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Acetyltransferase
Kinds of Acetyltransferase Terms modified by Acetyltransferase Selected AbstractsGENOTYPE AND ALLELE FREQUENCIES OF N -ACETYLTRANSFERASE 2 AND GLUTATHIONE S -TRANSFERASE IN THE IRANIAN POPULATIONCLINICAL AND EXPERIMENTAL PHARMACOLOGY AND PHYSIOLOGY, Issue 11 2007Anahita Torkaman-Boutorabi SUMMARY 1.,Xenobiotic-metabolizing enzymes constitute an important line of defence against a variety of carcinogens. Many are polymorphic, constituting the basis for the wide interindividual variation in metabolic capacity and possibly a source of variation in the susceptibility to chemical-induced carcinogenesis. The aim of the present study was to determine the frequencies of important allelic variants in the N- acetyltransferase 2 (NAT2) and glutathione S- transferase (GST) genes in the Iranian population and compare them with frequencies in other ethnic populations. 2Genotyping was performed in a total of 229 unrelated healthy subjects (119 men, 110 women) for NAT2 and 170 unrelated healthy subjects (89 men, 81 women) for GST from the general Tehran population. A combination of polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) was applied for typing of NAT2 polymorphisms. Detection of GSTM1 and GSTT1 null alleles was performed simultaneously using a multiplex PCR assay. 3The frequencies of specific NAT2 alleles were 0.299, 0.314, 0.380, 0.007 and 0.000 for *4 (wild-type), *5 (C481T, M1), *6 (G590A, M2), *7 (G857A, M3) and *14 (G191A, M4), respectively. The most prevalent genotypes were NAT2 *5/*6 (29.70%) and *4/*6 (21.40%). The GSTM1 - and GSTT1 -null alleles were detected in 44.7 and 21.2% of subjects, respectively. 4We found that Iranians resemble Indians with regard to allelic frequencies of the tested variants of NAT2. The predominance of slow (49.36%) and intermediate (41.47%) acetylation status compared with wild-type rapid acetylation status (9.17%) in the study group suggests the significant prevalence of the slow acetylator (SA) phenotypes in the Iranian population. Our data confirmed that Iranians are similar to other Caucasian populations in the frequency of both GSTM1 - and GSTT1 -null alleles. [source] Activation of Arylalkylamine N -Acetyltransferase by Phorbol Esters in Bovine Pinealocytes Suggests a Novel Regulatory Pathway in Melatonin SynthesisJOURNAL OF NEUROENDOCRINOLOGY, Issue 9 2004C. Schomerus Abstract In all mammalian species investigated, noradrenaline activates a ,-adrenoceptor/cAMP/protein kinase A-dependent mechanism to switch on arylalkylamine N -acetyltransferase and melatonin biosynthesis in the pineal gland. Other compounds which are known to influence the melatonin-generating system are phorbol esters. The effect of phorbol esters on regulation of melatonin synthesis has been mainly investigated in rat pinealocytes. In these cells, phorbol esters do not increase cAMP levels and arylalkylamine N -acetyltransferase on their own; however, phorbol esters potentiate the effects on cAMP and AANAT activity induced upon ,-adrenoceptor stimulation. In the present study, we investigated the effect of phorbol esters on the regulation of melatonin synthesis in bovine pinealocytes. We show that, in these cells, the phorbol esters 4,-phorbol 12-myristate 13-acetate (PMA) or phorbol 12,13-dibutyrate have a direct stimulatory effect and induced 4,10-fold increases in AANAT protein levels, AANAT activity and melatonin production. The extent of these effects was similar to those induced by noradrenaline. Notably, responses to PMA were not accompanied by increases in cAMP levels. Northern blot analysis showed that Aanat mRNA levels did not change upon PMA treatment indicating that phorbol esters control AANAT at a post-transcriptional level. The effects on AANAT and melatonin production were reduced by use of protein kinase C inhibitors, but not by blockade of the cyclic AMP/protein kinase A pathway. Our results point towards a novel mechanism in the regulation of melatonin production that is cAMP-independent and involves protein kinase C. The study is of particular interest because regulation of melatonin biosynthesis in bovines may resemble that in primates more closely than that in rodents. [source] Synthesis of Benzamides Related to Anacardic Acid and Their Histone Acetyltransferase (HAT) Inhibitory ActivitiesCHEMMEDCHEM, Issue 9 2008Abstract A group of benzamides related to anacardic acid amide CTPB with alkyl chains of defined length were prepared by a five-step sequence starting from 2,6-dihydroxybenzoic acid, and their activities were compared with those reported for the HAT inhibitor anacardic acid (AA). The subset of 4-cyano-3-trifluoromethylphenylbenzamides with shorter chains exhibited activities similar to that of AA, as they behaved as human p300 inhibitors, induced a decrease in histone acetylation levels in immortalized HEK cells, and counteracted the action of the HDAC inhibitor SAHA in MCF7 breast cancer cells. Moreover, an analogue with the shortest alkyl chain induced significant apoptosis at 50,,M in U937 leukemia cells. [source] What a Role did Histidine Residue Play in Arylamine N -Acetyltransferase 2 Acetylation?CHINESE JOURNAL OF CHEMISTRY, Issue 10 2006A Quantum Chemistry Study Abstract Arylamine N -acetyltransferases (NATs, EC 2.3.1.5) catalyze an acetyl group transfer from acetyl coenzyme A (AcCoA) to primary arylamines and play a very important role in the metabolism and bioactivation of drugs and carcinogens. Experiments revealed that His-107 was likely the residues responsible for mediating acetyl transfer. The full catalytic mechanism of acetylation process has been examined by density functional theory. The results indicate that, if the acetyl group is directly transferred from the donor, p -nitrophenyl acetate, to the acceptor, cysteine, the high activation energy will be a great hindrance. These energies have dropped in a little range of 20,25 kJ/mol when His-107 assisted the transfer process. However, when protonated His-107 mediated the reaction, the activation energies have been dropped about 73,85 kJ/mol. Our calculations strongly supported an enzyme acetylation mechanism that experiences