MHC class II sequences of an HLA-DR2 narcoleptic

Narcolepsy has a 98% association with the DR2-Dw2/DQw1 haplotype. To establish if a disease-specific allele is present in narcolepsy, a cDNA library was made from a B-cell line from a DR2,4/DQw1,3 narcoleptic. Clones encoding the two expressed DR2 chains, along with DQw1 and chains, were isolated and completely sequenced. The coding regions of these four genes were similar to published nucleotide and protein sequences from corresponding healthy controls, with some minor exceptions. The 3 untranslated region of one of the DR2 genes in the narcoleptic was extended by 42 bp. Complete sequences were not available for DQw1.2 or from healthy individuals, but first domain nucleotide sequences showed only a single nonproductive difference in DQ. Partial protein sequences of both DQ and from published data were identical. Although the effects of minor differences cannot be ruled out completely, it is concluded that there are probably no narcolepsy-specific DR or DQ / sequences, and that the alleles found in narcolepsy are representative of those found in the healthy population.     

13Cα and 13Cβ chemical shifts as a tool to delineate β-hairpin structures in peptides

Unravelling the factors that contribute to the formation and the stability of -sheet structure in peptides is a subject of great current interest. A -hairpin, the smallest -sheet motif, consists of two antiparallel hydrogen-bonded -strands linked by a loop region. We have performed a statistical analysis on protein -hairpins showing that the most abundant types of -hairpins, 2:2, 3:5 and 4:4, have characteristic patterns of ¹³C and ¹³C conformational shifts, as expected on the basis of their and angles. This fact strongly supports the potential value of ¹³C and ¹³C conformational shifts as a means to identify -hairpin motifs in peptides. Their usefulness was confirmed by analysing the patterns of ¹³C and ¹³C conformational shifts in 13 short peptides, 10–15 residues long, that adopt -hairpin structures in aqueous solution. Furthermore, we have investigated their potential as a method to quantify -hairpin populations in peptides.     

Evolutionary Role of Iron in Metabolism of Prokaryotes and in Biogeochemical Processes

The paper reviews data summarizing points of view about the conceptual role of iron in the appearance and evolutionary formation of the Earth and its biosphere. Participation of iron and its compounds in the appearance and development of processes of anaero- and aerobiosis as fundamental blocks of metabolism is presented as a hierarchical scheme. Magnetically arrayed iron compounds, in which the element is both in the Fe(II) and in the Fe(III) state, are considered a connecting link between the hierarchical levels. It is shown that the energy transformation Fe(II) Fe(III) is an oxidation–reduction energy core of the most important metabolic iron complexes and of processes of biogenesis both at the cellular level and in biogeosystems.     

The effects of collembola grazing on microbial activity in decomposing leaf litter

Summary Ground leaf litter was inoculated with the fungus Coriolus versicolor and incubated in respirometers for 6 days (fresh cultures) or 33 days (senescent cultures) before different number of Folsomia candida were added. Grazing by 5 animals stimulated O₂ consumption in both series of cultures but 10, 15 or 20 animals inhibited microbial respiration. The stimulatory effect was less marked in the senescent cultures. Bacterial and fungal standing crops increased in fresh cultures during the course of the experiment but grazing by collembola increased bacterial and reduced fungal standing, crops in proportion to the grazing intensity. Microbial standing crops were not determined for senescent cultures. Microarthropod feeding activities can therefore exert a strong differential effect on fungal and bacterial populations which has not been previously recognised.     

Hydrogen isotope discrimination in higher plants: Correlations with photosynthetic pathway and environment

Summary The ratio of deuterium to hydrogen (expressed as D) in hydrogen released as water during the combustion of dried plant material was examined. The D value (metabolic hydrogen) determined on plant materials grown under controlled conditions is correlated with pathways of photosynthetic carbon metabolism. C₃ plants show mean D values of-132 for shoots and -117 for roots; C₄ plants show mean D values of -91 for shoots and-77 for roots and CAM plants a D value of-75 for roots and shoots. The difference between the D value of shoot material from C₃ and C₄ plants was confirmed in species growing under a range of glasshouse conditions. This difference in D value between C₃ and C₄ species does not appear to be due to differences in the D value (tissue water) in the plants as a result of physical fractionation of hydrogen isotopes during transpiration. In C₃ and C₄ plants the hydrogen isotope discrimination is in the same direction as the carbon isotope discrimination and factors contributing to the difference in D values are discussed. In CAM plants grown in the laboratory or collected from the field D values range from-75 to +50 and are correlated with ¹³C values. When deprived of water, the D value (metabolic hydrogen) in both soluble and insoluble material in leaves of Kalanchoe daigremontiana Hamet et Perr., becomes less negative. These changes may reflect the deuterium enrichment of tissue water during transpiration, or in field conditions, may reflect the different D value of available water in areas of increasing aridity. Whatever the origin of the variable D value in CAM plants, this parameter may be a useful index of the water relations of these plants under natural conditions.     

