Wheat chromosome 2D carries genes controlling the activity of two DNA-degrading enzymes

DNA-degrading enzymes of 24.0 kDa and 27.0 kDa were observed to have different activities in two common wheat (Triticum aestivum L.) cultivars, Wichita and Cheyenne. A substrate-based SDS-PAGE assay revealed that these two enzymes were much more active in Wichita than in Cheyenne. Genes controlling the activities of these two enzymes were localized on chromosome 2D by testing DNA-degrading activities in reciprocal chromosome substitution lines between Wichita and Cheyenne. While the allele on Wichita chromosome 2D stimulated the activities of the 24.0- and 27.0-kDa enzymes in Cheyenne, the allele on Cheyenne chromosome 2D did not reduce the activities of the 24-kDa and 27-kDa enzymes in Wichita. Whether these genes code for the DNA-degrading enzymes themselves or for factors that regulate the enzyme activities remains unknown.This work was supported in part by USDA-Competitive Research Grants Office grant No. 90-37140-5426 to P.S.B. Contribution from Agricultural Research Division, University of Nebraska. Journal Series Number 10304     

Monocyte-derived dendritic cells that capture dead tumor cells secrete IL-12 and TNF-α through IL-12/TNF-α/NF-κB autocrine loop

This study focused on the question of how monocyte-derived dendritic cells (Mo-DCs) that capture dead tumor cells (Mo-DCs-Tum) secrete interleukin 12 (IL-12) and tumor necrosis factor (TNF-). Mo-DCs-Tum showed higher secretions of IL-12 and TNF- than were shown by Mo-DCs. Enhanced nuclear factor-kappa B (NF-B) activation was also induced in Mo-DCs-Tum within 6 h. The NF-B inhibitor, pyrrolidine dithiocarbamate (PDTC), suppressed both IL-12 and TNF- secretions from Mo-DCs-Tum. Administration of recombinant TNF- or IL-12 enhanced IL-12 or TNF- secretion respectively in Mo-DCs-Tum. Addition of anti-TNF- or anti-IL-12 neutralizing antibody decreased NF-B activation and IL-12 or TNF- secretion in Mo-DCs-Tum. These results suggest that TNF- or IL-12 secretion induces NF-B activation, and it stimulates further TNF- and IL-12 secretions, i.e., an IL-12/TNF-/NF-B autocrine loop, in Mo-DCs-Tum. Thus, Mo-DCs-Tum secrete a large amount of IL-12 and TNF- through accelerated NF-B activation induced by the IL-12/TNF-/NF-B autocrine loop.     

The reduction of tetrazolium salts by plant mitochondria

Summary Data has been obtained concerning the reduction of tetrazolium salts by mitochondria isolated from Jerusalem artichoke tubers with succinate as the substrate using a direct recording spectrophotometric method of assay. ATP was found to increase the rate of reduction of the tetrazolium salts, this being independent of the effect ATP had on the rate of oxygen uptake. The magnitude of the stimulation by ATP depended on the concentration of tetrazolium salts present and under certain circumstances was suppressed by the addition of azide and cyanide. The sites at which the tetrazolium salts were reduced along the electron transport chain were investigated. The role of ATP has been discussed in relation to the mechanism of tetrazolium reduction.Abbreviations TTC 2,3,5-triphenyl-2,1,3,4-tetrazolium chloride - BT 5,5-diphenyl-3,3-(3,3-dimethoxy-4,4-diphenylene)-ditetrazolium chloride - NT 2,2,5,5-tetraphenyl-3,3-(p-diphenylene)-ditetrazolium chloride - MTT 3-(4,5-dimethyl thiozolyl-2)-2,5-diphenyl tetrazolium bromide - INT 2-(p-iodophenyl)-3-p-dinitrophenyl-3-p-nitrophenyl-5-phenyl tetrazolium chloride - NBT 2,2-dinitrophenyl-5,5-diphenyl-3,3-dimethoxy-4,4-diphenylene)-ditetrazolium chloride - TNBT 2,2-5,5-tetra-p-nitrophenyl-3,3-dimethoxy-4,4-diphenylene) ditetrazolium chloride     

