 and Ser-180 in the S1 segment are critically important in Kv1.1 channel function.

Ile-177 and Ser-180 are conserved residues in the first transmembrane segment (S1) of the Shaker, Shab, Shaw, and Shal subfamilies of voltage-gated K+ channels. Here we report that the mutation of these residues in Kv1.1 to leucine, proline, or arginine abolished the expression of outward potassium currents in Xenopus oocytes. Co-injection of these mutant cRNAs and wild type Kv1.1 cRNA into Xenopus oocytes exerted a potent dominant negative effect resulting in the suppression of Kv1.1-encoded currents. Transient transfection experiments of COS-7 cells revealed that the S1 mutants directed the synthesis of Kv1.1 polypeptides. Quantitative co-immunoprecipitation assays revealed that most of the S1 mutants co-assembled and formed both homo- and heteromultimeric complexes. Furthermore, the mutated polypeptides could reach the plasma membranes of transfected Sol8 cells. We conclude that mutations of Ile-177 and Ser-180 do not interfere with either the assembly of multimeric channel complexes or the targeting of these complexes to the plasma membrane. It is likely that these residues are involved in helix-helix interactions that are critical to the proper functioning of voltage-gated potassium channels.

Amino Acid Sequence↗

PubMed1999

Capillary electrophoresis of some tetracycline antibiotics coupled with reductive fast cyclic voltammetric detection.

The separation and quantitative performance parameters for tetracycline, chlortetracycline and oxytetracycline antibiotics were investigated by capillary zone electrophoresis coupled with fast cyclic voltammetric detection. Optimization of pH and complexation with a boric acid-sodium tetraborate buffer provided good resolution of all compounds. Detection by electrochemical reduction using fast on-line cyclic voltammetric detection with a Hg-film-microm electrode gave detection limits (2 x peak-to-peak baseline noise) of 7 x 10(-7) mol/l for tetracycline and chlortetracycline, and 1.5 x 10(-6) mol/l for oxytetracycline. The influence of electrode material, potential range and scan rate was examined and discussed. Optimal electrochemical detection was obtained at a Hg-film electrode with a waveform that consisted of an initial constant potential of -0.6 V for 200 ms followed by a cyclic voltammetry (CV) scan at 300 V/s from - 0.6 V to a vertex potential of 1.7 V. The analytical signal was obtained by plotting the integrated values of the CV current from each applied waveform as a function of time. The calibration plot (peak areas) for each separated peak was found to be linear over three-orders of magnitude.

Anti-Bacterial Agents↗

PubMed1999

The efficacy of lamotrigine in rapid cycling and non-rapid cycling patients with bipolar disorder.

BACKGROUND: Patients with bipolar disorder (BD) who have rapid cycling features are often treatment refractory. Clear and conclusive evidence regarding effective treatments for this group is not available. METHODS: Patients with diagnoses of refractory bipolar disorder who were currently experiencing manic, mixed, depressive, or hypomanic episodes were treated with lamotrigine as add-on therapy (60 patients) or monotherapy (15 patients). We compared the efficacy of lamotrigine in the 41 rapid cycling and 34 non-rapid cycling patients with BD. RESULTS: Improvement from baseline to last visit was significant among both rapid cycling and non-rapid cycling patients for both depressive and manic symptomatology. For patients entering the study in a depressive episode, improvement in depressive symptomatology was equivalent in the two groups. Among patients entering the study in a manic, mixed, or hypomanic episode, those with rapid cycling improved less in manic symptomatology than did non-rapid cycling patients. Among rapid cycling patients with initial mild-to-moderate manic symptom severity, improvement was comparable to that in non-rapid cycling subjects; however, the subset of rapid cycling patients with severe initial manic symptomatology had little improvement in mania. Rapid cycling patients had earlier onset and more lifetime episodes of mania, depression, and mixed mania. CONCLUSIONS: Lamotrigine was generally effective and well tolerated in this group of previously non-responsive, rapid cycling bipolar patients.

Activity Cycles↗

PubMed1999

Adenylylcyclase increases responsiveness to catecholamine stimulation in transgenic mice.

