Molecular genetic studies on alcohol and aldehyde dehydrogenase: individual variation, gene mapping and analysis of regulation.
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Alcohol Dehydrogenase↗
Biomedical subjects
M Smith
Publications and source records attributed to M Smith.
Sequence and nitrate regulation of the Arabidopsis thaliana mRNA encoding nitrate reductase, a metalloflavoprotein with three functional domains.
The sequence of nitrate reductase (EC 1.6.6.1) mRNA from the plant Arabidopsis thaliana has been determined. A 3.0-kilobase-long cDNA was isolated from a lambda gt10 cDNA library of Arabidopsis leaf poly(A)+ RNA. The cDNA hybridized to a 3.2-kilobase mRNA whose level increased 15-fold in response to treatment of the plant with nitrate. An open reading frame encoding a 917 amino acid protein was found in the sequence. This protein is very similar to tobacco nitrate reductase, being greater than 80% identical within a section of 450 amino acids. By comparing the Arabidopsis protein sequence with other protein sequences, three functional domains were deduced: (i) a molybdenum-pterin-binding domain that is similar to the molybdenum-pterin-binding domain of rat liver sulfite oxidase, (ii) a heme-binding domain that is similar to proteins in the cytochrome b5 superfamily, and (iii) an FAD-binding domain that is similar to NADH-cytochrome b5 reductase.
Amino Acid Sequence↗
Influence of angiotensin II (AII) and angiotensin I (AI) on renal renin synthesis and release.
This study examined the effect of exogenous AI and AII on renin synthesis and release. AII or AII + AI were given using osmotic minipumps, over a 7 day period to rats receiving oral Enalapril. Enalapril caused a marked increase in renin release and synthesis. AII abolished the rise in plasma and renal renin observed with Enalapril. AI had no inhibitory or stimulatory effect on renin release or synthesis in rats receiving Enalapril and AII.
Angiotensin I↗
Renal enzyme activities in experimental ochratoxin A-induced porcine nephropathy: diagnostic potential of phosphoenolpyruvate carboxykinase and gamma-glutamyl transpeptidase activity.
Enzyme activities have been measured in needle biopsies from kidneys of pigs fed 1 ppm or 0.2 ppm of ochratoxin A for 1-5 wks. After feeding 1 ppm toxin for 1 wk, the activity of cytosolic phosphoenolpyruvate carboxykinase (PEPCK) was decreased by 40% and remained inhibited until the termination of the experiment (5 wk). The activity of gamma-glutamyl transpeptidase, a brush-border enzyme found in the proximal tubules, was reduced to a similar degree and remained inhibited. The activities of hexokinase, a cytosolic enzyme of general distribution in the nephron, and phosphate-dependent glutaminase, a distal tubule enzyme, were not affected. The biopsy results were confirmed by measurements in renal slices taken at the termination of the experiment, except that biopsy samples showed more variation in enzyme activity and a lower PEPCK activity. A guinea pig antibody against the cytosolic form of PEPCK was used to demonstrate that the mitochondrial form of the enzyme, which accounts for a considerable part of the total cellular activity, was not affected by ochratoxin A. When mitochondrial PEPCK activity present in the cytosolic fraction was accounted for, ochratoxin A was found to reduce PEPCK activity by 70-80%. The increase of ochratoxin A exposure from zero through 0.2 ppm to 1 ppm, which resulted in dose-dependent activity decrease of PEPCK and gamma-glutamyl transpeptidase, was accompanied by dose-dependent decrease of renal function, as measured by a reduction of maximal tubular excretion of para-aminohippurate per clearance of inulin (TmPAH/CIn) and an increase in glucose excretion. This suggest that these enzymes are sensitive indicators of ochratoxin A-induced porcine nephropathy. Assuming that porcine nephropathy represents a valid model of endemic (Balkan) nephropathy in humans, the measurement of cytosolic PEPCK and gamma-glutamyl transpeptidase activity in the kidney could be a sensitive test for ochratoxin A-induced disease in humans.
Animals↗
A simple and efficient procedure for generating random point mutations and for codon replacements using mixed oligodeoxynucleotides.
