Assignment of human ferritin genes to chromosomes 11 and 19q13.3→19qter

Summary Extracts of hamster-human and mouse-human hybrids, some with translocations involving chromosome 19, have been assayed for both human spleen ferritin (rich in L subunits) and human heart ferritin (rich in H subunits). Hybrid lines retaining part of the long arm of chromosome 19 including the region 19q13.319qter produced human L type ferritin. This confirms the previous assignment of the ferritin gene to chromosome 19 (Caskey et al. 1983). However, lines retaining chromosome 11 were found to contain human H type ferritin suggesting that the gene for the H subunit is on this chromosome. The presence of chromosome 6 was not necessary for the expression of either H or L type human ferritin. It thus seems unlikely that the gene for idiopathic haemochromatosis is a ferritin gene.     

Über das Verhalten von Champignonstämmen in Mischkultur

Zusammenfassung Mischungen aus dem Mycel sweier weißer oder zweier brauner Stämme des Kulturchampignons hatten keinen Einfluß auf den Fruchtkörperetrag.Bei Mischungen aus einem weißen und einem braunen Stamm was dagegen der Ertrag immer niedriger als erwartet und das Mycel verschwand verhältnismäßig schnell. Der Ertragsausfall beruht in der Regel auf einer Unterdrückung des braunen Stammes.Herrn Prof. von Sengbusch zum 65. Geburtstag in Dankbarkeit und Verehrung gewidment.     

Quantitative Veränderungen des Abscisinsäuregehaltes während der Fruchtentwicklung von Solanum lycopersicum L.

Summary The presence of abscisic acid (ABA) in methanol extracts from tomato fruits was determined by thinlayer chromatography, UV absorption, optical rotatory dispersion (ORD) and biological activity in different bioassays. In two growth periods (1968 and 1969), quantitative changes of the ABA content in growing fruits of the variety Moneymaker were measured by Milborrow's racemate dilution technique. The absolute content of ABA (g/l fruit) was increased during fruit development, reached a maximum, and then decreased in ripening fruits. The ABA concentration (g/kg) was also highest in unripe fruits and decreased during ripening. Similar results were obtained with the same variety and with the variety Haubners Vollendung by means of ORD and UV measurement only, without application of Milborrow's technique.     

Quantitative utilization of selected grasses by fall armyworm larvae

Food utilization by larvae of the fall armyworm (Spodoptera frugiperda [J. E. Smith]) (Lepidoptera: Noctuidae), showed greater consumption of corn (Zea mays L.) than pinto bean diet, Tifton 10, or Coastal bermudagrass (Cynodon dactylon [L.] Pers.). Transfer of larvae from diet to susceptible grasses such as corn, Tifton 10 or Coastal produced differences in growth rates as a result of food consumption rates. Transfer of larvae from diet to resistant grasses such as common centipedegrass (Eremochola ophiuroides [Munro] Hack) Tifton 292 bermudagrass, and zoysiagrass (Zoysia sp.) reduced larval growth as a result of low consumption rates and/or greater metabolic expenditures. Larvae initially fed Tifton 10, Coastal, or centipedegrass before feeding on corn grew significantly faster than when they fed continuously on corn.

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Résumé Des chenilles de S. frugiperda ont consommé plus de maïs (Zea mays), que d'un régime à base de Phaseolus vulgaris de la variété Pinto, ou de Tifton 10, ou de Cynodon dactylon de la variété Coastal. Des chenilles, transférées du régime artificiel sur des plantes sensibles comme le maïs, le Tifton 10 ou le Coastal, ont présenté des taux de croissance différents provenant de modifications de leur consommation. Le transfert du régime à des plantes résistantes, telles que Eremochola ophiuroides, Tifton 292, C. dactylon et Zoysia sp., a entraîné une diminution de la croissance larvaire par suite d'une plus faible consommation et/ou de dépenses métaboliques plus élevées. Des chenilles ayant consommé du Tifton 10, du Coastal ou de l'E. ophiuroides avant de consommer du maïs, se sont développées significativement plus vite que celles qui s'étaient alimentées continuellement sur maïs.