a thiolate-imidazolium pair, and verified the presumption from experiments. [source] Apparent mitochondrial asymmetry in Xenopus eggsDEVELOPMENTAL DYNAMICS, Issue 4 2003Natalia Volodina Abstract Cell polarity is manifest along the animal/vegetal axis in eggs of the frog, Xenopus laevis. Along this axis, maternal cytoplasmic components are asymmetrically distributed and are thought to underlie specification of distinct cell fates. To ascertain the molecular identities of such cytoplasmic components, we have used a monoclonal antibody that specifically stains the vegetal hemisphere of Xenopus eggs. The antigenic protein Vp67 (vegetal protein of 67 kDa) was identified through purification and cloning as a Xenopus homolog of the mitochondrial protein dihydrolipoamide acetyltransferase, a component of the pyruvate dehydrogenase complex. The identification of Vp67 as a mitochondrial protein could indicate that populations of mitochondria are asymmetrically distributed in Xenopus eggs. Developmental Dynamics 226:654,662, © 2003 Wiley-Liss, Inc. [source] Prediction of polymorphic N -acetylation of new drug candidates by correlation with human NAT1 and NAT2DRUG DEVELOPMENT RESEARCH, Issue 1 2002Katalin Jemnitz Abstract Due to interindividual variation in N -acetyltransferase 2 (NAT2) activity, pharmaceutical companies face the problem of polymorphic metabolism in drugs that are metabolized mainly or exclusively by this enzyme. An in vitro method has been developed to predict in vivo polymorphic N -acetylation at an early stage of drug development. Two new type 5H-2,3-benzodiazepine derivatives, Nerisopam (NER) with anxiolytic activity and GYKI47261 with antiepileptic activity, are metabolized mainly by N -acetylation in the rat and human. The selectivity of human N -acetyltransferases (NAT1,2) to form the acetylated metabolites has been investigated by correlation analysis. Twelve human liver samples were characterized for NAT1 and NAT2 phenotype based on their enzyme activity toward two selective NAT1 (p -aminobenzoic acid, PABA; p -aminosalicylic acid, PAS) and two selective NAT2 (sulfamethazine, SMZ; procainamide, PROC) substrates. Significant correlation was found between enzyme activities NAT1PABA/NAT1PAS and NAT2SMZ/NAT2PROC, respectively, and no correlation was observed comparing enzyme activities toward NAT1PABA/NAT2PROC. Enzyme activities using NER and GYKI 47261 as substrates were compared to activities obtained with NAT1 and NAT2 selective substrates, and the correlation coefficients were calculated. Good correlation was established between the rates of acetylation of the two drugs and that of the NAT2 selective substrate (NER/NAT2SMZ, r2=0.91, GYKI 47261/NAT2SMZ, r2=0.91). In contrast, no correlation was found between the rate of conjugation of the drugs and that of NAT1 selective substrate (NER/NAT1PABA, r2=0.022, GYKI 47261/NAT1PABA, r2=0.0004), suggesting polymorphic in vivo metabolism, since both drugs are acetylated preferably by NAT2. According to our results, correlation analysis based on in vitro acetylation activity may be used to predict in vivo polymorphic metabolism. Drug Dev. Res. 56:17,22, 2002. © 2002 Wiley-Liss, Inc. [source] Association of prostate cancer with rapid N -acetyltransferase 1 (NAT1*10) in combination with slow N -acetyltransferase 2 acetylator genotypes in a pilot case-control studyENVIRONMENTAL AND MOLECULAR MUTAGENESIS, Issue 3 2002David W. Hein Abstract N -acetyltransferase-1 (NAT1) and N -acetyltransferase-2 (NAT2) are important in the metabolism of aromatic and heterocyclic amine carcinogens that induce prostate tumors in the rat. We investigated the association of genetic polymorphisms in NAT1 and NAT2, alone and in combination, with human prostate cancer. Incident prostate cancer cases and controls in a hospital-based case-control study were frequency-matched for age, race, and referral pattern. The frequency of slow acetylator NAT1 genotypes (NAT1*14, *15, *17) was 5.8% in controls but absent in cases. In contrast, in comparison with all other NAT1 genotypes the putative rapid acetylator NAT1 genotype (NAT1*10) was significantly higher in prostate cancer cases than controls (OR, 2.17; 95% CI, 1.08,4.33; P = 0.03). Combinations of NAT1*10 with NAT2 slow acetylator genotypes (OR, 5.08; 95% CI, 1.56,16.5; P = 0.008) or with NAT2 very slow (homozygous NAT2*5) acetylator genotypes (OR, 7.50; 95% CI, 1.55,15.4; P = 0.016) further increased prostate cancer risk. The results of this small pilot study suggest increased susceptibility to prostate cancer for subjects with combinations of NAT1*10 and slow (particularly very slow) NAT2 acetylator genotypes. This finding should be investigated further in larger cohorts and in other ethnic populations. Environ. Mol. Mutagen. 40:161,167, 2002. © 2002 Wiley-Liss, Inc. [source] Expression of cell fate determinants and plastic changes after neurotoxic lesion of adult mice spinal cord by cholera toxin-B saporinEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 8 2010Rosario Gulino Abstract Recent studies have attempted to repair the damaged spinal cord (SC) by stimulating neurogenesis or neuroplasticity. Sonic hedgehog (Shh), Notch-1 and Numb are involved in the stem cell functioning. Additionally, Notch-1 has a role as modulator of synaptic plasticity. However, little is known about the role of these proteins in the adult SC after removal of motoneurons. In this study, we have injected cholera toxin-B saporin into the gastrocnemius muscle to induce a depletion of motoneurons within the lumbar SC of adult mice, and analysed the expression of choline acetyltransferase (ChAT), Synapsin-I, Shh, Notch-1 and Numb proteins. The functional outcome of the lesion was monitored by grid walk and rotarod tasks. The neurotoxin lesion determined a motoneuron depletion and a transient decrease of ChAT, Synapsin-I, Shh and Numb levels in the lumbar SC. ChAT was associated with Synapsin-I expression and motor performance at 1 week but not 1 month after lesion, suggesting that the recovery of locomotion could depend on synaptic plasticity, at least in an early phase. Shh and Notch-1 were associated with Synapsin-I levels, suggesting a role in modulating