Immunosurgical treatment of stage I malignant melanoma

Summary We treated thirty seven patients suffering from a clinical stage I malignant melanoma by immunosurgery and observed them for a three-year period ending December 1975. Prior to surgery the tumor was submitted to the application of DNCB. Patients were divided between those who had or had not been subjected to biopsy before starting treatment. Accordingly, 23 patients treated by immunosurgery as the first treatment, were called the primitive treatment group and 14 patients subjected to a previous biopsy, the secondary treatment group. In the primitive treatment group, all 23 patients survived without any sign of recurrence. Relapse was observed in 4 out of 14 patients of the biopsy group. Extensive histological investigations have been made by light and electron microscopes. The tumors were infiltrated and disrupted by a large number of mononucleated cells. The majority of infiltrating cells were small lymphocytes. True lymphatic follicles have been observed in the skin around the melanoma.     

Production of a novel syrup containing neofructo-oligosaccharides by the cells of Penicillium citrinum

A novel syrup containing neofructo-oligosaccharides was produced from sucrose (Brix 70) by whole cells of Penicillium citrinum. The efficiency of fructo-oligosaccharides production was more than 55% and those of the main carbohydrate components, 1-kestose (Fruf 21Fruf 21 Glc), nystose (Fruf 21Fruf 21 Fruf 21 Glc) and neokestose (Fruf 26 Glc12 Fruf), were 22, 14 and 11%, respectively.     

Different ζ globin gene deletions among Black Americans

Summary Four types of chromosomes with a deletion between the human embryonic and globin genes were identified among 2.8% of 321 Black Americans from Georgia. Two deletions of approximately 11 kb which differed by about 300 bp occurred on chromosomes with or without a polymorphic Xba I site 5 to the globin gene [(X⁺) or (X^-)]. The deletions are identifiable in Xba I digests of genomic DNA using an or a globin gene probe which yield fragments of 23 kb from (X⁺)–^* chromosomes or 27 kb from (X^–)–^* chromosomes. Digestion with other enzymes and probing with both and probes gave fragments typical of the two globin gene deletions previously identified in Polynesians. Among Black Americans, these globin gene deletions have been found in combination with globin gene deletions in trans but not in cis. Homozygotes have not been found. Hematologic data on carriers of the globin gene deletions in association with Hb AS, SS, and SC suggest that these deletions have no effect on the function of the adult globin genes.     

The inflammatory cytokines tumor necrosis factor α and interleukin-1β stimulate phosphatidylcholine secretion in primary cultures of rat type II pneumocytes

Tumor necrosis factor and interleukin-1 increase surfactant secretion in type II pneumocytes in a time- and dose-dependent manner. This stimulatory effect was additive to that of lipopolysaccharide, suggesting that cytokines and lipopolysaccharide may exert their actions through different signal transduction pathways. Tumor necrosis factor and interleukin-1 did not modify the increase on phosphatidylcholine secretion induced by the direct protein kinase C activator tetradecanoylphorbol 13-acetate, whereas this effect was inhibited by the protein kinase C inhibitors bisindolylmaleimide (2 × 10-6M) and 1-(5-isoquinolinylsulphonyl)-2-methyl piperazone (10-4M). In addition, the stimulatory effect of tumor necrosis factor and interleukin-1 was not suppressed by the intracellular Ca2+ chelator BAPTA (5 × 10-6M) or by KN-62 (3 × 10-5M), a specific inhibitor of Ca2+-calmodulin-dependent protein kinase. These results suggest that tumor necrosis factor or interleukin-1 stimulate phosphatidylcholine secretion via protein kinase C activation in a Ca2+ -independent manner.     

