Characterization of two β-glucosidase genes fromClostridium thermocellum

Summary Two -glucosidase genes, designatedbglA andbglB, were isolated from a gene bank ofClostridium thermocellum DSM 1237. The coding sequences forbglA andbglB were located on non-homologous DNA fragments of 3.2– and 3.4-kb, respectively. Both genes direct inEscherichia coli the synthesis of cytoplasmic -glucosidases, which differ with respect to substrate specificity and temperature profile. The properties of thebglA-encoded -glucosidase A closely resemble that of a -glucosidase previously isolated fromC. thermocellum cultures.     

Linear differentation of cereal chromosomes

Summary The BSG test was used in an investigation of the linear differentiation in rye variety Zhitkinskaya, common wheat variety Aurora and two secondary Triticale namely AD-196 and F-1239.Chromosomes of Aurora variety and wheat chromosomes within Triticale may be easily divided into constant and variable chromosomes as described previously (lordansky et al. 1977; Zurabishvili et al 1977).It is necessary to emphasize that the diversity of variable chromosomes underlies karyotypical polymorphism within wheat and Triticale species. The polymorphism observed exists in parallel with strict homomorphism of homologous chromosomes.In IB chromosomes of Aurora variety, the short arm is substituted by the rye chromosome arm. The karyotype of Triticale AD-196 consists of six pairs of rye chromosomes and fifteen pairs of wheat chromosomes.     

Organogenesis versus embryogenesis from long-term suspension cultures of cucumber (Cucumis sativus L.)

Suspension cultures were initiated from leaf explant-derived callus of cucumber,Cucumis sativus cv. Hokus, and maintained under two different conditions; (I) continuously in medium with 5 M 2,4-D + 5 M BA, and (II) alternately three cultures in medium containing 5 M NAA + 5 M BA and one culture in 5 M 2,4-D + 5 M BA. After plating on solid medium with 0.5 M KIN + 0.1 M IAA, suspension aggregates from long-term culture in medium with 2,4-D developed into callus, and subsequently formed somatic embryos. These embryos, however, hardly developed into plants. They showed growth arrest and several structural abnormalities. In contrast, organogenesis took place when suspension aggregates from NAA containing medium were plated on solid medium with 0.5 M KIN + 0.1 M IAA. Numerous adventitious buds were regenerated, which quite normally developed into plants. Sucrose at low concentration of 1% improved plant formation. On the average thirty complete plants were obtained from each ml of suspension. It is discussed why adventitious buds develop into plants so well, whereas somatic embryos are prone to growth arrest and abnormal development.Abbreviations BA 6-benzylaminopurine - KIN kinetin - IAA indole-3-acetic acid - NAA 1-naphthaleneacetic acid - 2,4-D 2,4-dichlorophenoxyacetic acid     

Sterol conjugates of two phenotypically different calli of Beta vulgaris

Steryl glycosides are the predominant form of sterol at 88% of the total sterol in non-betalain producing calli of Beta vulgaris. The total sterol decreases and sterol form shifts from steryl glycosides to 97% free sterol upon the transition of non-betalain to betalain producing calli. A substantial decrease in stigmasterol (24--ethylcholesta-5,22E-dien-3-ol) and sitosterol (24-ethylcholest-5-en-3-ol) levels is observed during this transition, and alters the ratio of ⁷:⁵ sterols. Spinasterol (24- ethyl-5-cholesta-7,22E-dien-3-ol) is the dominant sterol at 43% and 95% of the total sterol in non-betalain producing and betalain producing calli. The level of 22-dihydrospinasterol (24-ethyl-5-cholest-7-en-3-ol) is reduced in both calli to 3% from 25% in leaves. Lanosterol (4,4,14-trimethyl-cholesta-8(9),24-dien-3-ol) and cycloartenol (9,19-cyclopropyl-4,4,14-trimethyl-cholest-24-en-3-ol) were identified in betalain and nonbetalain producing callus respectively.     