A very simple and highly efficient procedure for the generation of single and multiple substitutions in segments of DNA is described which has no requirements for conveniently placed restriction sites, and allows all DNA sequences to be equally accessible. A mixed pool of oligodeoxynucleotides is synthesized by contaminating the monomeric nucleotides with low levels of the other three nucleotides such that the full-length oligonucleotide contains on the average one to two changes per molecule. This pool is used in priming in vitro synthesis of the complementary strand of cloned DNA fragments in M13 or pEMBL vectors which have previously been passed through a dut-, ung- Escherichia coli host. Strong selection for the newly synthesized strand is provided by transforming the heteroduplex into a dut+, ung+ host. Single and multiple substitutions in the carboxy-terminal coding region of the MATa1 gene of Saccharomyces cerevisiae are introduced at high efficiency (25-55%) and the changes are identified by direct sequencing alone. The same principle can be used to generate multiple sets of changes at any specified codon.
Codon↗
Local radiotherapy for pedal manifestations of HLA-B27-related arthropathy.
Four males with characteristic foot manifestations of HLA-B27-related arthropathy are reported. The severity of this problem and its recalcitrance to standard therapies led to the use of local radiotherapy. The beneficial outcome and minimal short-term side effects of this treatment are discussed in relation to previous experience with radiotherapy in ankylosing spondylitis.
Adolescent↗
Effect of coronary artery revascularization on left and right ventricular ejection fractions during rest and exercise.
We have reported a case of a 55-year-old man who had improvement of both left and right ventricular ejection fractions during rest and exercise after coronary revascularization. Ventricular performance was assessed using radionuclide angiography. Although coronary artery bypass grafting has previously been shown to improve left ventricular function, we believe this is the first report of improved right ventricular function after coronary revascularization.
Coronary Artery Bypass↗
The structure of Marek's disease virus DNA: amplification of repeat sequence in IRs and TRs.
The structure of Marek's disease virus (MDV) DNA was investigated by using Southern blot hybridization analysis. A heterogeneous region was observed in the inverted repeats region, IRs and TRs, as well as in the TRL and IRL. The results of DNA sequencing of the heterogeneous region showed that the heterogeneity of IRS and TRS was due to amplification of a 178-bp repeat sequence. Amplification of IRS and TRS was found in viral DNA from both pathogenic and nonpathogenic strains. The structure of DNA from the latent MDV genome present in established lymphoblastic cells was also determined. Amplification of the 132-bp repeat sequence in IRL and TRL was not found in latent MDV DNA of established lymphoblastic cells, whereas amplification of the 178-bp repeat sequence in IRS and TRS was found in the same DNA.
Base Sequence↗
H2 antagonists and bupivacaine clearance.
The effect of the H2 receptor antagonists cimetidine and ranitidine on bupivacaine clearance was assessed in women scheduled to undergo elective Caesarean section under epidural anaesthesia. Thirty-six women were randomly allocated to receive either no medication, cimetidine 400 mg or ranitidine 150 mg on the night prior to and on the morning of surgery. No significant difference was found between the peak bupivacaine levels: the mean (SD) values were 0.74 (0.17) microgram/ml, 0.81 (0.38) microgram/ml and 0.70 (0.24) microgram/ml in the control, cimetidine and ranitidine groups, respectively. Similarly, the H2 receptor antagonists did not alter the plasma bupivacaine against time curves, half-life or bupivacaine clearance in the three groups studied.
Adult↗
Inhibition of glucose-induced insulin secretion by calcium channel blocking drugs in-vitro but not in-vivo in the rat.