synaptic plasticity. Numb expression also appeared reduced after lesion and linked to motor performance. Moreover, unlike other lesion models, we observed glial reaction but no evidence of cell proliferation within the depleted SC. Given the mentioned roles of Shh, Notch-1 and Numb, we believe that an in vivo manipulation of their signalling after lesion could represent a suitable way to improve functional recovery by modulating synaptic plasticity and/or neurogenesis. [source] Expression of PRiMA in the mouse brain: membrane anchoring and accumulation of ,tailed' acetylcholinesteraseEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 7 2003Noël A. Perrier Abstract We analysed the expression of PRiMA (proline-rich membrane anchor), the membrane anchor of acetylcholinesterase (AChE), by in situ hybridization in the mouse brain. We compared the pattern of PRiMA transcripts with that of AChE transcripts, as well as those of choline acetyltransferase and M1 muscarinic receptors which are considered pre- and postsynaptic cholinergic markers. We also analysed cholinesterase activity and its molecular forms in several brain structures. The results suggest that PRiMA expression is predominantly or exclusively related to the cholinergic system and that anchoring of cholinesterases to cell membranes by PRiMA represents a limiting factor for production of the AChE tailed splice variant (AChET),PRiMA complex, which represents the major AChE component in the brain. This enzyme species is mostly associated with cholinergic neurons because the pattern of PRiMA mRNA expression largely coincides with that of ChAT. We also show that, in both mouse and human, PRiMA proteins exist as two alternative splice variants which differ in their cytoplasmic regions. [source] Dedifferentiation of intrinsic response properties of motoneurons in organotypic cultures of the spinal cord of the adult turtleEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 7 2000Jean-François Perrier Abstract Explant cultures from the spinal cord of adult turtles were established and used to study the sensitivity of the intrinsic response properties of motoneurons to the changes in connectivity and milieu imposed by isolation in culture. Transverse sections 700 ,m thick were explanted on cover slips and maintained in roller-tube cultures in medium containing serum and the growth factors brain-derived neurotrophin factor (BDNF), neurotrophin-3 (NT3), glial cell line-derived neurotrophic factor (GDNF) and ciliary neurotrophic factor (CNTF). The gross morphology of acute sections was maintained after 4 weeks in culture. Cell bodies of motoneurons remained stainable in fixed cultures with an antibody against choline acetyltransferase (ChAT) throughout the culture period. During culture, motoneurons maintained stable resting membrane potentials and were contacted by functional synapses. The ability to generate action potentials was also preserved as was delayed inward rectification and generation of calcium spikes in the presence of tetra-ethyl ammonium (TEA). In response to depolarization, however, motoneurons presented strong outward rectification, and only 41% of the cells recorded from maintained the ability to fire repetitively. By the second week in culture, a fraction of motoneurons displayed fast and slow transient outward rectification and low-threshold calcium spikes, features not seen in turtle motoneurons in acute slices. On the other hand, properties mediated by L-type Ca2+ channels disappeared during the first few days in culture. Our observations show that the phenotypical intrinsic response properties of mature spinal motoneurons are modified in explant cultures. The properties acquired resemble the properties in juvenile motoneurons in several species of terrestrial vertebrates. [source] Overexpression of spermidine/spermine N1 -acetyltransferase in transgenic mice protects the animals from kainate-induced toxicityEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 2 2000Kyllikki Kaasinen Abstract We recently generated a transgenic mouse line with activated polyamine catabolism through overexpression of spermidine/spermine N1 -acetyltransferase (SSAT). A detailed analysis of brain polyamine concentrations indicated that all brain regions of these animals showed distinct signs of activated polyamine catabolism, e.g. overaccumulation of putrescine (three- to 17-fold), appearance of N1 -acetylspermidine and decreases in spermidine concentrations. In situ hybridization analyses revealed a marked overexpression of SSAT-specific mRNA all over the brain tissue of the transgenic animals. The transgenic animals appeared to tolerate subcutaneous injections of high-dose kainate substantially better as their overall mortality was less than 50% of that of their syngenic littermates. We used the expression of glial fibrillary acidic protein (GFAP) as a marker of brain injury in response to kainate. In situ hybridization analysis with GFAP oligonucleotide up to 7 days after the administration of sublethal kainate doses showed reduced GFAP expression in transgenic animals in comparison with their non-transgenic littermates. This difference was especially striking in the cerebral cortex of the transgenic mice where the exposure to kainate hardly induced GFAP expression. The treatment with kainate likewise resulted in loss of the hippocampal (CA3) neurons in non-transgenic but not transgenic animals. These results support our earlier findings indicating that elevated concentrations of brain putrescine, irrespective whether derived from an overexpression of ornithine decarboxylase, or as shown here, from an overexpression of SSAT, play in all likelihood a neuroprotective role in brain injury. [source] Transcriptional regulation of tumor necrosis factor-, in keratinocytes mediated by interleukin-1, and tumor necrosis factor-,EXPERIMENTAL DERMATOLOGY, Issue 6 2002S. Lisby Abstract: Irritant contact dermatitis (ICD) is an inflammatory skin reaction in which cytokines are thought to play a crucial role. In particular, tumor necrosis factor-, (TNF-,) has been implicated in the mechanism of this reaction. We report that interleukin-1, (IL-1,) that has been