Theoretical studies on the necessary number of components in mixtures

Summary Theoretical studies on the optimal numbers of components in mixtures (for example multiclonal varieties or mixtures of lines) have been performed according to phenotypic yield stability (measured by the parameter variance). For each component i, i = 1, 2,..., n, a parameter u_i with 0 u_i 1 has been introduced reflecting the different survival and yielding ability of the components. For the stochastic analysis the mean of each u_i is denoted by u ₁ and its variance by _i ² For the character total yield the phenotypic variance V can be explicitly expressed dependent on 1) the number n of components in the mixture, 2) the mean of the _i ² 3) the variance of the _i ² 4) the ratio and 5) the ratio _i ² /² where denotes the mean of the u _i and _u ² is the variance of the u _j. According to the dependence of the phenotypic stability on these factors some conclusions can be easily derived from this V-formula. Furthermore, two different approaches for a calculation of necessary or optimal numbers of components using the phenotypic variance V are discussed: A. Determination of optimal numbers in the sense that a continued increase of the number of components brings about no further significant effect according to stability. B. A reduction of b % of the number of components but nevertheless an unchanged stability can be realized by an increase of the mean of the u _i by 1% (with and _u ² assumed to be unchanged). Numerical results on n (from A) and 1 (from B) are given. Computing the coefficient of variation v for the character total yield and solving for the number n of components one obtains an explicit expression for n dependent on v and the factors 2.-5. mentioned above. In the special case of equal variances, _i ² = _o ² for each i, the number n depends on v, x = (₀/)² and y = (_u/)². Detailed numerical results for n = n (v, x, y) are given. For x 1 and y 1 one obtains n = 9, 20 and 79 for v = 0.30, 0.20 and 0.10, respectively while for x 1 and arbitrary y-values the results are n = 11, 24 and 95.This publication is an extended version of a lecture given at the 1984-EUCARPIA meeting (Section Biometrics in Plant Breeding) in Stuttgart-Hohenheim (Federal Republic of Germany)     

Synthesis of Nα,Nδ-dicarbobenzoxy-L-ornithyl-β-alanine benzyl ester,a derivative of salty peptide,in 1,1,1-trichloroethane with polyethylene glycol-modified papain

Summary N,N-Dicarbobenzoxy-L-ornithyl--alanine benzyl ester, a derivative of salty peptide, was synthesized from N,N-dicarbobenzoxy-L-ornithine ethyl ester and -alanine benzyl ester in 1,1,1-trichloroethane using papain modified with polyethylene glycol. The peptide bond formation proceeded in a transparent organic solvent at room temperature and the product was obtained as precipitates from the reaction system.     

Die Feinstruktur der Zirbeldrüse normaler,trächtiger und experimentell beeinflußter Meerschweinchen

Zusammenfassung In der Meerschweinchenzirbeldrüse lassen sich elektronenmikroskopisch helle und dunkle Pinealzellen sowie einzelne Gliazellen nachweisen. In den bei weitem überwiegenden hellen Pinealzellen zeichnet sich ein Teil der vesicle-crowned rodlets (VCR) durch lokale Auftreibungen aus. Von VCR deutlich abzugrenzen sind die vesicle-crowned balls (VCB). Erstmalig beschrieben wird das Vorkommen von sog. Zylindern, die als Vorstufen von VCB aufgefaßt werden. In den relativ seltenen dunklen Pinealzellen, die sich durch chromatinreiche Kerne und elektronendichtes Zytoplasma auszeichnen, sind Vesikel, VCR, VCB und Zylinder seltener als in hellen Pinealzellen. Die reichlich vorhandenen marklosen Nervenfasern finden sich vor allem in perivasculären Räumen, seltener im Parenchym. Synapsen zwischen Nerven und Pinealzellen wurden nicht beobachtet. In den Zirbeldrüsen trächtiger Meerschweinchen zeichnen sich in der 2. Hälfte der Tragzeit die hellen Pinealzellen durch stärkere Lappung der Kerne, gehäuftes Auftreten von laktiven Zonen, Vermehrung von Mitochondrien, glattem ER, agranulären Vesikeln, VCR, VCB und Zylindern aus. Die dunklen Pinealzellen nehmen während der Tragzeit an Zahl zu. Post partum bilden sich diese Veränderungen innerhalb einer Woche zurück. Längerer Aufenthalt der Tiere in Dunkelheit führt zu einer Aktivierung der hellen Pinealzellen mit auffallender Vermehrung der VCR und zu einer Zunahme der dunklen Zellen. Unter Dauerbelichtung kommt es in den hellen Zellen zu einer Abnahme fast aller Zellorganellen und zu einer starken Vermehrung der VCR, die nach 70 Tagen auch Formveränderungen aufweisen. Nach Reserpinbehandlung beobachtet man eine Verminderung und degenerative Veränderungen der VCR. Es wird diskutiert, daß die VCR als prae- bzw. postsynaptische Strukturen der Erregungsübertragung von Nerven zu Pinealzellen bzw. von Pinealzellen untereinander dienen könnten.