Extent of heterosis in rice (Oryza sativa L.) under cold stress conditions — yield and its components

Summary The nature and extent of heterosis for grain yield and its component characters was estimated in a seven-parent F₁-diallel cross in rice under the cold stress conditions of Palampur, representing sub-temperate climate of the mid hills of Himachal Pradesh. The highest heterotic effects for grain yield were observed in Himalaya 1/Phul Patas 72 and China 988/Himdhan hybrids, which gave 60.36 and 32.48% heterosis, respectively, over mid parent, 38.76 and 26.86% heterosis, respectively, over high parent and 32.30 and 26.86% heterosis, respectively, over the best variety, Himdhan. Heterosis for grain yield in these crosses was due to an increase in tiller number, panicle length, spikelets/panicle and 1,000-grain weight. Large number of crosses exhibited significant heterosis for high spikelet sterility and the majority of them had significant heterosis for late flowering, taking more than 120 days to flower, which resulted in lack of heterosis for grain yield in such crosses due to cold stress at the reproductive stage.Part of the thesis submitted by the senior author to Himachal Pradesh Agricultural University, Palampur (H.P.) India, in partial fulfilment of the requirements for the degree of Doctor of Philosophy in Agriculture     

Isoform-Specific Membrane Translocation of Protein Kinase C After Ischemic Preconditioning

Mild cerebral anoxic/ischemic/stress insults promote tolerance and thereby protect the brain from subsequent lethal anoxic/ischemic insults. We examined whether specific activation of PKC , , , or isoforms is associated with ischemic preconditioning (IPC) in rat brain. IPC was produced by a 2-minute global cerebral ischemia. Membrane and cytosolic fractions of the hippocampi were immunoblotted using specific antibodies for PKC, , , and . PKC showed a significant translocation to the membrane fraction from 30 min to 4 h and PKC at 4 h following IPC. In contrast, the membrane/cytosol ratio of PKC showed a tendency to decrease at 30 min and 8 h, and the membrane/cytosol ratio of PKC was significantly decreased from 30 min to 24 h following IPC. These findings indicate PKC isoform-specific membrane translocations in the hippocampus after brief global brain ischemia and suggest that activation of PKC and PKC may be associated with IPC-induced tolerance in the rat hippocampus.     

Variability in the 11S globulin fraction of seed storage proteins ofHelianthus (Asteraceae)

The seed storage globulins from sixHelianthus and four hybrids were studied using mono and bidimensional gel SDS electrophoresis (+ 2 mercaptoethanol). The polypeptide composition of each subunit was determined. Different pairs are specifically expressed according to the species studied. Three typical patterns were discriminated. All the studied species exhibit five subunits: two of them are expressed in all the species (₁₁ and ₂₂). The subunit corresponding to the ₁₁ pair is present inH. petiolaris and in the three populations ofH. annuus studied. The _2b2 pair is common toH. annuus andH. argophyllus. H. petiolaris presents two specific _2a2 and ₄₄ pairs andH. annuus a specific ₃₃ pair. InH. argophyllus _{11 33} or ₄₄ are never observed but are replaced by ₁₃ and ₃₁ pairs. Some globulins, poorly represented, are of forms but present chains of higher molecular weights (in the range 54–56 kDa). Expressing variations in the banding patterns between these species by the use of a similarity index reveals complete identity between the three populations ofH. annuus. Identity between the twoH. petiolaris studied is also observed.H. annuus andH. argophyllus appear to be closer to each other thanH. petiolaris concerning the seed storage globulins.     