reported up-regulated in many inflammatory skin conditions is capable of increasing TNF-, mRNA and protein expression in murine keratinocytes. Furthermore, we show that TNF-, is capable of up-regulating itself in keratinocytes most likely in an autocrine manner. The signalling mechanisms involved in both IL-1,- and TNF-,-mediated regulation of TNF-, are critically dependent upon protein kinase C (PKC), as demonstrated by blocking studies using protein kinase inhibitors. Furthermore, the increase in TNF-, mRNA expression seen after stimulation with rTNF-, and rIL-1, involved increased transcription of TNF-, mRNA. This was demonstrated in a chloramphenicol acetyltransferase (CAT) assay using a CAT-construct containing the full-length TNF-, promoter. These observations support the notion of keratinocytes functioning as an amplifier of pro-inflammatory cytokine generation in the epidermis during ICD and other inflammatory skin conditions. [source] Cloning of Xenopus orthologs of Ctf7/Eco1 acetyltransferase and initial characterization of XEco2FEBS JOURNAL, Issue 24 2008Masatoshi Takagi Sister chromatid cohesion is important for the correct alignment and segregation of chromosomes during cell division. Although the cohesin complex has been shown to play a physical role in holding sister chromatids together, its loading onto chromatin is not sufficient for the establishment of sister chromatid cohesion. The activity of the cohesin complex must be turned on by Ctf7/Eco1 acetyltransferase at the replication forks as the result of a specific mechanism. To dissect this mechanism in the well established in vitro system based on the use of Xenopus egg extracts, we cloned two Xenopus orthologs of Ctf7/Eco1 acetyltransferase, XEco1 and XEco2. Both proteins share a domain structure with known members of Ctf7/Eco1 family proteins. Moreover, biochemical analysis showed that XEco2 exhibited acetyltransferase activity. We raised a specific antibody against XEco2 and used it to further characterize XEco2. In tissue culture cells, XEco2 gradually accumulated in nuclei through the S phase. In nuclei formed in egg extract, XEco2 was loaded into the chromatin at a constant level in a manner sensitive to geminin, an inhibitor of the pre-replication complex assembly, but insensitive to aphidicolin, an inhibitor of DNA polymerases. In both systems, no specific localization was observed during mitosis. In XEco2-depleted egg extracts, DNA replication occurred with normal kinetics and efficiency, and the condensation and sister chromatid cohesion of subsequently formed mitotic chromosomes was unaffected. These observations will serve as a platform for elucidating the molecular function of Ctf7/Eco1 acetyltransferase in the establishment of sister chromatid cohesion in future studies, in which XEco1 and XEco2 should be dissected in parallel. [source] Acetyl-CoA:1- O -alkyl- sn -glycero-3-phosphocholine acetyltransferase (lyso-PAF AT) activity in cortical and medullary human renal tissueFEBS JOURNAL, Issue 14 2003Tzortzis N Nomikos Platelet-activating factor (PAF) is one of the most potent inflammatory mediators. It is biosynthesized by either the de novo biosynthesis of glyceryl ether lipids or by remodeling of membrane phospholipids. PAF is synthesized and catabolized by various renal cells and tissues and exerts a wide range of biological activities on renal tissue suggesting a potential role during renal injury. The aim of this study was to identify whether cortex and medulla of human kidney contain the acetyl-CoA:1- O -alkyl- sn -glycero-3-phosphocholine acetyltransferase (lyso-PAF AT) activity which catalyses the last step of the remodeling biosynthetic route of PAF and is activated in inflammatory conditions. Cortex and medulla were obtained from nephrectomized patients with adenocarcinoma and the enzymatic activity was determined by a trichloroacetic acid precipitation method. Lyso-PAF AT activity was detected in both cortex and medulla and distributed among the membrane subcellular fractions. No statistical differences between the specific activity of cortical and medullary lyso-PAF AT was found. Both cortical and medullary microsomal lyso-PAF ATs share similar biochemical properties indicating common cellular sources. [source] ORF6 from the clavulanic acid gene cluster of Streptomyces clavuligerus has ornithine acetyltransferase activityFEBS JOURNAL, Issue 8 2002Nadia J. Kershaw The clinically used beta-lactamase inhibitor clavulanic acid is produced by fermentation of Streptomyces clavuligerus. The orf6 gene of the clavulanic acid biosynthetic gene cluster in S. clavuligerus encodes a protein that shows sequence homology to ornithine acetyltransferase (OAT), the fifth enzyme of the arginine biosynthetic pathway. Orf6 was overexpressed in Escherichia coli (at ,,15% of total soluble protein by SDS/PAGE analysis) indicating it was not toxic to the host cells. The recombinant protein was purified (to >,95% purity) by a one-step technique. Like other OATs it was synthesized as a precursor protein which underwent autocatalytic internal cleavage in E. coli to generate , and , subunits. Cleavage was shown to occur between the alanine and threonine residues in a KGXGMXXPX--(M/L)AT (M/L)L motif conserved within all identified OAT sequences. Gel filtration and native electrophoresis analyses implied that the ORF6 protein was an ,2,2 heterotetramer and direct evidence for this came from mass spectrometric analyses. Although anomalous migration of the , subunit was observed by standard SDS/PAGE analysis, which indicated the presence of two bands (as previously observed for other OATs), mass spectrometric analyses did not reveal any evidence for post-translational modification of the , subunit. Extended denaturation with SDS before PAGE resulted in observation of a single major , subunit band. Purified ORF6 was able to catalyse the reversible transfer of an acetyl group from N -acetylornithine to glutamate, but not the formation of N -acetylglutamate from glutamate and acetyl-coenzyme A, nor (detectably) the hydrolysis of N -acetylornithine. Mass spectrometry also revealed the reaction proceeds via acetylation of the , subunit. [source] RBP2 