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The fine structure of the pineal gland of normal, pregnant and experimentally affected guinea-pigs

Summary By means of electron microscopy light and dark pinealocytes can be distinguished in the guinea-pig pineal gland. Glial cells are rare. In the light pinealocyte. the most frequent cell type, some vesicle-crowned rodlets (VCR) show circumscribed thickenings. From these structures vesicle-crowned balls (VCB) have to be clearly distinguished. Furthermore cylinders occur, which, it is suggested, are precursors of VCB. Dark pinealocytes characterized by chromatin-rich nuclei and electron-dense cytoplasm are rare and contain fewer vesicles, VCR, VCB and cylinders than light pinealocytes. Numerous non-myelinated nerve fibres are situated within perivascular spaces, a few also in the parenchyma. Synapses between nerve fibres and pinealocytes were not observed. In the pineal gland of pregnant guinea-pigs the following changes can be observed in the second half of gestation. The light cells show many nuclear indentations and an increase of active zones, mitochondria, smooth ER, agranular vesicles, VCR, VCB, and cylinders respectively. The dark cells increase in number. After birth these changes reverse to normal within one week. Constant darkness leads to an activation of the light cells accompanied by an increase of the VCR and to an increase in number of the dark cells. Under constant illumination the light cells show a decrease of their organelles and a strong increase of the VCR. After 70 days the VCR also show a change in shape. Following reserpine treatment the VCR decrease in number and show signs of degeneration. It is discussed that the VCR function as pre- or postsynaptic structures and that they are involved either in transmitting impulses from nerve fibres to pinealocytes or from one pinealocyte to the other.

Untersuchung unter Leitung von Univ.-Doz. Dr. L. Vollrath.     

Conservation status of parrot populations in an Atlantic rainforest area of southeastern Brazil

A census of four species of syntopic parrots was carried out using distance sampling methods on São Sebastião island, SE Brazil. Most of the 33593 ha island is covered by mature and secondary Atlantic rainforest. Almost 80% of these forests are within the Ilhabela Park. Although the species counted have marked differences in size and weight, density (individuals/km²) and estimated population size in 23500 ha of well-preserved forests were similar: Amazona farinosa (13.82±5.94; 3247±1395), Pionus maximiliani (15.79±7.04; 3712±1654), Brotogeris tirica (15.05±4.87; 3537±1143) and Pyrrhura frontalis (13.06±5.53; 3068±1298). Encounter rates of Forpus crassirostris and Pionopsitta pileata were very low, which suggests that there is only a small population of these species on the island. The São Sebastião forests still support healthy populations of parrots. Although woodpecker population estimates on the island are large enough to provide nesting sites for parrots, competition for holes with other secondary cavity nesters such as toucans, flycatchers and tytiras, and the selective cutting of dead trees for canoe construction, which is a common practice on the island, may limit hole availability for parrots.     

Induction of β galactosidase in the yeast Kluyveromyces lactis

Summary The inductive effect of lactose, -methyl-thio-D-galactopyranoside, (TMG) and glucose on galactosidase synthesis in Kluyveromyces lactis has been studied. Whereas TMG gave a five fold stimulation of the rate of -galactosidase synthesis, lactose only gave a small stimulation. Glucose caused represssion at levels above 10^-3M but stimulated -galactosidase synthesis when added at lower concentrations.     

