FT Ⅰ, a novel positive myeloid-lineage-specific transcription regulatory element within the mouse myeloperoxidase gene enhancer, En 1

INTanDUCTI0NMyeloidcelldifferentiati0n,inwhichmultip0tentialprogenitorcellsarec0nvertedint00neofthesixmaturedifferentiatedcells,i.e.,erythr0cytes,platelets(megakary-ocytes),macr0phages,neutr0phils,e0sinophilsandbas0phils,involvestemporalre-gulati0nofexpression0fanumberoflineage-anddifferentiationstage-specificgenes.Understandingthedevel0pmentalspecificationoflineageaJswellasmaturationstageassociatedpatterns0fgeneexpressioninmyel0idcelldifferentiationrequiresanin-sightintothecontrol0findivid…     

Phytochelatin-mediated cadmium tolerance inschizosaccharomyces pombe

Summary Plants and certain fungi respond to heavy metal toxicity with the induced synthesis of metal-binding peptides known as phytochelatins (PCs). With cadmium, PCs can bind the metal to form a low molecular weight PC-Cd complex and a high molecular weight PC-Cd-S²⁻ complex. The sulfide ions enhance the stability and Cd-binding capacity of the metal chelate, and formation of this sulfide-containing complex is associated with enhanced tolerance to cadmium. Molecular analyses of two fission yeast mutants that fail to produce a wild type level of the PC-Cd-S²⁻ complex have determined that a vacuolar membrane transporter and several enzymes of the purine biosynthesis pathway are necessary in vivo for formation of the PC- Cd-S²⁻ complex. A model based on vacuolar sequestration of the PC-Cd complex by an ATP-binding cassette-type transporter and its subsequent maturation into the stable PC-Cd-S²⁻ complex via the actions of two purine biosynthetic enzymes is described. Presented in the Session-in-Depth Bioremediation through Biotechnological Means at the 1993 Congress on Cell and Tissue Culture, San Diego, CA, June 5–9, 1993.     

Utilization of adenine and guanine as nitrogen sources by Chlamydomonas reinhardtii cells

Chlamydomonas reinhardtü Dangeard, adenine or guanine can be used as the sole nitrogen source for growth by means of an inducible system which is repressed by ammonia. Cells grown on either adenine or guanine were able to take up both purines, although the adenine uptake rate was always about 40% of the guanine uptake rate. Both adenine and guanine were taken up by an inducible system(s) exhibiting hyperbolic kinetics with identical apparent A, values of 3-2 mmol m^?3 for adenine and 3-2mmol m^?3 for guanine. Adenine and guanine utilization depended on pH, with similar optimal pH values of 7·3 and 7·4, respectively. Adenine and guanine each acted as a competitive inhibitor of the other's uptake, and their utilization was also inhibited by hypoxanthine, xanthine and urate. Inhibition of adenine uptake by guanine and hypoxanthine was competitive, with A′, values of 5·5 and 1. 6 mmol m^?3 respectively. Guanine uptake was also inhibited competitively by adenine (K₁= 1·3mmol m^?3) and hypoxanthine (K₁= 3. 3 mmol m^?3). Utilization of both adenine and guanine was inhibited by cyanide, azide, 3-(3,4-dichlorophenyl)-1,1-dimethyl urea, 2,4-dinitrophenol and carbonylcyanide m-chlorophenylhydrazone, and was also sensitive to p-hydroxymercuribenzoate and N-ethyl-maleimide. On the basis of these results, taken together, the possibility that adenine and guanine are translocated into Chlamydomonas by a common system is discussed.     

Incomplete cerebral ischemia in the rat provokes increase of tissue and plasma malondialdehyde

Short-term incomplete cerebral ischemia was induced in the rat by bilaterally clamping for 5 min the common carotid arteries; subsequent reperfusion of 10 min was obtained by removing carotid occlusion. At the end of ischemia or reperfusion, animals were sacrificed by decapitation. A control group was represented by sham-operated rats. Peripheral venous blood samples were withdrawn from the femoral vein from rats subjected to cerebral reperfusion 5 min before ischemia, at the end of ischemia, and 10 min after reperfusion. A highly sensitive HPLC method for the direct determination of malondialdehyde, oxypurines, and nucleosides was used on 200 μL of brain tissue and plasma extracts. Incomplete cerebral ischemia induced the, appearance of a significant amout of tissue malondialdehyde (undetectable in control animals) and a decrease of ascorbic acid. A further 6.6-fold increase of malondialdehyde and a 18.5% decrease of ascorbic acid occurred after 10 min of reperfusion. Plasma malondialdehyde, which was present in minimal amount before ischemia, significantly increased after 5 min of ischemia, being strikingly augmented after 10 min of reperfusion. A similar trend was observed for oxypurines and nucleosides. From these data, it can be affirmed that tissue concentrations of malondialdehyde and ascorbic acid, and plasma levels of malondialdehyde, oxypurines, and nucleosides, reflect both the oxygen radical-mediated tissue injury and the depression of energy metabolism thus representing early biochemical markers of short-term incomplete brain ischemia, and reperfusion in the rat.     