is an MRG15 complex component and down-regulates intragenic histone H3 lysine 4 methylationGENES TO CELLS, Issue 6 2007Tomohiro Hayakawa MRG15 is a conserved chromodomain protein that associates with histone deacetylases (HDACs) and Tip60-containing histone acetyltransferase (HAT) complexes. Here we further characterize MRG15-containing complexes and show a functional link between MRG15 and histone H3K4 demethylase activity in mammalian cells. MRG15 was predominantly localized to discrete nuclear subdomains enriched for Ser2 -phosphorylated RNA polymerase II, suggesting it is involved specifically with active transcription. Protein analysis of the MRG15-containing complexes led to the identification of RBP2, a JmjC domain-containing protein. Remarkably, over-expression of RBP2 greatly reduced the H3K4 methylation in culture human cells in vivo, and recombinant RBP2 efficiently removed H3K4 methylation of histone tails in vitro. Knockdown of RBP2 resulted in increased H3K4 methylation levels within transcribed regions of active genes. Our findings demonstrate that RBP2 associated with MRG15 complex to maintain reduced H3K4 methylation at transcribed regions, which may ensure the transcriptional elongation state. [source] Genetic polymorphism of N -acetyltransferase 2 in the susceptibility to laryngeal squamous cell carcinomaHEAD & NECK: JOURNAL FOR THE SCIENCES & SPECIALTIES OF THE HEAD AND NECK, Issue 12 2005Murat Ünal MD Abstract Background. The purpose of this study was to investigate whether polymorphism of N -acetyltransferase 2 (NAT2) genotypes are associated with the risk of laryngeal squamous cell carcinoma (SCC). Methods. The study group consisted of 45 white patients with laryngeal SCC (42 men, with a mean age of 54 years [range, 37,70 years] and three women, with a mean age of 47 years [range, 32,55 years]) and 104 control subjects (68 men and 36 women; mean age, 50 years; range, 28,73 years). All of the patients were primarily treated with surgical intervention. Blood samples (5 mL) were obtained before surgery or from the patients under follow-up to 5 years after surgery (mean follow-up, 27 months; range, 6,48 months). DNA was extracted from the lymphocytes by high pure template preparation kit. NAT2*5A, NAT2*6A, NAT2*7A/B, and NAT2*14A were detected by use of LightCycler- NAT2 mutation detection kit by real-time polymerase chain reaction with Light Cycler instruments. The association between NAT2 polymorphisms and laryngeal SCC was prospectively modeled through multivariate logistic regression analysis. Results. We found that the risk of laryngeal SCC was 7.3-fold higher in individuals with NAT2*5 mutant allele, 3.8-fold higher in subjects with NAT2*6 heterozygote allele, and 38.3-fold higher in NAT2*6 mutant allele. We also found that individuals with NAT2*7 heterozygote allele had a 0.2-fold less risk for the development of laryngeal SCC (p = .018). Conclusion. In this population, patients with NAT2*5 mutant and *6 heterozygous and mutant genotypes had a significantly higher risk for development of laryngeal SCC. © 2005 Wiley Periodicals, Inc. Head Neck27: XXX,XXX, 2005 [source] Ontogenetic effects of MAO-A inhibition on rat pineal n -acetylserotonin and melatonin during the first month of neonatal lifeHUMAN PSYCHOPHARMACOLOGY: CLINICAL AND EXPERIMENTAL, Issue 8 2000Gregory F Oxenkrug Abstract Inhibitors of monoamine oxidase A (MAO-A) but not MAO-B stimulate the activity of pineal serotonin N -acetyltransferase (AANAT) in the adult rat pineal leading to increased formation of N -acetyl serotonin (NAS) and melatonin (MEL). The pineal gland of the neonatal rat has AANAT activity, but the second enzyme in melatonin biosynthesis, HIOMT (hydroxyindole- O -methyltransferase) converting NAS to MEL, is absent during the first week of neonatal life. In this study we examined the effects of acute clorgyline treatment in vitro and in vivo, on pineal indoles over the first month of neonatal life. The results show that clorgyline stimulates NAS production by pineal both in vitro and in vivo from day five on with a marked increase between day 14 and day 21. In contrast, MEL is not increased until day 21, with a sharp rise thereafter. Copyright © 2000 John Wiley & Sons, Ltd. [source] FDB2 encodes a member of the arylamine N -acetyltransferase family and is necessary for biotransformation of benzoxazolinones by Fusarium verticillioidesJOURNAL OF APPLIED MICROBIOLOGY, Issue 2 2009A.E. Glenn Abstract Aims:, To clone and characterize genes from the mycotoxigenic fungus, Fusarium verticillioides, which are associated with its ability to biotransform allelopathic benzoxazolinones produced by maize, wheat, and rye. Methods and Results:, Suppression subtractive hybridization identified F. verticillioides genes up-regulated in response to 2-benzoxazolinone (BOA), including a cluster of genes along chromosome 3. One of these genes, putatively encoding an arylamine N -acetyltransferase (NAT), was highly represented in the subtracted library and was of particular interest since previous analyses identified the FDB2 locus as possibly encoding transferase activity. The gene was subcloned and complemented a natural fdb2 mutant. Conversely, disruption of the gene eliminated the ability of F. verticillioides to metabolize BOA. Other genes in the cluster also were assessed using a complementation assay. Metabolic profiles of fdb2 mutants suggest that minor acylation activity occurred independently of the NAT activity encoded by FDB2. Conclusions:, The previously defined FDB2 locus was functionally associated with the gene encoding putative NAT activity, and the FDB2 gene was essential for biotransformation of BOA. The flanking gene FDB3 encodes a putative Zn(II)2Cys6 transcription factor and contributes to efficient BOA biotransformation but was not essential. Significance and Impact of the Study:, Biotransformation of benzoxazolinones by F. verticillioides may enhance its ecological fitness in maize field environments and our results provide greater understanding of the genes that modulate the biotransformation process. Additionally, this is the first homologue of the NAT gene family to be characterized in a filamentous