Aldimine to ketoamine isomerization (Amadori rearrangement) potential at the individual nonenzymic glycation sites of hemoglobin a: Preferential inhibition of glycation by nucleophiles at sites of low isomerization potential

The relative roles of the two structural aspects of nonenzymic glycation sites of hemoglobin A, namely the ease with which the amino groups could form the aldimine adducts and the propensity of the microenvironments of the respective aldimines to facilitate the Amadori rearrangement, in dictating the site selectivity of nonenzymic glycation with aldotriose has been investigated. The chemical reactivity of the amino groups of hemoglobin A forin vitro reductive glycation with aldotriose is distinct from that in the nonreductive mode. The reactivity of amino groups of hemoglobin A toward reductive glycation (i.e., propensity for aldimine formation) decreases in the order Val-1(), Val-1(), Lys-66(), Lys-61(), and Lys-16(). The overall reactivity of hemoglobin A toward nonreductive glycation decreased in the order Lys-16(), Val-1(), Lys-66(), Lys-82(), Lys-61(), and Val-1(). Since the aldimine is the common intermediate for both the reductive and nonreductive modification, the differential selectivity of protein for the two modes of glycation is clearly a reflection of the propensity of the microenvironments of nonenzymic glycation sites to facilitate the isomerization reaction (i.e., Amadori rearrangement). A semiquantitative estimate of this propensity of the microenvironment of the nonenzymic glycation sites has been obtained by comparing the nonreductive (nonenzymic) and reductive modification at individual glycation sites. The microenvironment of Lys-16() is very efficient in facilitating the rearrangement and the relative efficiency decreases in the order Lys-16(), Lys-82(), Lys-66(), Lys-61(), Val-1(), and Val-1(). The propensity of the microenvironment of Lys-16() to facilitate the Amadori rearrangement of the aldimine is about three orders of magnitude higher than that of Val-1() and is about 50 times higher than that of Val-1(). The extent of nonenzymic glycation at the individual sites is modulated by various factors, such as thepH, concentration of aldotriose, and the concentration of the protein. The nucleophiles—such as tris, glycine ethyl ester, and amino guanidine—inhibit the glycation by trapping the aldotriose. The nonenzymic glycation inhibitory power of nucleophile is directly related to its propensity to form aldimine. Thus, the extent of inhibition of nonenzymic glycation at a given site by a nucleophile directly reflects the relative role ofpK _a of the site in dictating the glycation at that site. The nonenzymic glycation of an amino group of a protein is an additive/synergestic consequence of the propensity of the site to form aldimine adducts on one hand, and the propensity of its microenvironment to facilitate the isomerization of the aldimines to ketoamines on the other. The isomerization potential of microenvironment plays the dominant role in dictating the site specificity of the nonenzymic glycation of proteins.     

A model of associative memory in the brain

A model of associative memory for time varying spatial patterns is proposed and simulated on a digital computer. This is a network composed of many neuron-like elements, and shows an ability for associative memory similar to that of the brain.Suppose a number of sequences of spatial patterns are presented to this network, for example, 12345, ABC, and so on. Then, these patterns are memorized in the network. After that, if any part of one of these sequences, say 23, is presented to the circuit, the rest of the sequence, 45, is recalled following to it. It resembles to such a situation — if we hear a part of a melody which we have memorized in the past, the rest of the melody is recalled even after it is stopped half-way. Although the recalled patterns are not always 100% correct, they are not completely destroyed even if the presented patterns are imperfect.     

Electron microscopic demonstration of phospholipase B activity in the liver and the kidney of the mouse

Summary A method has been developed for electron microscopic histochemical demonstration of phospholipase B, (lecithinase B, E C 3.1.1.5, lysolecithin acyl hydrolase), which hydrolyzes - and -positions of phospholipids in mouse liver, kidney and adrenal tissues. Tissues either fixed in cold 1% paraformaldehyde or unfixed were cut into 40 m frozen sections and were incubated at 37° C in a medium at pH 6.6 or 4.5 containing 2 M lysolecithin and 0.25 mM CaCl₂ for 20 min. The fatty acids liberated by enzymatic hydrolysis were trapped as calcium precipitate and were converted to lead precipitate by treatment with lead nitrate. The reaction products were observed by electron microscopy to be localized on the end of the smooth endoplasmic reticulum at pH 6.6 and in lysosomes and lipid droplets at pH 4.5. It is concluded that the products showed the localization of phospholipase B activity.     