The adaptive significance of cultural behavior: Comments and reply

Summary Fundamentally, theoretically, there is only one process underlying genetic and cultural evolution: natural selection. Organism fitness-enhancement (adaptive significance) is one of its practical mechanisms; group formation and maintenance is another, often but not always through fitness-enhancement; and need-fulfillment is still another. If Durham can accept that formulation, and switch from organism-thinking to instruction-thinking (Cloak, 1975: 178), he will free himself from two handicaps: First, he can forget his worries about reductionism and determinism (1976a: 100, 101). Under this general theory of natural selection, cultural evolutionis biological evolution, continued by other (nongenetic) means. Second, he will spare himself the appearance of anthropomorphism, mentalism, and wishy-washiness attendant on his discussion of kinds of significance, other than adaptive significance, of cultural behaviors (1976a: 102–106, 115).     

Organization of α-chain genes among Hb G-Philadelphia heterozygotes in association with Hb S, β-thalassemia,and α-thalassemia-2

The percentages of the -chain variant Hb G-Philadelphia (Hb G) or ₂ 68 AsnLys₂ were evaluated in 84 adult and 18 newborn heterozygotes. These included members of three families who were studied in more detail by nucleic acid hybridization techniques. The adult heterozygotes fell in two categories, one with a higher proportion of Hb G [46.5±1.0% (SD), N=21] and another with lower values (33.9±3.4%, N=63). Among the newborn heterozygotes, two babies fell in the category with the higher proportion of Hb G while 16 babies gave values between 25 and 34%. Studies of -chain gene organization on the parents of one neonate with a Hb G level of 27% at birth and 37% at 8 months excluded the presence of chromosomes with triplicated -chain genes which could lead to the ^0G/ genotype. Rather, these studies on five Hb G heterozygotes from three families confirmed the linkage between Hb G and a specific type of -thalassemia-2 associated with the presence of a 16-kbp Bgl II fragment which most probably carries the ^G locus since it has been found in 19 Hb G heterozygotes studied to date. The presence of an -thal-2 heterozygosity and three -chain genes (^0G/) was confirmed among Hb G heterozygotes with lower proportions of this variant. It is likely that the even lower values found in some newborn could arise through defective assembly of ^G- dimers. The presence of an -thal-2 homozygosity and two active -chain genes, one on each chromosome (^0G/⁰), was confirmed among heterozygotes with the higher proportion of Hb G. One of each of these categories was present in each of the three families investigated. This type of variability in the number of active -chain genes due to a heterozygosity or a homozygosity for -thalassemia-2 explains the trimodality of Hb S percentages among heterozygotes and the atypical hematological or biosynthetic features among patients with -thalassemia and sickle-cell syndromes.This research was supported by USPHS Research Grants HLB-05168 and HLB-15158 and by designated research funds of the Veterans Administration. This is Contribution No. 0693 of the Department of Cell and Molecular Biology, Medical College of Georgia, Augusta.     

Development of an L6 myoblast in vitro model of moniliformin toxicosis

L6 myoblasts were used as an in vitro model to investigate the role of moniliformin and its interaction with monensin in turkey knockdown syndrome and sudden death syndromes in poultry. Cell viability and microscopic and ultrastructural alterations noted in L6 myoblasts cultured in the presence of moniliformin (0.0–0.3 g/l) were compared to those observed in parallel cultures also containing one of the following compounds: selenium (0–0.004 ng/l), thiamine (0–0.3 g/l), or pyruvate (0–0.46 g/l). Marked dilation of the RER, membranous whorls, glycogen deposition, membrane-bound cytoplasmic inclusions and necrosis were observed in myoblasts exposed to 0.03/2-0.30 g moniliformin/l medium. Supplementation of medium with thiamine and pyruvate, or selenium, provided significant protection to cells exposed to 0.0–0.3 g/l or 0.0–0.15 g moniliformin/l, respectively. Dose-dependent differences in protein and ATP production were not detected. Myoblasts grown in medium containing 0–0.15 g moniliformin/l and 7.5–50.0 M A23187, beauvericin or monensin had degrees of cytotoxicity similar to parallel cultures receiving only an ionophore. L6 myoblasts were a useful model of moniliformin toxicosis. The findings of this study suggest cytotoxicity due to moniliformin in L6 myoblasts may be due in part to oxidative damage and altered pyruvate metabolism, and that moniliformin does not predispose myoblasts to ionophore toxicosis. This study supports the results of in vivo investigations in poultry that moniliformin and monensin do not act synergistically to induce knockdown or monensin toxicosis.     