fungus. [source] TCDD suppresses insulin-responsive glucose transporter (GLUT-4) gene expression through C/EBP nuclear transcription factors in 3T3-L1 adipocytesJOURNAL OF BIOCHEMICAL AND MOLECULAR TOXICOLOGY, Issue 2 2006Phillip Chin-Chen Liu Abstract TCDD is known to reduce significantly the level of the functionally active form of glucose transporter type 4 (GLUT4) in vivo in adipose tissue and muscles. To study the mechanistic basis of this phenomenon, we conducted transient transfection and DNA deletion analysis in 3T3-L1 cells using chloramphenicol acetyltransferase (CAT) reporter plasmids containing the GLUT4 promoter joined to the bacterial CAT. It was found that in transfected control samples, CAT activity was significantly higher in cells transfected with p469CAT and p273CAT than those with p78CAT, indicating that the region between ,78 and ,273 contained elements that play major roles in transactivation of this gene. Treatment with TCDD decreased CAT activity with p469CAT and p273CAT, but not with p78CAT, indicating the same region to contain the element(s) affected by TCDD. A gel-shift (EMSA) analysis result indicated that TCDD shows the profound effect only on the nuclear proteins binding to the [32P]-labeled probe containing C/EBP response element equivalent of the ,265 to ,242 stretch of the GLUT4 promoter. The results of supershift analysis showed that TCDD caused a decrease in the tier of C/EBP, and an increase in that of C/EBP, among the proteins bound to this C/EBP response element. We studied the effect of TCDD in cells overexpressing either C/EBP,, C/EBP,, or C/EBP, through transient transfection of p273CAT or p469CAT. The results clearly showed that the effect of TCDD to suppress the CAT activity of p273 or p469 disappeared in those cells overexpressing C/EBP, or C/EBP,. These results implicate the C/EBP proteins to be the main mediator of suppressive action of TCDD on GLUT4 gene expression in 3T3-L1 cells. © 2006 Wiley Periodicals, Inc. J Biochem Mol Toxicol 20:79,87, 2006; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.20120 [source] The polymine spermine regulates osteogenic differentiation in adipose stem cells,JOURNAL OF CELLULAR AND MOLECULAR MEDICINE, Issue 5a 2008G.S. Tjabringa Abstract For bone tissue engineering, it is important that mesenchymal stem cells (MSCs) differentiate into osteoblasts. To develop a method for differentiation of adipose tissue-derived mesenchymal stem cells (AT-MSCs) along the osteogenic lineage, we studied the effect of polyamines, which are organic cations implicated in bone growth and development, on differentiation of AT-MSCs. Treatment of goat-derived AT-MSCs with 1,25-dihydroxyvitamin-D3 (1,25(OH)2D3), which stimulates osteogenic differentiation, for 7 days induced gene expression of the polyamine-modulated transcription factor-1 (PMF-1) and spermidine/spermine N (1)-acetyltransferase (SSAT), which are both involved in polyamine metabolism, suggesting that polyamines are involved in osteogenic differentiation of AT-MSCs. Furthermore, treatment of AT-MSCs with the polyamine spermine-regulated gene expression of runx-2, a transcription factor involved in early stages of osteogenic differentiation, and that of osteopontin, a bone matrix protein expressed in later stages of osteogenic differentiation. Runx-2 gene expression was increased 4 and 14 days after a short 30 min. treatment with spermine, while osteopontin gene expression was only increased 4 days after spermine treatment. Finally, alkaline phosphatase activity, which is intimately involved in the formation of extracellular matrix of bone, was increased 4 weeks after the 30 min.-spermine treatment of AT-MSCs. In conclusion, this study shows for the first time that the polyamine spermine regulates differentiation of AT-MSCs along the osteogenic lineage, which can be used as a new method for differentiation of AT-MSCs along the osteogenic lineage. Therefore, polyamines may constitute a promising tool for bone tissue engineering approaches using AT-MSCs, such as a one-step surgical procedure for spinal interbody fusion. [source] Polyamine metabolism and cancerJOURNAL OF CELLULAR AND MOLECULAR MEDICINE, Issue 2 2003Thresia Thomas Abstract Polyamines are aliphatic cations present in all cells. In normal cells, polyamine levels are intricately controlled by biosynthetic and catabolic enzymes. The biosynthetic enzymes are ornithine decarboxylase, S-adenosylmethionine decarboxylase, spermidine synthase, and spermine synthase. The catabolic enzymes include spermidine/spermine acetyltransferase, flavin containing polyamine oxidase, copper containing diamine oxidase, and possibly other amine oxidases. Multiple abnormalities in the control of polyamine metabolism and uptake might be responsible for increased levels of polyamines in cancer cells as compared to that of normal cells. This review is designed to look at the current research in polyamine biosynthesis, catabolism, and transport pathways, enumerate the functions of polyamines, and assess the potential for using polyamine metabolism or function as targets for cancer therapy. [source] GSTT1 and GSTM1 gene deletions are not associated with hepatotoxicity caused by antitubercular drugsJOURNAL OF CLINICAL PHARMACY & THERAPEUTICS, Issue 4 2010S. Chatterjee MD Summary Background and objective:, Susceptibility to antitubercular drug (ATD)-induced hepatotoxicity may be genetically mediated, with variant alleles of genes such as N -acetyltransferase (NAT2) and CYP2E1 reported as risk factors. Two studies of Asian populations have reported that GSTM1*0/*0 (null) genotype was a likely predictor of hepatotoxicity, whereas another of a Caucasian population implicated GSTT1*0/*0. We undertook a prospective case,control study to investigate whether GSTM*0/*0 and GSTT1*0/*0 were risk factors for ATD-induced hepatotoxicity. Methods:, Pulmonary tuberculosis patients on isoniazid, rifampicin and pyrazinamide who developed hepatotoxicity using defined criteria were prospectively identified. These cases were then matched with at least one control subject on the same drugs but without hepatotoxicity. Genotyping for GSTM1 and GSTT1 was performed by multiplex PCR on