The organic calcium channel blocking drugs darodipine, nicardipine, diltiazem and verapamil inhibited glucose-induced (8.3 mmol L-1) insulin secretion by rat isolated islets in a concentration-dependent manner. The IC50 values (mumol L-1) for nicardipine, diltiazem and verapamil were 0.0025, 1.94 and 3.6, respectively. There were no significant differences between the potencies of any one drug when compared at two different glucose concentrations (27.8 and 8.3 mmol L-1). Isolated islets were also responsive to the calcium channel activating drugs BAY K 8644 and CGP 28392, which enhanced glucose-induced insulin secretion and prevented the inhibitory effect of verapamil. BAY K 8644 was more potent than CGP 28392. In the anaesthetized rat, neither verapamil (1 mg kg-1 i.v. 10 min before glucose) nor nicardipine (0.2 mg kg-1 i.v.) modified the glucose or insulin response to intravenous glucose whilst producing marked cardiovascular effects. The plasma concentrations of nicardipine (3.9 X 10(-8) M at 10 min post injection) were similar to those producing effects in-vitro whereas the plasma concentrations of verapamil (5 X 10(-7) M) were lower. It thus appears that the islet B cell calcium channel is less sensitive to calcium channel blocking drugs in-vivo than in-vitro. Moreover, in-vivo, the cardiovascular system is more sensitive to these drugs than are the islet B cells, although the potencies of the calcium channel blocking drug in isolated islets are similar to those reported for cardiac muscle and vascular smooth muscle in-vitro.
3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
Constitutive expression of a c-myb cDNA blocks Friend murine erythroleukemia cell differentiation.
A full-length human c-myb cDNA clone has been isolated from a CCRF-CEM leukemia cell cDNA library. The plasmid vector contains simian virus 40-derived promotor, splice, and polyadenylation sequences as well as a transcription unit for a dihydrofolate reductase cDNA. We have introduced this construct into Friend erythroleukemia (F-MEL) cells and have isolated a number of clones which contain intact and transcriptionally active human c-myb sequences. F-MEL clones expressing the highest levels of the human c-myb mRNA differentiate poorly in response to dimethyl sulfoxide. Two clones which initially expressed low levels of human c-myb transcripts and which differentiated normally were subsequently inhibited in their ability to differentiate when grown in successively higher concentrations of methotrexate, due to amplification and enhanced expression of plasmid sequences. The inhibitory effect on F-MEL differentiation appeared to be independent of the early decline in c-myc transcripts which were normally regulated in all cases examined. Our results indicate that constitutive expression of a nontruncated human c-myb cDNA can exert profound effects on erythroid differentiation and argue for a causal role of c-myb in the F-MEL differentiation process.
Animals↗
Cytoplasmic Mg2+ concentration in platelets: implications for determination of Ca2+ with aequorin.
The concentration of cytoplasmic ionized Mg2+ ([Mg2+]i) varies considerably among different cell types. It has not been measured in platelets. Incorrect estimates of this value could markedly affect many intracellular investigations, including calibration of measurements of platelet cytoplasmic ionized Ca2+ concentration ([Ca2+]i) with the photoprotein aequorin and other Ca2+-sensitive probes. [Mg2+]i was measured in washed, gel-filtered human platelets suspended in modified Tyrode buffer by two methods: 31P-nuclear magnetic resonance (NMR) spectroscopy of intact platelets and null-point titration in platelets selectively permeabilized with digitonin. The 31P-NMR spectra demonstrated that the [Mg2+]i, as calculated from the chemical shift values of ATP resonances, was 0.23 +/- 0.02 (SD) mM in unstimulated platelets. The mean [Mg2+]i as determined by null-point titration was 0.3 +/- 0.1 mM (range: 0.1-0.6 mM). When this [Mg2+]i value was used to construct a Ca2+-calibration curve for aequorin, the indicated [Ca2+]i values in resting and stimulated platelets were lower than those obtained from curves based on previously assumed values for [Mg2+]i (1.0-1.25 mM). This finding largely resolves the discrepancy between resting [Ca2+]i as determined by aequorin or by quin2, fura-2, and indo-1.
Aequorin↗
Report of the committee on the genetic constitution of chromosomes 9 and 10.
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Chromosome Mapping↗
Augmentation of glucose induced insulin secretion by pertussis vaccine, phentolamine and benextramine: involvement of mechanisms additional to prevention of the inhibitory actions of catecholamines in rats.