Conformational transmission in ATP synthase during catalysis: Search for large structural changes

Escherichia coli ATP synthase has eight subunits and functions through transmission of conformational changes between subunits. Defective mutation at Gly-149 was suppressed by the second mutations at the outer surface of the subunit, indicating that the defect by the first mutation was suppressed by the second mutation through long range conformation transmission. Extensive mutant/pseudorevertant studies revealed that / and / subunits interactions are important for the energy coupling between catalysis and H⁺ translocation. In addition, long range interaction between amino and carboxyl terminal regions of the subunit has a critical role(s) for energy coupling. These results suggest that the dynamic conformation change and its transmission are essential for ATP synthase.     

Problems in estimating growth rates of marine phytoplankton from short-term14C assays

Growth rate estimates () of phytoplankton populations that were sampled from nitrogen-limited continuous cultures and then incubated for short durations in batch culture with added¹⁴C-HCO₃ ^– were significantly different than steady-state growth rates () for 3 of 5 marine phytoplankton species. Two diatoms,Thalassiosira weissflogii andChaetoceros simplex, displayed virtually identical growth rates (=) over a wide range of, whereas for a third diatom,Phaeodactylum tricornutum, was overestimated by an average of 40% compared to. In contrast, was underestimated by the¹⁴C technique for the two remaining species: up to 40% at a steady-state of 1.0 day^–1 for the chlorophyteDunaliella tertiolecta and up to 100% at of 1.4 day^–1 for the haptophytePavlova lutheri. For the latter two species the divergence between and appeared to increase with increasing steady-state. A simple model of labeled and total carbon flow between the aqueous phase and cellular biomass was constructed to demonstrate that respiration was negligible when=, but was significant when>. In the cases in which<, a rapid physiological alteration presumably took place once the steady state was disturbed and cells were placed in the incubation chambers, which perhaps was related to the nutritional state of the cultures at the time of sampling. Questions thus are raised regarding our ability to measure accurately primary productivity from shipboard experiments with confined samples of phytoplankton from nutrient-impoverished waters that probably are less hardy than the laboratory cultures used in these studies.     

Zur Dynamik feiner Pseudopodien von Hochmoor-Amöben

Zusammenfassung Es werden verschiedene Vorgänge an sehr zarten Pseudopodien beschrieben und mit Hilfe der an submikroskopischen Fibrillen (schraubig gebauten Mikrofibrillen und Mikroröhrchen der Elektronenmikroskopie) auftretenden Torsionsspannungen und Rotationen gedeutet.

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On the dynamics of slender pseudopodia of bog amebae

Summary Different processes which occur on very slender pseudopods are described. They are interpreted with the help of torsional forces and revolutions of the submicroscopic fibrils (helical microfibrils and microtubules in electron microscopy).

     

Increased radioresistance of cells in cultured multicell spheroids. I. Dependence on cellular interaction

Summary Monolayers of six different cell lines were investigated with respect to ionic coupling using micro-electrode techniques. In parallel, survival after Co--irradiation of monolayer- and spheroid cultures of these lines was compared. It was found that spheroids of coupled cell lines were more radioresistant than monolayers (contact effect). However, cell coupling did not enhance the survival of monolayers over single cells. This suggests that the contact effect is a tissue phenomenon requiring cellular interaction but is expressed only under conditions of three-dimensional growth.     