genomic DNA. The odds ratios for the frequency of specific GSTM1 and GSTT1 homozygotes in the case and control subjects were calculated to test for association between the genotypes and hepatotoxicity. Results and discussion:, Hundred and fifty-one subjects (51 cases, 100 controls) were enrolled. Odds ratio for GSTM1 null genotype was 1·00 (95% CI 0·51,1·97) and GSTT1 null was 2·02 (95% CI 0·39,10·39), respectively, showing that these genotypes are not associated with hepatotoxicity. Conclusion:,GSTM1 *0/*0 or GSTT1 *0/*0 or both null genotypes, do not appear to be associated with ATD-induced hepatotoxicity in our Indian population. [source] Comparison of genetic polymorphisms of the NAT2 gene between Korean and four other ethnic groupsJOURNAL OF CLINICAL PHARMACY & THERAPEUTICS, Issue 6 2009T. S. Kang MS Summary Background and objective:,N -acetyltransferase 2 (NAT2) is responsible for the acetylation of numerous drugs and in the transformation of aromatic and heterocyclinc amines into carcinogenic intermediates. Polymorphism of NAT2 may contribute to interindividual variability in such acetylation. The aim of this study was to determine the allele frequencies of polymorphisms of the NAT2 gene, analyse linkage disequilibrium (LD) block and haplotypes in Koreans and compare them with those of other ethnic groups. Methods:, We analysed genetic polymorphisms in all functional promoter and exons of the NAT2 gene by direct sequencing of genomic DNA from 192 healthy Korean subjects. The LD and haplotype blocks of these subjects were constructed from genotype data using an expectation,maximization algorithm. We compared these allele frequencies, LD block and haplotype structure with those of other ethnic groups registered on the International HapMap database. Results and discussion:, We identified 33 polymorphisms including six novel single nucleotide polymorphisms, ,10778T>C, ,10777A>G, ,10351A>G, ,10199C>T and ,10104G>T in promoter and 578C>T in exon2 (T193M) in the Korean subjects tested. All allele frequencies reported in the Koreans were similar to those of Asians except for one allele (rs4345600, ,9306A>G), whereas African and European groups had different frequencies in exon2. The haplotype structure and LD block among the five groups also revealed significant differences. Conclusion:, Ethnic differences in the NAT2 genotype frequencies may be one of the important factors explaining variability in cancer incidence and drug toxicity. Our observations could be useful in assessing the susceptibility of different populations to cancer and contribute to better predictions of the pharmacokinetics and pharmacodynamics of drugs that are metabolized by NAT2, in different populations. [source] Medullary motor neurones associated with drinking behaviour of Japanese eelsJOURNAL OF FISH BIOLOGY, Issue 1 2003T. Mukuda A fluorescent dye, Evans blue (EB), was injected into the following seven drinking-associated muscles of the Japanese eel Anguilla japonica: the sternohyoid, third branchial, fourth branchial, opercular, pharyngeal, upper oesophageal sphincter and oesophageal body muscles. The sternohyoid muscle promotes ,ingestion', and the remaining muscles contribute to ,swallowing'. All neurones stained by EB were located ipsilaterally in the caudal medulla oblongata (MO) of the Japanese eel. Neurones projecting into the sternohyoid muscle were identified as those in the spino-occipital motor nucleus (NSO), and neurones projecting into the remaining muscles as those in the glossopharyngeal,vagal motor complex (GVC). Within the GVC, the neuronal arrangement was topological, and hence, ,swallowing' will be completed if the GVC neurones ,fire' progressively from rostral to caudal. These neurones in the NSO and GVC may use acetylcholine (ACh) as a neurotransmitter, as the EB-positive neurones in both nuclei were immunoreactive against anticholine acetyltransferase (anti-ChAT) antibody. Besides the MO, some somata in a ganglion of the vagal nerve were also stained by EB injected into the pharyngeal, the upper oesophageal sphincter and the oesophageal body muscles. The localization and the shape of the somata suggest that they are sensory neurones. These sensory neurones were not ChAT-immunoreactive. Combining these results, based on a model for ,swallowing' in mammals, a plausible model for central organization of ,drinking' in the Japanese eel is proposed, which suggests that ,drinking' in the fishes is regulated by the neuronal circuit for ,swallowing' in mammals. [source] Modulation of Aanat gene transcription in the rat pineal glandJOURNAL OF NEUROCHEMISTRY, Issue 2 2010Anthony K. Ho Abstract The main function of the rat pineal gland is to transform the circadian rhythm generated in the suprachiasmatic nucleus into a rhythmic signal of circulating melatonin characterized by a large nocturnal increase that closely reflects the duration of night period. This is achieved through the tight coupling between environmental lighting and the expression of arylalkylamine- N -acetyltransferase, the rhythm-controlling enzyme in melatonin synthesis. The initiation of Aanat transcription at night is controlled largely by the norepinephrine-stimulated phosphorylation of cAMP response element-binding protein by protein kinase A. However, to accurately reflect the duration of darkness, additional signaling mechanisms also participate to fine-tune the temporal profile of adrenergic-induced Aanat transcription. Here, we reviewed some of these signaling mechanisms, with emphasis on the more recent findings. These signaling mechanisms can be divided into two groups: those involving modification of constitutively expressed proteins and those requiring synthesis of new proteins. This review highlights the pineal gland as an excellent model system for studying neurotransmitter-regulated rhythmic gene expression. [source] Temporal coupling of cyclic AMP and Ca2+/calmodulin-stimulated adenylyl cyclase to the circadian clock in chick retinal photoreceptor cellsJOURNAL OF NEUROCHEMISTRY, Issue 4 2006Shyam S. Chaurasia Abstract cAMP signaling pathways play crucial roles in photoreceptor cells and other retinal cell types. Previous studies demonstrated