Characterization of the putative α subunit of a heterotrimeric G protein in rice

A recombinant protein with a cDNA that encodes the putative subunit of a rice heterotrimeric G protein was synthesized in Escherichia coli and purified. The recombinant protein (rGrice ) with an apparent molecular mass of 45 kDa was bound with guanosine 5-(3-O-thio)triphosphate with an apparent association constant (kapp) of 0.36. The protein also hydrolyzed GTP and its Kcat was 0.44. rGrice was ADP-ribosylated by activated cholera toxin.Monoclonal antibodies raised against rGrice reacted with a 45 kDa polypeptide localized in the plasma membrane of rice seedlings. The peptide map of this polypeptide after digestion with V8 protease was identical to that of rGrice . A 45 kDa polypeptide in the plasma membrane, as well as rGrice , was ADP-ribosylated by activated cholera toxin. The GTPase activity of the plasma membrane was stimulated 2.5-fold by mastoparan 7 but not mastoparan 17. These properties were similar to those of the subunits of heterotrimeric G proteins in animals, suggesting that the putative subunit is truly the subunit itself.     

Measurement of H2'-C2' and H3'-C3' dipolar couplings in RNA molecules

The quality of nucleic acid solution structures can be significantly improved using residual dipolar coupling data. However, many of the one-bond couplings that could be used for this purpose are difficult to measure. Conventional 2D experiments are often unable to reveal one-bond H2-C2 and H3-C3 couplings in large RNA molecules due to spectral overlap. Here we show how to use 3D HCcH-COSY and Relay HCcH-COSY to measure one-bond H2-C2 and H3-C3 couplings which improved the precision of the structures obtained recently for a 42 nucleotide RNA.     

α-Globin gene deletion causes α-thalassemia syndromes in two German families

Summary Restriction endonuclease mapping of chromosomal DNA has been used to determine whether the -globin gene deletion or non-deletion form of -thalassemia is the underlying molecular defect in individuals of two unrelated German families with -thalassemia syndromes. The obtained DNA pattern in all cases indicated loss of -globin genes resulting in-/,--/, and--/- genotypes in thalassemia-2, -thalassemia-1, and Hb H individuals respectively. The chromosomes showing loss of one -globin gene in -thalassemia-2 and Hb H disease were characterized by the so-called rightward deletion form exhibiting loss of a 3.7 kb DNA fragment in the -gene cluster.     

Partial purification of myosin from lily pollen tubes by monitoring with in vitro motility assay

Summary Myosin in pollen tubes ofLilium longiflorum was partially purified, using an in vitro motility assay as a monitor. The main components in the partially purified preparation had molecular masses of 110, 120, and 140 kDa in SDS-PAGE. They became bound to actin filaments in an ATP-dependent manner. Among the components, only that of 120 kDa became bound to ATP and was concluded to be the heavy chain of pollen tube myosin.Abbreviations ATP adenosine-5-triphosphate - DTT dithiothreitol - EB extraction buffer - EGTA ethyleneglycol-bis-(-aminoethylether) N, N, N, N-tetraacetic acid - PAGE polyacrylamide gel electrophoresis - PIPES piperazine-N,N-bis-(2-ethanesulfonic acid) - PMSF phenylmethylsulfonyl fluoride - SDS sodium dodecylsulfate - TBS Tris buffered saline - TEB Tris-EGTA buffer     

The composition of Erythrosins, Fluorescein, Phloxine and Rose Bengal: a study using thin-layer chromatography and solvent extraction