     

A use of Ramachandran potentials in protein solution structure determinations

A strategy is developed to use database-derived - constraints during simulated annealing procedures for protein solution structure determination in order to improve the Ramachandran plot statistics, while maintaining the agreement with the experimental constraints as the sole criterion for the selection of the family. The procedure, fully automated, consists of two consecutive simulated annealing runs. In the first run, the database-derived - constraints are enforced for all aminoacids (but prolines and glycines). A family of structures is then selected on the ground of the lowest violations of the experimental constraints only, and the - values for each residue are examined. In the second and final run, the database-derived - constraints are enforced only for those residues which in the first run have ended in one and the same favored - region. For residues which are either spread over different favored regions or concentrated in disallowed regions, the constraints are not enforced. The final family is then selected, after the second run, again only based on the agreement with the experimental constraints. This automated approach was implemented in DYANA and was tested on as many as 12 proteins, including some containing paramagnetic metals, whose structures had been previously solved in our laboratory. The quality of the structures, and of Ramachandran plot statistics in particular, was notably improved while preserving the agreement with the experimental constraints.     

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.     

Physiological consequences of mitochondrial antibiotic-resistant mutations in Paramecium: growth-rates, cytochromic defects and cyanide-insensitive respiration of mutant and erythromycin-treated wild-type strains.

Summary A set of mitochondrial antibiotic-resistant mutants of Paramecium have been analyzed with respect to their growth-rates, cytochromic content and respiratory properties. The mutants could be arranged in a continuous series ranging from strains equivalent to wild-type to severely affected ones; affected strains display longer generation times, reduced amount of cytochrome oxidase and very high levels of cyanideinsensitive respiration. Perfect phenocopies of the mutants were obtained by treating wild-type cells with low concentrations of erythromycin suggesting that the mutations exert their pleiotrophic effect by perturbating mitochondrial protein synthesis in agreement with the idea that these mutations affect the mitochondrial ribosomes. In the mitochondria of some of the mutants, electrons can be channelled with equal efficiency into the classical cyanide-sensitive pathway and the alternate cyanide insensitive (and SHAM_sensitive) one, providing direct demonstration of the branching of these two respiratory pathways. In the absence of any added inhibitor, however, electrons tend to be channelled in the cyanide-sensitive pathway.All the physiological data fit perfectly the genetic data concerning the stability of the various mutations in mixed mitochondrial populations, i.e., markers that were known to be strongly counter-selected with respect to wild-type in such populations correspond to severely affected strains, while markers that were known to be stable correspond to healthy strains. A more quantitative analysis of the data shows that that there is little or no complementation between wild-type and mutated mitochondria in mixed cells indicating a high extent of functional autonomy of mitochondria in Paramecium.     

Cyclodextrins as a useful tool for bioconversions in plant cell biotechnology

The application of cyclodextrins as precursor solubilizers in biotechnological processes, in which plant cells are involved, is new. In this paper the possibilities for cyclodextrin facilitated bioconversions by freely suspended and/or immobilized plant cells or plant enzymes are demonstrated. After complexation with -cyclodextrin, the phenolic steroid 17-estradiol could be ortho-hydroxylated into a catechol, mainly 4-hydroxyestradiol, by a phenoloxidase from in vitro grown cells of Mucuna pruriens. By complexation with -cyclodextrin the solubility of the steroid increased from almost insoluble to 660 M. In addition, by complexation with -cyclodextrin, a solution of 3 mM coniferyl alcohol could be fed to cell cultures of Podophyllum hexandrum in order to enhance the accumulation of podophyllotoxin. Finally, the glucosylation of podophyllotoxin by cell cultures derived from Linum flavum was investigated. Four cyclodextrins: -cyclodextrin, -cyclodextrin, hydroxypropyl--cyclodextrin and dimethyl--cyclodextrin were used to improve the solubility of podophyllotoxin. Dimethyl--cyclodextrin met our needs the best and the solubility of podophyllotoxin could be enhanced from 0.15 to 1.92 mM. Podophyllotoxin--d-glucoside was formed at a rate of 0.51 mmol l^-1 suspension per day by the L. flavum cells growing in the presence of 1.35 mM podophyllotoxin, complexed with dimethyl--cyclodextrin.Abbreviations DW dry weight - E2 17-estradiol - FW fresh weight - PCV packed cell volume     

Retroviral transduction of interferon-γ cDNA into a nonimmunogenic murinefibrosarcoma: generation of T cells in draining lymph nodes capable of treating established parental metastatic tumor