a circadian rhythm of cAMP level in chick photoreceptor cell cultures that drives the rhythm of activity of the melatonin synthesizing enzyme arylalkylamine N -acetyltransferase and the rhythm of affinity of the cyclic nucleotide-gated channel for cGMP. Here, we report that the photoreceptor circadian clock generates a rhythm in Ca2+/calmodulin-stimulated adenylyl cyclase activity, which accounts for the temporal changes in the cAMP levels in the photoreceptors. The circadian rhythm of cAMP in photoreceptor cell cultures is abolished by treatment with the l -type Ca2+ channel antagonist nitrendipine, while the Ca2+ channel agonist, Bay K 8644, increased cAMP levels with continued circadian rhythmicity in constant darkness. These results indicate that the circadian rhythm of cAMP is dependent, in part, on Ca2+ influx. Photoreceptor cell cultures exhibit a circadian rhythm in Ca2+/calmodulin-stimulated adenylyl cyclase enzyme activity with high levels at night and low levels during the day, correlating with the temporal changes of cAMP in these cells. Transcripts encoding two of the Ca2+/calmodulin-stimulated adenylyl cyclases, type 1 and type 8 (Adcy1 and Adcy8), displayed significant daily rhythms of mRNA expression under a light,dark cycle, but only the Adcy1 transcript rhythm persisted in constant darkness. Similar rhythms of Adcy1 mRNA level and Ca2+/calmodulin-stimulated adenylyl cyclase activity were observed in retinas of 2-week-old chickens. These results indicate that a circadian clock controls the expression of Adcy1 mRNA and Ca2+/calmodulin-stimulated adenylyl cyclase activity; and calcium influx into these cells gates the circadian rhythm of cAMP, a key component in the regulation of photoreceptor function. [source] Choline acetyltransferase activity at different ages in brain of Ts65Dn mice, an animal model for Down's syndrome and related neurodegenerative diseasesJOURNAL OF NEUROCHEMISTRY, Issue 2 2006Andrea Contestabile Abstract Ts65Dn mice, trisomic for a portion of chromosome 16 segmentally homologous to human chromosome 21, are an animal model for Down's syndrome and related neurodegenerative diseases, such as dementia of the Alzheimer type. In these mice, cognitive deficits and alterations in number of basal forebrain cholinergic neurons have been described. We have measured in Ts65Dn mice the catalytic activity of the cholinergic marker, choline acetyltransferase (ChAT), as well as the activity of the acetylcholine-degrading enzyme acetylcholinesterase (AChE), in the hippocampus and in cortical targets of basal forebrain cholinergic neurons. In mice aged 10 months, ChAT activity was significantly higher in Ts65Dn mice, compared to 2N animals, in the hippocampus, olfactory bulb, olfactory cortex, pre-frontal cortex, but not in other neocortical regions. At 19 months of age, on the other hand, no differences in ChAT activity were found. Thus, alterations of ChAT activity in these forebrain areas seem to recapitulate those recently described in patients scored as cases of mild cognitive impairment or mild Alzheimer's disease. Other neurochemical markers putatively associated with the disease progression, such as those implicating astrocytic hyperactivity and overproduction of amyloid precursor protein family, were preferentially found altered in some brain regions at the oldest age examined (19 months). [source] Effects of NGF on different phenotypes and genotypes of cholinergic murine SN56 cellsJOURNAL OF NEUROCHEMISTRY, Issue 2003H. Bielarczyk Nerve growth factor (NGF) is important for differentiation and maintenance of septal cholinergic neurons. It caused concentration-dependent increase of choline acetyltransferase (ChAT) activity ([EC50%] 1 ng/mL), acetylcholine (ACh) content and morphologic maturation of SN56TrkA(+)p75(+) but not TrkA(,)p75(+) cells. NGF added with cyclic AMP altered significantly differential effects of the latter neither in TrkA(,) nor TrkA(+). However, when cyclic AMP-predifferentiated cells were treated with NGF alone, it caused suppression of the cholinergic phenotype in both cell lines. Anti-p75 antibodies totally reversed inhibitory effects of NGF on ChAT activity. Differentiation was accompanied by increase whereas its reversal by decrease of intracellular Ca content. These data indicate that NGF may exert opposite effects on phenotype of cholinergic neurons by p75 receptor signaling pathways and changes in intracellular Ca. Acknowledgement: Supported by KBN project 6P05A 01020. [source] Dephosphorylation of pCREB by protein serine/threonine phosphatases is involved in inactivation of Aanat gene transcription in rat pineal glandJOURNAL OF NEUROCHEMISTRY, Issue 1 2003Marco Koch Abstract The rat pineal gland is a suitable model to investigate neurotransmitter-controlled gene expression, because it is well established that the stimulation of melatonin biosynthesis by norepinephrine (NE) depends on the activation of the gene that encodes arylalkylamine N -acetyltransferase (AANAT), the melatonin rhythm enzyme. The mechanisms responsible for downregulation of Aanat transcription are less clear. In this in vitro study we investigated the role of pCREB dephosphorylation for termination of Aanat gene transcription. Immunosignals for pCREB, strongly induced after NE stimulation, rapidly decreased after withdrawal of NE. The immunoreactivity of the inhibitory transcription factor ICER increased twofold after NE treatment for 6 h, but did not change within 30 min after removal of the stimulus. Application of protein serine/threonine phosphatase (PSP) inhibitors prevented pCREB dephosphorylation and blocked the decreases in Aanat mRNA levels, AANAT protein amount and melatonin biosynthesis all of which occurred rapidly after NE withdrawal. PSPs in the rat pineal gland were characterized by immunocytochemistry and immunoblotting. NE-stimulation for 8 h induced accumulation of PSP1-catalytic subunit (CSU) in pinealocyte nuclei, but did not affect the distribution of PSP2A-CSU. The results identify dephosphorylation of pCREB by PSPs as an essential mechanism for downregulation of Aanat transcription in the rat pineal gland. [source] | |||||||||||||||||||||||||||||||||||||