T-cell repertoire in a strain of transgenic C57BL/6 mice with the HLA-DRA gene on the X-chromosome

We have established a strain of transgenic mice in which the HLA-DRA gene was integrated into the X-chromosome and the xenogeneic mixed isotype molecule, DRE^b, was expressed in a cell type-specific manner, although the transgenic DRA gene contained only 268 base pairs of the 5-flanking region. The DRE^b molecules expressed in the transgenic mice functioned as major histocompatibility complex (MHC) class II to select T-cell repertoire, and to stimulate mixed lymphocyte reaction. In female transgenic mice homozygous for HLA-DRA (DR-B6-F-homo) and male transgenic mice (DR-B6-M), DRE^b molecules were expressed in almost all of the MHC class II A^b-positive cells. In contrast, the expression of DRE^b molecules in female transgenic mice hemizygous for HLA-DRA (DR-B6-F-hemi) was found only in part of the A^b positive cells, and the proportion of cells expressing the DRE^b molecules varied due to random inactivation of one of the X-chromosomes. Clonal deletions of the T cells and mature thymocytes bearing Tcrb-V5 and Tcrb-V11, which are eliminated from the peripheral repertoire in mice expressing self-superantigen and MHC class II E molecules, were incomplete in DR-B6-F-hemi as compared with those in DR-B6-F-homo, and were correlated with the proportion of DRE^b-positive spleen cells. These observations suggested that the number of bone marrow-derived cells expressing DRE^b molecules was critical for clonal deletions of Tcrb-V5⁺ and Tcrb-V11⁺ T cells in the thymus.     

Δ5-3β-hydroxysteroid dehydrogenase/Δ5-Δ4-ketosteroid isomerase (3β-HSD), a possible enzyme of cardiac glycoside biosynthesis,in cell cultures and plants of Digitalis lanata EHRH

Summary The in vitro transformation of pregnenolone into progesterone in Digitalis lanata tissues was shown to be catalyzed by a 3-hydroxysteroid dehydrogenase/ketosteroid isomerase (3-HSD). Product formation was monitored by HPLC. The enzyme could be partially characterized and 3-HSD activities were measured in various Digitalis lanata tissues and in cell cultures of other plant species. Since no correlation was observed between biosynthetic competence of the tissue and 3-HSD activity, it was concluded that this enzyme does not play a major role in regulating cardenolide biosynthesis.Abbreviations BA benzyladenine - 2,4-D 2,4-dichlorophenoxyacetic acid - DMSO dimethyl sulfoxide - DMF N,N-dimethylformamide - IAA indole-3-acetic acid     

Immobilized Lotus tetragonolobus agglutinin binds oligosaccharides containing the Lex determinant

A defined set of oligosaccharides and glycopeptides containing -linked fucose were used to examine the specificity of the immobilized fucose-binding lectin Lotus tetragonolobus agglutinin (LTA1), also known as lotus lectin. Glycans containing the Lewis x determinant (Lex) Gal1-4[Fuc1-3]GlcNAc1-3-R were significantly retarded in elution from high density LTA-Emphaze columns. The lectin also bound the fucosylated lacdiNAc trisaccharide GalNAc1-4[Fuc1-3]GlcNAc. The lectin did not bind glycans containing either sialylLex or VIM-2 determinants, nor did it bind the isomeric Lea, Gal1-3[Fuc1-4]GlcNAc-R. Although 2-fucosyllactose Fuc1-2Gal1-4Glc) was retarded in elution from the columns, larger glycans containing the H-antigen Fuc1-2Gal1-3(4)GlcNAc-R interacted poorly with immobilized LTA. Our results demonstrate that immobilized LTA is effective in isolating glycans containing the Lex antigen and is useful in analyzing specific fucosylation of glycoconjugates. Abbreviations: LTA, Lotus tetragonolobus agglutinin; UEA-1, Ulex europaeus agglutinin-I; LNT, AAL, Aleuria aurantia agglutinin; Gal1-3GlcNAc1-3Gal1-3Glc; LNnT, Gal1-4GlcNAc1-3Gal1-3Glc; Lex, Lewis x antigen; Lea, Lewis a antigen; GDPFuc, guanosine 5-diphosphate--L-fucose