Gamma-tubulin colocalizes with microtubule arrays and tubulin paracrystals in dividing vegetative cells of higher plants

Summary The distribution of -tubulin throughout cell division is studied in several taxa of higher plants. -Tubulin is present along the whole length of microtubules (Mts) in every cell stage-specific Mt array such as the preprophase band, the preprophase-prophase perinuclear Mts, the kinetochore Mt bundles, the phragmoplast, and the telophase-interphase transition Mt arrays. -Tubulin follows with precision the Mt pattern, being absent from any other, Mt-free, cell site. In cells treated with anti-Mt drugs, -tubulin is present only on degrading or on reappearing Mt arrays, while it is totally absent from cells devoid of Mts. -Tubulin is also present in tubulin paracrystals, which are formed in colchicine-treated cells. These observations support the view that in higher plants -tubulin may not be a microtubule-organizing-center-specific protein, but it may play a certain structural and/or functional role being related to - and -tubulin.Abbreviations Mt microtubule - MTOC microtubule-organizing center - PPB preprophase band     

Binding of Amyloid Beta-Protein to Intracellular Brain Proteins in Rat and Human

Amyloid beta-protein (A), in its soluble form, is known to bind several circulatory proteins such as apolipoprotein (apo) E, apo J and transthyretin. However, the binding of A to intracellular proteins has not been studied. We have developed an overlay assay to study A binding to intracellular brain proteins. The supernatants from both rat and human brains were found to contain several proteins that bind to A 1–40 and A 1–42. No major difference was observed in the A binding-proteins from brain supernatants of patients with Alzheimer's disease and normal age-matched controls. Binding studies using shorter amyloid beta-peptides and competitive overlay assays showed that the binding site of A to brain proteins resides between 12–28 amino acid sequence of A. The presence of several intracellular A-binding (AB) proteins suggests that these proteins may either protect A from its fibrillization or alternatively promote A polymerization. Identification of these proteins and their binding affinities for A are needed to assess their potential role in the pathogenesis of Alzheimer's disease.     

Modulation of natural killer activity by thymosin alpha 1 and interferon

Summary A single injection of -interferon (-IFN) (30 000 units/mouse), a major biological modifier of natural killer (NK) cytolytic activity, strongly stimulated NK activity in normal mice, as expected, while the same treatment did not statistically alter the NK response in cyclophosphamide (CY)-suppressed animals.We investigated the possibility of thymosin ₁ cooperating with -IFN in boosting NK activity in CY-suppressed animals.The results show that treatment with thymosin ₁ (200 g/kg) for 4 days, followed by a single injection of -IFN 24 h before testing, strongly restored NK activity in CY-suppressed mice. Thymosin ₁ was, moreover, able to accelerate the recovery rate of NK activity in bone marrow reconstituted murine chimeras.Taken together the data support the concept that the synergic effect between thymosin ₁ and -IFN could be the result of effects on differentiation of the NK lineage at different levels.     

A coupled pacemaker-slave model for the insect photoperiodic clock: interpretation of ovarian diapause data in Drosophila melanogaster

A coupled circadian oscillator model for the insect photoperiodic clock is described which consists of a hierarchically arranged pacemaker and slave. The pacemaker is self-sustained, temperature compensated, and entrainable by the light cycle; the slave is a damping oscillation receiving entrainment from two sources, from the pacemaker via a coupling factor, and also directly from the light. The damping slave oscillation is seen as the photoperiodic oscillator, equivalent to that proposed earlier by Lewis and Saunders (1987). The present simulations describe the effect of the strength of the coupling factor between hypothetical short- and long-period pacemaker oscillations (modelled on the clock mutants per ^sand per ^L2in Drosophila melanogaster) and a slave oscillation with a period of about 24 hours. The output is presented in terms of photoperiodic response curves and Nanda-Hamner, or resonance, plots. With a high coupling strength, the pacemakers strongly entrain the slave, but with a low coupling strength the slave's properties are more evident. The model is presented as a possible explanation for recent ovarian diapause data in D. melanogaster clock mutants (Saunders 1990), but also as a more general model for the role of the insect circadian system in seasonal time measurement.     