Gene modification of tumor cells with the cDNA for interferon (IFN) has been shown to increase the immunogenicity of some tumor cells. In order to explore further the possible therapeutic relevance of these previous findings, two clones of the nonimmunogenic MCA-102 fibrosarcoma of C57BL/6 origin were retrovirally transduced with the cDNA encoding murine IFN: 102.4JK (4JK), a clone with relatively high major histocompatibility complex (MHC) class I expression, and 102.24JK (24JK), a clone with low expression of surface MHC class I molecules. Retroviral transduction of tumor cells with the cDNA encoding for IFN resulted in a substantial up-regulation of MHC class I surface expression in the 24JK clone but little change of class I in the 4JK clone. In an attempt to generate antitumor lymphocytes, these gene-modified cells were inoculated into mouse footpads and draining lymph nodes (DLN) were removed, dispersed, and cultured in vitro for 10 days with irradiated tumor cells and interleukin-2. DLN from mice bearing either unmodified tumor or tumor transduced with cDNA encoding neomycin resistance (Neo ^R) or IFN, were used to treat recipients harboring 3-day pulmonary metastases induced by the parental, unmodified tumor. Treatment with DLN cells obtained following the injection of 24JK tumor cells modified with the gene for IFN significantly reduced the number of pulmonary metastases in four separate experiments, compared to groups treated by DLN cells generated from inoculation of either the unmodified, parental 24JK clone or the same clone transduced with theNeo ^R gene only. In contrast, DLN cells induced either by IFN-transduced 4JK (high expression of MHC class I) or an unmodified 4JK tumor (moderate expression of MHC class I) had significant but equal therapeutic efficacy. Although the in vitro growth rate of tumor cell lines was unaffected by the insertion of the mouse IFN cDNA, their in vivo (s.c.) growth rates were significantly slower than those of the nontransduced tumors. Thus, after retroviral transduction of the murine IFN cDNA into a nonimmunogenic tumor with a very low level of surface expression of MHC class I, modified tumor cells could elicit therapeutic T cells from DLN capable of successfully treating established pulmonary metastases upon adoptive transfer. This strategy significantly confirms previous observations on the potential therapeutic effects of gene modification of tumor cells with IFN and extends the realm of therapeutic possibilities to include the use of DLN cells for the development of T-cell based immunotherapies against nonimmunogenic human tumors.     

A solvable selfreproductive hypercycle model for the selection of biological molecules

Summary By introducing a modified enzyme coupling in Eigen's hypercycle equations we have produced an exactly solvable selfreproductive hypercycle model. The model shows explicitly how collections of different macromolecular information carriers may coexist, through internal couplings, in the presence of constraints. We show how the selective value concept is manifested in the model and give simple criteria for selection among competing hypercycles.     

Identification of a specific protein in the mitochondrial fraction of the rat heart whose phosphorylation is inhibited by taurine

Summary It was previously reported that the mitochondrial fraction of the rat heart contained a specific protein with a molecular weight of approximately 44kDa whose phosphorylation was inhibited by taurine (Lombardini,1994a). Isolation of the 44kDa phosphoprotein on a 1-dimensional polyacrylamide gel using traditional glycine buffers followed by re-electrophoresing the cut out proportion of the gel which corresponds to the 44kDa protein on a tricine-buffered gel resulted in sufficient pure protein for sequence analysis. The results indicate that the 44kDa phosphoprotein is pyruvate dehydrogenase.     

Inactivation of voltage-dependent calcium current in an insulinoma cell line

We have studied the mechanism of Ca current inactivation in the -cell line HIT-T15 by conventional and perforated patch recording techniques, using two pulse voltage protocols and a combination of current and tail current measurements. In 5 mM Ca, from a holding potential of - 80 mV, the maximum current showed a complex time course of inactivation: a relatively fast, double exponential inactivation (_h1 12 ms and _h2 60 ms) and a very slowly inactivating component ( > 1 s). The faster component (_h1) was due to the voltage-dependent inactivation of a low-threshold-activated (LVA), T-type current, which deactivates more slowly ( 3–5 ms) than the other components ( 0.2–0.3 ms). The intermediate component (_h2) was due to the Ca-dependent inactivation of a portion of the high-threshold-activated (HVA) current. A saturating dose of the dihydropyridine (DHP) nifedipine (10 M) did not affect the LVA current, but inhibited by 68 ± 5% the transient, Ca-sensitive portion of the HVA current and by 33 ± 12% the long lasting component. We suggest that three components of the calcium current can be resolved in HIT cells and the main target of DHPs is a HVA current, which inactivates faster than the DHP-resistant HVA component and does so primarily through calcium influx. Correspondence to: C. Marchetti     

Studies on imperfect fungi I. Impact of component stereo-isomers on the synthesis of oligosaccharides from sucrose by pathogenic fungi

Summary The impact of extraneously furnished D(+)-glucose and D(–)-fructose, on the synthesis of new oligosaccharides from sucrose solution byC. lycopersici andF. semitectum, is presented in the present paper. The results are significant, in that they show, that the behaviour of the organisms is determined by the shell-wrapped molecular memory enshrined in the nucleus of the cell; and the signal of optical rotatory power is interpreted by the operator-structural genes determining the configuration of the enzyme-fructosidase.     