Fast atom bombardment combined with tandem mass spectrometry for the determination of nucleosides

The positive and negative ion fast atom bombardment (FAB) mass spectra and fast atom bombardment collisionally activated decomposition (CAD) spectra of a series of nucleosides and two dinucleotides are reported. The nucleosides studied are substituted forms of guanosine, adenosine, nebularine, tubercidin, uridine, and related pyrimidines. The FAB and CAD data both contain similar information. The CAD spectra are found to provide some structural information not found in the FAB mass spectra. Tandem mass spectrometry also allows emphasis to be put on weak fragments which are either not observed in the FAB mass spectrum or are lost in the matrix ion signals.     

Template-directed reactions of aminonucleosides

Summary We have studied the reactions between adenosine 5-phosphorimidazolide and 9-(2-amino-2-deoxyxylofuranosyl) adenine (I) or 3-methylamino-3-deoxyadenosine (II), both with and without a poly (U) template. We find that both amino compounds react much more rapidly than does adenosine, in the absence of a template. The rate of reaction is greatly enhanced by a poly (U) template in the case of I, but the enhancement is slight in the case of II.Abbreviations A adenosine - xylo A_NH2 9-(2-amino-2-deoxy--D-xylofuranosyl) adenine - A_NHMe 3-methylamino-3-deoxyadenosine - ImpA adenosine 5-phosphorimidazolide - A³ pA adenylyl-[35]-adenosine - A² pA adenylyl-[25]-adenosine - U_NPA adenylyl-[52]-2-amino-2-deoxyuridine - xylo A_NPA 9-[adenylyl-(52)-2-amino-2-deoxy--D-xylofuranosyl]adenine - A_(NMe)pA adenylyl-[53]-3-methylamino-3-deoxyadenosine - pA adenosine 5phosphate - AppA P₁, P₂-diadenosine 5pyrophosphate - (pA)_n n = 2, 3 [2-5]-linked oligomers of pA - A² pA² pA [2-5]-linked trinucleoside diphosphate of A - poly (U) polyuridylic acid     

Inner mitochondrial membrane anion channel is present in brown adipocytes but is not identical with the uncoupling protein

Summary Vesicles of inner mitochondrial membrane, mitoplasts, from rat brown adipose tissue were prepared by osmotic swelling and studied using the patch-clamp technique. Current events of a 107.8±8.7 pS (n=16, 21°C) channel were recorded in the mitoplast-attached mode. This channel was selective for anions and its kinetics resembled those of channels previously found in liver and heart mitochondria of mouse and ox. In whole-mitoplast mode each of five purine nucleotides (20 m) blocked the channel. This is the first demonstration of pharmacological blockade of this type of channel. Although a similar anion channel in mouse and ox mitochondria was suggested to be the uncoupling protein (UCP) associated with nonshivering thermogenesis, we present several arguments against this possibility. Thus we describe a high-conductance, purine-nucleotide-binding, anion selective mitochondrial channel, that is not the UCP.     

The metabolism of 6-(2,3,4-trihydroxy-3-methylbutylamino) purine by soybean callus

6-(2,3,4-trihydroxy-3-methylbutylamino) purine (trihydroxyzeatin) applied to soybean callus is metabolised slowly. After 48 h only one peak of biological activity which co-eluted with the applied cytokinin was detected. When the callus was incubated on a medium which contained 10^–5 M trihydroxyzeatin, spiked with 8 {¹⁴C} trihydroxyzeatin, for 28 days, three peaks of biological activity and three peaks of radioactivity were detected. One of the biologically active and radioactive peaks co-eluted with zeatin. Another of the radioactive peaks co-eluted with N-(purin-6-yl) glycine. From the data obtained it apears that trihydroxyzeatin can be both oxidized and reduced by soybean callus. The potential to be converted to zeatin may explain why trihydroxyzeatin and its parent compound, which is usually rapidly metabolised by living material, are equally active in the soybean callus bioassay. From the radioactive data obtained it appears that trihydroxyzeatin is susceptible to oxidation to form N-(purin-6-yl) glycine.     

Cloning and restriction map of the E. coli apt gene

The apt gene coding for Escherichia coli adenine phosphoribosyl transferase has been cloned in pBR322. The restriction map of a 1.6-kb fragment containing the apt gene is presented.