BACKGROUND: The cellular content of cAMP generated by activation of adenylylcyclase (AC) through the beta-adrenergic receptor (betaAR) is a key determinant of a cell's response to catecholamine stimulation. We tested the hypothesis that increased AC content, independently of betaAR number, increases responsiveness to catecholamine stimulation in vivo. METHODS AND RESULTS: Transgenic mice with cardiac-directed expression of ACVI showed increased transgene AC expression but no change in myocardial betaAR number or G-protein content. When stimulated through the betaAR, cardiac function was increased, and cardiac myocytes showed increased cAMP production. In contrast, basal cAMP and cardiac function were normal, and long-term transgene expression was not associated with abnormal histological findings or deleterious changes in cardiac function. CONCLUSIONS: The amount of AC sets a limit on cardiac beta-adrenergic signaling in vivo, and increased AC, independent of betaAR number and G-protein content, provides a means to regulate cardiac responsiveness to betaAR stimulation. Overexpressing an effector (AC) does not alter transmembrane signaling except when receptors are activated, in contrast to receptor/G-protein overexpression, which yields continuous activation and has detrimental consequences. Our findings establish the importance of AC content in modulating beta-adrenergic signaling in the heart, suggesting a new target for safely increasing cardiac responsiveness to betaAR stimulation.

Adenylyl Cyclases↗

PubMed1999

[Study on the correlation of silicosis with antioxidant and antioxidase].

Plasma vitamin C(P-VC), vitamin E(P-VE) and beta-carotene(P-beta-CAR) contents and the activities of superoxide dismutase(E-SOD), catalase(E-CAT) and glutathione peroxidase (E-GSH-Px) in erythrocyte in 73 silicosis patients and 60 healthy control subjects were measured. The average levels of P-VC, P-VE, P-beta-CAR, E-SOD, E-CAT and E-GSH-Px of patients were significantly lower than those of the controls (P < 0.001). All indexes were correlated to the course, condition and pulmonary function of silicosis patients. The results analyzed by stepwise regression showed that the correlation between course, condition and pulmonary function of patients and P-VE and E-SOD was close. The balance between oxidation and the antioxidation in silicosis patients may be disturbed, and oxygen free radical reaction may be pathologically exacerbated.

Adult↗

PubMed1999

Insulin-like growth factor 1 is required for G2 progression in the estradiol-induced mitotic cycle.

Insulin-like growth factor 1 (IGF1) has been proposed as a "G1-progression factor" and as a mediator of estradiol's (E2) mitogenic effects on the uterus. To test these hypotheses, we compared E2's mitogenic effects on the uteri of Igf1-targeted gene deletion (null) and wild-type littermate mice. The proportion of uterine cells involved in the cell cycle and G1- and S-phase kinetics were not significantly different in wild-type and Igf1-null mice. However, the appearance of E2-induced mitotic figures and cell number increases were profoundly retarded in Igf1-null uterine tissue. There was a significant increase in nuclear DNA concentration in Igf1-null cells, consistent with a G2 arrest. Interestingly, apoptotic cells were also significantly reduced in abundance, and the normal massive apoptotic response to E2 withdrawal was absent in the Igf1-null uterus. These data show that Igf1 is an essential mediator of E2's mitogenic effects, with a critical role not in G1 progression but in G2 progression.

Animals↗

PubMed1999

Calmodulin mediates calcium-dependent activation of the intermediate conductance KCa channel, IKCa1.

Small and intermediate conductance Ca2+-activated K+ channels play a crucial role in hyperpolarizing the membrane potential of excitable and nonexcitable cells. These channels are exquisitely sensitive to cytoplasmic Ca2+, yet their protein-coding regions do not contain consensus Ca2+-binding motifs. We investigated the involvement of an accessory protein in the Ca2+-dependent gating of hIKCa1, a human intermediate conductance channel expressed in peripheral tissues. Cal- modulin was found to interact strongly with the cytoplasmic carboxyl (C)-tail of hIKCa1 in a yeast two-hybrid system. Deletion analyses defined a requirement for the first 62 amino acids of the C-tail, and the binding of calmodulin to this region did not require Ca2+. The C-tail of hSKCa3, a human neuronal small conductance channel, also bound calmodulin, whereas that of a voltage-gated K+ channel, mKv1.3, did not. Calmodulin co-precipitated with the channel in cell lines transfected with hIKCa1, but not with mKv1. 3-transfected lines. A mutant calmodulin, defective in Ca2+ sensing but retaining binding to the channel, dramatically reduced current amplitudes when co-expressed with hIKCa1 in mammalian cells. Co-expression with varying amounts of wild-type and mutant calmodulin resulted in a dominant-negative suppression of current, consistent with four calmodulin molecules being associated with the channel. Taken together, our results suggest that Ca2+-calmodulin-induced conformational changes in all four subunits are necessary for the channel to open.

Amino Acid Sequence↗