Dual-mode potential oscillations on an immobilized acetylcholinesterase membrane system

We have studied by numerical stimulations the effects of varying external salt concentration on a multibarrier kinetic model of a synthetic membrane system bearing AChE and BSA. For certain parameters, we find a spiky high frequency modulation imposed on a bursting low frequency carrier. In the absence of added salt, oscillations with only one mode arise, consistent with the experimental result of Friboulet and Thomas. From this study, we suggest how periodic ionic permeability changes of ions may arise in excitable membranes.     

On the signal transducing mechanisms involved in the synergistic interaction between interleukin-1 and bradykinin on prostaglandin biosynthesis in human gingival fibroblasts

Recombinant human interleukin-1 (IL-1) and bradykinin (BK) synergistically stimulate prostaglandin E₂ (PGE₂) formation in human gingival fibroblasts cultured for 24 h. Neither BK or IL-1 per se, nor their combinations, caused any acute stimulation of cellular cyclic AMP accumulation. BK, but not IL-1, caused a rapid, transient rise of intracellular Ca²⁺ concentration ([Ca²⁺]_i), as assessed by recordings of fura-2 fluorescence in monolayers of prelabelled gingival fibroblasts. IL-1 did not change the effect of BK on [Ca²⁺]_i. Ionomycin and A 23187, two calcium ionophores, synergistically potentiated the stimulatory effect of IL-1 on PGE₂ formation. Three different phorbol esters known to activate protein kinase C also synergistically potentiated the action of IL-1 on PGE₂ formation. Exogenously added arachidonic acid significantly enhanced the basal formation of PGE₂. In IL-1 treated cells, the enhancement of PGE₂ formation seen after addition of arachidonic acid, was synergistically upregulated by IL-1. These data show that i) the synergistic interaction between IL-1 and BK on PGE₂ formation is not due to an effect linked to an upregulation of cyclic AMP or [Ca²⁺]_i; ii) the signal transducing mechanism by which BK interacts with IL-1, however, may be linked to a BK induced stimulation of [Ca²⁺]_i and/or protein kinase C; iii) the mechanism involved in the action of IL-1 may, at least partly, be due to enhancement of the biosynthesis of prostanoids mediated by an upregulation of cyclooxygenase activity.     

High frequency somatic embryogenesis and plant regeneration from papaya hypocotyl callus

High frequency somatic embryogenesis in papaya (Carica papaya L.) tissue cultures was achieved by culturing hypocotyl sections from ten-day-old seedlings on half-strength Murashige and Skoog salts (MS) medium containing modified MS vitamins, 2.3 to 112.5 M 2,4-dichlorophenoxyacetic acid (2,4-d), 400 mg l^-1 glutamine, and 6% sucrose. Four hermaphroditic Hawaiian cultivars produced embryogenic calluses after ten to 14 weeks of culture at 27°C in the dark. Efficiency in embryogenic response of genotypes differed, Kapoho > Sunset > Sunrise > Waimanalo. The frequency of embryogenesis in induction medium containing 4.5 M 2,4-d was lowest with 3% sucrose and highest with 7% sucrose. Somatic embryos developed directly from embryogenic calluses on induction medium, or, more often, they differentiated from calluses subcultured on a medium devoid of growth regulators. Between 50 and 500 embryos were produced from each 2-mm hypocotyl section after at least two months on induction medium and two months on maturation medium. Embryos subsequently developed into normal-looking plants on MS medium. Shoot cuttings from germinated embryos and micropropagated plants were rooted with 5.0 M indole-3-butyric acid (IBA), grown in the greenhouse, and transferred to the field.Journal Series no. 3732 of the Hawaii Institute of Tropical Agriculture and Human Resourees     