Synopsis Commercial samples of Erythrosin B (CI 45430), Erythrosin Y (CI 45425), Fluorescein (CI 45350), Phloxine (CI 45410) and Rose Bengal (CI 45440) have been analysed by thin-layer chromatography. The Erythrosins were found to be mixtures consisting in the main of 4-iodofluorescein, 4,5-di-iodofluorescein, 2,4,5-triiodofluorescein and 2,4,5,7-tetraiodofluorescein, in some instances together with 2,4,5-tri-iodo-4,5,6,7-tetrachlorofluorescein and 2,4,5,7-tetraiodo-4,5,6,7-tetrachlorofluorescein. Samples of Fluorescein were mixtures of the nominal dye usually with traces of several unidentified, fluorescent components. Those of Phloxine consisted mainly of mixtures of 4-bromo-4,5,6,7-tetrachlorofluorescein, 4,5-dibromo-4,5,6,7-tetrachlorofluorescein, 2,4,5-tribromo-4,5,6,7-tetrachlorofluorescein and 2,4,5,7-tetrabromo-4,5,6,7-tetrachlorofluorescein, often with 4,5,6,7-tetrachlorofluorescein Samples of Rose Bengal were mixtures of 4-iodo-4,5,6,7-tetrachlorofluorescein, 4,5-di-iodo-4,5,6,7-tetrachlorofluorescein, 2,4,5-tri-iodo-4,5,6,7-tetrachlorofluorescein and 2,4,5,7-tetraiodo-4,5,6,7-tetrachlorofluorescein together with some unidentified components.Most of the commercial dye samples gave an insoluble residue when extracted with methanol. This residue was usually inorganic carbonate or halide. Some possible practical consequences of the various impurities are discussed.     

Use of a fluorescence proble for hydrophobic groups, anilinonaphthalene sulphonic acid, in the supravital study of unusual slug oocyte nucleoli

Synopsis Oocytes of the slug,Limax flavus, display cytoplasmic and nucleolar fluorescence in the presence of 8-anilino-1-naphthalene sulphonic acid, a fluorescent probe for hydrophobic groups. When nuclei are isolated in a balanced salt solution, or in 0.25 m sucrose, chromosomes also become fluorescent, but chromosomes never display fluorescence in intact cells. In the complex multiform nucleolus characteristic of slug oocyte nuclei, the nucleolar cap displays conspicuously high levels of fluorescence and patterns of differential fluorescence are obvious. It would appear that this and similar reagents could be of significant value in the investigation of nucleoli in supravital and unfixed preparations of isolated nuclei.     

The level of free amino acids in erythrocytes of different breeds of hen

Summary The content of free amino acids was determined in erythrocytes of adult Leghorn (Lg, White Rock (WR) and Cornish (Cr) hens, bred under identical conditions. The concentration of total amino acids was twice as high in the erythrocytes as in plasma, amounting to 396 m/100 ml, 424 m/100 ml and 475m/100 ml in White Rock, Cornish and Leghorn hens, respectively.Significant differences were found in the ratio of basic amino acids to acidic amino acids. These values were 0.76, 1.75 and 3.19 in White Rick, Leghorn and Cornish hens, respectively; in the plasma of all 3 breeds the ratio was 1. Statistically significant interbreed differences were expressed more distinctly in erythrocyte than in plasma amino acid concentrations. For absolute concentrations the differences were significant in the case of 9 amino acids.     

Carotenoid pigments of Sorangium compositum (Myxobacterales) including two new carotenoid glucoside esters and two new carotenoid rhamnosides

Summary The carotenoid pigments of the myxobacterium Sorangium compositum were analyzed by chromatographical and chemical techniques and by visible, infra red, and mass spectroscopy. Besides -carotene, neurosporene, torulene, lycopene, and 1,2-dihydro-1-hydroxy--carotene, four new carotenoid glycosides were found. These pigments were identified as 1,2-dihydro-1-hydroxy-torulene glucoside ester (I), 1,2-dihydro-3,1-dihydroxy-torulene glucoside ester (III), 1,2-dihydro-1-hydroxy-torulene rhamnoside (II), and 1,2-dihydro-3,1-dihydroxytorulene rhamnoside (IV).Fifth communication on the carotenoids of myxobacteria. Fourth communication see Arch. Mikrobiol. 76, 364–380 (1971).     