Silver nitrate and aminoethoxyvinylglycine affect <Emphasis Type="Italic">Agrobacterium</Emphasis>-mediated apple transformation

In Fuji, the production of ethylene was increased with the addition of AgNO₃ and inhibited with the addition of 10 M aminoethoxyvinylglycine (AVG). The addition of 80 M AgNO₃ to transformed explants of Fuji cultured on selection medium resulted in increased ethylene production (20 l l^–1) at 3 weeks. Under examining the effect of AgNO₃ in Fuji, the 40 M AgNO₃ showed with higher 33.8% and 6.5% in the efficiency of regeneration and transformation. However, ethylene production in Gala explants treated with 10M AgNO₃ (3 l l^–1) decreased after 2 weeks compared with the control (5 l l^–1). Although the regeneration efficiency of Gala with 10 M AgNO₃ was higher (41.1%) than the control (20.1%), there was no significant difference in the transformation efficiency at the same concentration. Shoot regeneration of Fuji and Gala was completely inhibited with 10 M AVG. These results suggest that the addition of AgNO₃ affects the efficiency of Agrobacterium-mediated gene transfer in Fuji.Eun Soo Seong, Ill Min Chung- These two Authors Contributed equally to this work     

Histoenzymatic characterization of sub-types of Type I fibres in fast muscles of rats

Summary The histochemical activities of succinic dehydrogenase (SDH), myofibrillar Adenosine triphosphatase (ATPase) and alpha glycerophosphate dehydrogenase were studied in serial sections of rat vastus lateralis (red) (RVL), gastrocnemius and diaphragm muscles. Three main fibre-types were distinguished. The Type I fibres of RVL and gastrocnemius muscles fell into two distinct groups: one category-Type IA showed very low ATPase activity. The second category of Type IB fibres displayed moderate ATPase reaction. The Type IA fibres were divisible into two sub-groups when tested for SDH reaction. Type IA₁ fibres possessed a homogenous distribution of diformazan·granules throughout the fibre: Type IA₂ fibres displayed characteristic moth-eaten pattern of diformazan localization. The diaphragm muscle did not show either Type IB or Type IA₂ varieties. The great majority of Type I fibres were sub-type IA₁ in the three fast muscles studied. It is also demonstrated here that an inherent heterogeneity exists between Type I fibres of diaphragm and leg muscles in regard to -GPD localization. This histochemical data emphasizes the fact that subdivision of Type I striated muscle fibres of mammalian animals into two sub-types is only approximate and that a further subcategorization is possible.     

Trophic Assemblages of the Rocky Intertidal Zone in Vostok Bay,Sea of Japan

The ratios between the indices of relative abundance for different trophic life forms have been used to characterize bionomical types of a rocky intertidal zone. It has been shown that the distribution of life forms is determined by the geomorphological peculiarities of the surveyed intertidal areas. A brief critical historical review has been provided of the terms bionomy and bionomical type.     

Enrichment and efficient screening of ES cells containing a targeted mutation: the use of DT‐A gene with the polyadenylation signal as a negative selection maker

Gene targeting in embryonic stem (ES) cells via homologous recombination can occur at very low frequency. In order to enrich homologous recombinants before screening, a negative selection marker, such as thymidine kinase (TK) and diphtheria toxin A fragment (DTA), has been commonly used. In this study, we developed a negative selection marker using DTA gene with polyadenylation signal and it was designated DTApA. To determine the difference in targeting efficiency of the negative selections, we constructed three different targeting vectors for each negative selection (first, TK at the 3 end, second, TK at both the 5 and 3 ends <2 X TK>, and third, DTApA at the 5 end of the homologous sequences). Gene targeting experiments using these constructs clearly showed that negative selection using DTApA was more efficient than that using TK for homologous recombination and that negative selection using DTApA was as efficient as that using 2 X TK. Considering the fact that the use of DTApA is more convenient for construction of targeting vectors than that of 2 X TK, DTApA is an efficient negative selection marker.In addition, we examined long and accurate PCR (LAPCR) for screening gene targeted clones. The use of LAPCR with genomic DNAs from ES cell clones facilitated simple detection of homologous recombinants, suggesting that the screening with LAPCR is compatible with the use of longer homologous sequences of both arms in vector design. Our results indicate that the use of DTApA for negative selection together with the application of LAPCR for screening ensures efficient and timesaving screening for homologous recombinants.