Pertussis vaccine, pertussis toxin, and the alpha-adrenoceptor blocking drug phentolamine augment glucose-induced insulin secretion. The present study was carried out to determine the relationship between this action and the ability of these agents to prevent the inhibitory actions of adrenaline. Pertussis vaccine augmented glucose-induced insulin secretion in rat islets ex vivo and prevented the inhibitory actions of adrenaline and clonidine. Incubation of islets with phentolamine or the irreversible alpha-adrenoceptor blocking agent benextramine also augmented glucose-induced insulin secretion. However, the alpha-adrenoceptor blocking drugs idazoxan, yohimbine or phenoxybenzamine, in concentrations that prevented the inhibitory effects of adrenaline and/or clonidine, did not modify glucose-induced insulin release in vitro. Benextramine (1 X 10(-5) mol/l) blocked the inhibitory effect of clonidine, whilst having no significant effect on the response to adrenaline. It is concluded that stimulation of insulin secretion by certain alpha-adrenoceptor blocking drugs can be dissociated from their alpha-adrenoceptor properties. The ability of pertussis vaccine, phentolamine or benextramine to augment glucose-induced insulin release in vitro is unlikely to be due to the prevention of the inhibitory action of endogenous catecholamines.
Adrenergic alpha-Antagonists↗
Menstrual migraine: a double-blind trial of percutaneous estradiol.
The present study investigated whether administration of percutaneous estradiol for the 7 days encompassing menstruation (the paramenstruum) would be effective in alleviating menstrual migraine. The study was a double-blind cross-over placebo comparison of percutaneous estradiol in gel form. Twenty-two women who suffered from regular recurring menstrual migraine were studied during 2 assessment menstrual cycles, 4 treatment cycles (2 of estradiol gel, 2 of placebo gel), and 1 follow-up (no treatment) cycle. Women completed daily records of the occurrence and severity of migraine and medication used. Eighteen women completed the study. There was a significant reduction in the frequency of migraine in the paramenstruum and in the amount of medication taken during use of percutaneous estradiol. Women expressed a significant preference for continuation of therapy with percutaneous estradiol.
Administration, Cutaneous↗
Pairwise linkage analysis of 11 loci on human chromosome 4.
New RFLPs are described for INP10 and interleukin 2. The 55 pairwise genetic linkage relationships for these two loci and nine additional loci on the long arm of chromosome 4 (4q) are reported. Fifteen new linkages are established, and new data are added to the four previously reported linkages on 4q. Tight linkage of interleukin 2 (T-cell growth factor), epidermal growth factor, and alcohol dehydrogenase is described. Significant differences were observed between male and female recombination rates. The female rate was estimated to be 1.27 times the male rate. On the basis of these pairwise results, the order for the 11 loci is D4S35-GC-(ALB/AFP)-MT2P1-D4S1-INP10-ADH3-( EGF/IL2)-(FBB/FBA/FBG)-MNS. This preliminary order can serve as a starting point for more detailed multipoint analysis.
Chromosome Mapping↗
Catecholamines and plasma potassium.
There are many circumstances in which disease or its treatment may influence potassium homeostasis, and yet serious consequences resulting from potassium changes are uncommon. This article explores the mechanisms involved in short-term potassium control and describes their clinical implications.
Adrenergic Agonists↗
Molecular cloning and analysis of P. 69, a vir-controlled protein from Bordetella pertussis.
The cloning sequencing and analysis of an important antigenic component of Bordetella pertussis is described. The gene for P.69, in common with a variety of other so called "virulence" genes, (e.g., adenylate cyclase (AC), pertussis toxin (PT) and filamentous haemagglutinin (FHA)), is under control of the vir locus. The protein P.69 is externally localised on cells and protein preparations are protective as judged by the mouse intra-cerebral challenge test. The gene encoding the P.69 antigen was isolated by hybridization of mixed oligonucleotide probes against B. pertussis genomic DNA. These oligonucleotides were designed from the protein sequence data obtained from a cyanogen bromide digest of the P.69 protein. DNA sequence analysis reveals a G:C rich gene capable of encoding a protein of 910 amino acids and Mr of 93478, whose likely promoter and ribosome binding sites show little homology to their E.coli counterpart. In common with some of the genes in the PT operon the sequence 5'-CCTGG-3' was found 5' to the ATG initiation codon. At the 3', end 29 bases after the TAA stop codon, the sequence 5'GTTTTTCCT-3' was found in an equivalent position to the same sequence in the PT operon. Examination of the protein sequence reveals two regions with directly repeated elements (GGAVP)3(GGFGP)2 and (PQP)5.
Amino Acid Sequence↗