Guinea Pigs as a Nontransgenic Model for APP Processing in Vitro and in Vivo

Alzheimer's disease (AD) is characterized, amongst others, by the appearance of vascular and parenchymal -amyloid deposits in brain. Such aggregates are mainly composed of -amyloid peptides, which are derived by proteolytic processing of a larger amyloid precursor protein (APP). APP is highly conserved among mammalian species, but experimental studies in rodents are often hampered by the humble APP-processing in the amyloidogenic pathway and by the inability of rodent -amyloid peptides to form higher molecular aggregates such as soluble oligomers and insoluble -amyloid plaques. Thus, there is need for in vitro and in vivo model systems that allow identification of factors that increase amyloidogenic APP processing and accelerate -amyloid plaque formation and testing the potency of pharmacological manipulations to ameliorate -amyloid load in brain. Transgenic mice that overexpress human APP containing AD-associated mutations that favor the amyloidogenic pathway of APP processing represent such a model. However, mutations of the APP gene are not frequent in AD and, therefore, the mechanisms of -amyloid plaque formation, the composition of -amyloid plaques, and the accompanying tissue response in brain of these animals may be different from that in AD. In contrast, guinea pigs express -amyloid peptides of the human sequence and appear to represent a more physiological model to examine the long-term effects of experimental manipulations on APP processing and -amyloid plaque formation in vivo. Additionally, APP processing in guinea pig primary neuronal cultures has been shown to be similar to cultures of human origin. In this article we highlight the advantages and limitations of using guinea pigs as experimental models to study APP processing.     

Unusual amino acids VIII. Asymmetric hydrogenation of some heteroaryl-N-CBZ and N-BOC aminocinnamic acid derivatives

Summary (Z)--[(Benzyloxy)- or (tert.-butyloxy)carbonylamino]- (thienyl)-or (furyl)-acrylic acids and their esters were prepared by known methods and hydrogenated to the corresponding optically active alanine derivatives with optical yields in the range of 58–93% ee using the cationic rhodium complex of PROPRAPHOS.     

Do Casuarina cunninghamiana seedlings dependent on symbiotic N2 fixation have higher phosphorus requirements than those supplied with adequate fertilizer nitrogen?

P requirements of Casuarina cunninghamiana seedlings inoculated with Frankia and reliant on symbiotic N2 fixation were compared with those of uninoculated seedlings grown with 4 levels of fertilizer N in a solution culture system.Growth responses to increasing P supply depended on the N treatment that had been imposed. At the two lowest levels of N supply (0 t M and 100 t M) growth was relatively poor and there was no response to increasing P supply above 10 t M and 50 t M P respectively. In contrast, inoculated seedlings and those with higher levels of combined N (500 t M and 1000 t M) had significantly greater shoot dry weights (except at 0.1 t M P), and they responded to increasing P supply to between 50 and 100 t M P. At each level of P supply, the dry weights of seedlings in these 3 N treatments were similar. Nitrogen concentrations in shoots of seedlings provided with fertilizer N decreased with increasing P supply to 50 t M and then remained unchanged. In contrast, N concentrations in shoots of inoculated seedlings increased with increasing P supply to 25 t M P. At the 2 highest levels of P supply, N concentrations in shoots of inoculated plants were significantly higher than those in seedlings provided with fertilizer N. In all N treatments, P concentrations in shoots increased with increasing P supply; concentrations were similar in inoculated seedlings and those with the 2 highest levels of fertilizer N across all levels of P supply.Alleviation of P deficiency in inoculated seedlings increased nodule number, nodule dry weight, N2 fixation g-1 nodule dry weight (nodule 'efficiency'), P concentration in nodules, proportion of total seedling biomass allocated to nodules and average nodule size. However, all these parameters reached their maximum values at levels of P supply at, or below, those required for maximum host-plant growth (50 to 100 t M P).The results indicate that the P requirement for host plant growth per se is similar to, or higher than, that required for symbiotic N2 fixation processes.     

Charles Darwin's theory of evolution: A review of our present understanding

The paper characterizes Darwin's theory, providing a synthesis of recent historical investigations in this area. Darwin's reading of Malthus led him to appreciate the importance of population pressures, and subsequently of natural selection, with the help of the wedge metaphor. But, in itself, natural selection did not furnish an adequate account of the origin of species, for which a principle of divergence was needed. Initially, Darwin attributed this to geographical isolation, but later, following his work on barnacles which underscored the significance of variation, and arising from his work on botanical arithmetic, he supposed that diversity allowed more places to be occupied in a given region. So isolation was not regarded as essential. Large regions with intense competition, and with ample variation spread by blending, would facilitate speciation. The notion of place was different from niche, and it is questioned whether Darwin's views on ecology were as modern as is commonly supposed. Two notions of struggle are found in Darwin's theory; and three notions of variation. Criticisms of his theory led him to emphasize the importance of variation over a range of forms. Hence the theory was populational rather than typological. The theory required a Lamarckian notion of inheritable changes initiated by the environment as a source of variation. Also, Darwin deployed a use/habit theory; and the notion of sexual selection. Selection normally acted at the level of the individual, though kin selection was possible. Group selection was hinted at for man. Darwin's thinking (and also the exposition of his theory) was generally guided by the domestic-organism analogy, which satisfied his methodological requirement of a vera causa principle.