Biological chromatology. The laws of colour and design in nature

Summary Biochromatology can be defined as the study of the relationships between colours, and between colours and design in nature. Certain laws concerning biochromatics can be formulated in the terminology of the Colour Circle. One of them is that planes in warm colours and planes in cold colours never adjoin. Another law is that design is never polychromatic. It seems that the observed biochromatical facts can be explained with the help of the principle optimal manifestation of colour, which in turn is based on the formative principles of matter itself.     

Formation of the imidazolides of dinucleotides under potentially prebiotic conditions

Summary Imidazolides of dinucleotides such as ImpApA can be formed from the corresponding dinucleotides in a two-stage process, which gives up to 15% yields under potentially prebiotic conditions. First a solution of the dinucleotide and sodium trimetaphosphate is dried out at constant temperature and humidity. This produces polyphosphates such as pnApA in excellent yield (80%). The products are dissolved in water, imidazole is added, and the solution is dried out again. This yields the 5-phosphorimidazolides.Abbreviations P₃! trimetaphosphate - A adenosine - U uridine - EDTA ethylenediaminetetraacetic acid - Ap adenosine 2(3)-phosphate - Ap! adenosine cyclic 2:3-phosphate - pA adenosine 5-phosphate - pA²p adenosine 2, 5-diphosphate - pA³p adenosine 3, 5-diphosphate - pAp! 5-phospho-adenosine cyclic 2:3-phosphate - ATP adenosine 5-triphosphate - ImpA adenosine 5-phosphorimidazolide - A²pA adenylyl-[25]-adenosine - A³pA adenylyl-[35]-adenosine - A²pU adenylyl-[25]-uridine - A³pU adenylyl-[35]-uridine - pA²pA 5-phosphoadenylyl-[25]-adenosine - pA³pA 5-phospho-adenylyl-[35]-adenosine - pA²pU 5-phospho-adenylyl-[25]-uridine - pA³pU 5-phospho-adenylyl-[35]-uridine - pApN (N= A, U) 5-phosphate of a dinucleoside phosphate - p_nApN (N = A, U; n = 2, 3, 4.) 5-polyphosphate of a dinucleoside phosphate - ImpA²pA imidazolide of pA²pA - ImpA³pA imidazolide of pA³pA - ImpA²pU imidazolide of pA²pU - ImpA³pU imidazolide of pA³pU - ImpApN imidazolide of pApN     

The structure of the incompatibility factors of Schizophyllum commune: Constitution of the three classes of B factors

Summary The B factors of Schizophyllum commune are of 3 classes: The high recombining class I has 7 alleles and 7 alleles; the low recombining classes are class II with 7 allels and probably 2 alleles and class III with probably 2 (or also 2) alleles and 7 allels. A fourth hypothetical class (-) was not found and either does not exist or is indistinguishable from class III by the tests employed. The and alleles differ from and by either (a) mutations affecting both mating specificity and recombination frequency, or (b) deletions involving most of the B region.The research was supported by a grant from the Atomic Energy Commission of the U.S. No. (30-1)-3875 and was performed at the Biological Laboratories, Harvard University, Cambridge, Mass., U.S.A.     

Sequence analysis of 22 kDa-like α-coixin genes and their comparison with homologous zein and kafirin genes reveals highly conserved protein structure and regulatory elements

Several genomic and cDNA clones encoding the 22 kDa-like -coixin, the -prolamin of Coix seeds, were isolated and sequenced. Three contiguous 22 kDa-like -coixin genes designated -3A, -3B and -3C were found in the 15 kb -3 genomic clone. The -3A and -3C genes presented in-frame stop codons at position +652. The two genes with truncated ORFs are flanking the -3B gene, suggesting that the three -coixin genes may have arisen by tandem duplication and that the stop codon was introduced before the duplication.Comparison of the deduced amino acid sequences of -coixin clones with the published sequences of 22 kDa -zein and 22 kDa-like -kafirin revealed a highly conserved protein structure. The protein consists of an N-terminus, containing the signal peptide, followed by ten highly conserved tandem repeats of 15–20 amino acids flanked by polyglutamines, and a short C-terminus. The difference between the 22 kDa-like -prolamins and the 19 kDa -zein lies in the fact that the 19 kDa protein is exactly one repeat motif shorter than the 22 kDa proteins.Several putative regulatory sequences common to the zein and kafirin genes were identified within both the 5 and 3 flanking regions of -3B. Nucleotide sequences that match the consensus TATA, CATC and the ca. –300 prolamin box are present at conserved positions in -3B relative to zein and kafirin genes. Two putative Opaque-2 boxes are present in -3B that occupies approximately the same positions as those identified for the 22 kDa -zein and -kafirin genes. Southern hybridization, using a fragment of a maize Opaque-2 cDNA clone as a probe, confirmed the presence of Opaque-2 homologous sequences in the Coix and sorghum genomes.The overall results suggest that the structural and regulatory genes involved in the expression of the 22 kDa-like -prolamin genes of Coix, sorghum and maize, originated from a common ancestor, and that variations were introduced in the structural and regulatory sequences after species separation.     

Differences in the heat-shock response between thermotolerant and thermosusceptible cultivars of hexaploid wheat

Summary Heat-shock protein (HSP) gene expression in two wheat lines cv Mustang (heat-tolerant) and cv Sturdy (heat-susceptible) were analyzed to determine if wheat genotypes differing in heat tolerance also differ in in-vitro HSP synthesis (translatable HSP mRNAs) and steady-state levels of HSP mRNA. Several sets of mRNA were isolated from seedling leaf tissues which had been heat-stressed at 37 °C for various time intervals. These mRNAs were hybridized with HSP cDNA or genomic DNA probes (HSP17, 26, 70, 98, and ubiquitin). Protein profiles were compared using in-vitro translation and 2-D gels. The Northern slot-blot data from the heat-stress treatment provide evidence that the heat-tolerant cv Mustang synthesized low molecular weight (LMW) HSP mRNA earlier during exposure to heat shock and at a higher level than did the heat-susceptible cv Sturdy. This was especially true for the chloroplast-localized HSP. The protein profiles shown by 2-D gel analysis revealed that there were not only quantitative differences of individual HSPs between the two wheat lines, but also some unique HSPs which were only found in the Mustang HSP profiles. The high level of RFLP between the two wheat lines was revealed by Southern blot hybridization utilizing a HSP17 probe. These data provide a molecular basis for further genetic analysis of the role of HSP genes in thermal tolerance in wheat.     

Microsatellite markers in the hexaploid Prunus domestica species and parentage lineage of three European plum cultivars using nuclear and chloroplast simple-sequence repeats

In a previous study, cDNA microsatellite markers were described in apricot (Prunus armeniaca L.). Specific PCR primers were designed to amplify the microsatellite-containing regions from genomic DNA in different Prunus species. In the present work, cDNA microsatellite markers were developed in the hexaploid Prunus domestica L. species and polymorphism was ascertained in a segregating plum population. Co-dominant mendelian segregation of alleles was demonstrated and microsatellite polymorphism displayed up to 6 alleles per SSR locus per individual. Parentage lineage of three full-sib European plum cultivars (cv. Cacanska najbolja, Cacanska rana and Cacanska lepotica) was reconstructed by the analysis of the above nuclear SSR markers, completed by four chloroplastic microsatellite loci. The six most informative nuclear loci enabled discrimination between the three Cacak cultivars and unrelated individuals as well as the previously proposed parents, Wangenheim and Pozegaca. Data obtained support previous evidence that these cultivars originated from the Stanley cultivar. However, SSR analysis finally excluded Wangenheim as the other possible parent. Based on the results obtained with nuclear and chloroplast SSR loci, we propose the origin of those three Cacak cultivars in a cross between Stanley as the mother plant and Ruth gerstetter as the pollinator. Furthermore, we demonstrate the utility of these apricot SSR markers for genotype fingerprinting of the hexaploid plum cultivars.