Microbial transformation of ginsenoside Rb1 by Rhizopus stolonifer and Curvularia lunata

Of 49 microbial strains screened for their capabilities to transform ginsenoside Rb₁, Rhizopus stolonifer and Curvularia lunata produced four key metabolites: 3-O-[-d-glucopyranosyl-(1,2)--d-glucopyranosyl]- 20-O-[-d-glucopyranosyl]-3,12, 20(S)-trihydroxydammar-24-ene (1), 3-O-[-d-glucopyranosyl-(1,2)--d- glucopyranosyl]-20-O-[-d-glucopyranosyl]-3,12, 20(S)-trihydroxydammar-24-ol (2), 3-O-[-d-gluco- pyranosyl-(1,2)--d-glucopyranosyl]-3, 12, 20(S)-trihydroxydammar-24-ene (3), and 3-O--d-glucopyranosyl-3, 12, 20(S)-trihydroxydammar-24-ene (4), identified by TOF-MS, ¹H- and ¹³C-NMR spectral data. Metabolites 1, 3 and 4 were from the incubation with R. stolonifer, and 1 and 2 from the incubation with C. lunata. Compound 2 was identified as a new compound.     

Molecular studies of the 5S RNA genes of Drosophila melanogaster. I. Production of chromosomes containing duplications and deficiencies

Summary Aneuploids have been produced for the 5S RNA genes of Drosophila melanogaster by three different methods. Tandem duplications were produced at a frequency of about 0.05% by irradiation of oocytes and selection for anti-Minutes. Other Minute suppressors were recovered and localized to the region between nu ^Dand Pu ²on the genetic map. One, Su ^M1, reduced crossing-over as a heterozygote and is tentatively identified as a duplication. Deficiencies for the 5S RNA genes were also produced, which showed no dominant phenotype. A Minute deficiency was localized to region 56F-57A on the salivary map and was shown to be allelic to M(2) 173. The anti-Minute method of selection should be useful for producing tandem duplications throughout the genome.Postdoctoral investigator supported by subcontract No. 3322 from the Biology Division of Oak Ridge National Laboratory to the University of Tennessee.Operated by Union Carbide Corporation for the U.S. Atomic Energy Commission.     

Gene Conversion and Functional Divergence in the β-Globin Gene Family

Different models of gene family evolution have been proposed to explain the mechanism whereby gene copies created by gene duplications are maintained and diverge in function. Ohta proposed a model which predicts a burst of nonsynonymous substitutions following gene duplication and the preservation of duplicates through positive selection. An alternative model, the duplication–degeneration–complementation (DDC) model, does not explicitly require the action of positive Darwinian selection for the maintenance of duplicated gene copies, although purifying selection is assumed to continue to act on both copies. A potential outcome of the DDC model is heterogeneity in purifying selection among the gene copies, due to partitioning of subfunctions which complement each other. By using the d_N/d_S () rate ratio to measure selection pressure, we can distinguish between these two very different evolutionary scenarios. In this study we investigated these scenarios in the -globin family of genes, a textbook example of evolution by gene duplication. We assembled a comprehensive dataset of 72 vertebrate -globin sequences. The estimated phylogeny suggested multiple gene duplication and gene conversion events. By using different programs to detect recombination, we confirmed several cases of gene conversion and detected two new cases. We tested evolutionary scenarios derived from Ohtas model and the DDC model by examining selective pressures along lineages in a phylogeny of -globin genes in eutherian mammals. We did not find significant evidence for an increase in the ratio following major duplication events in this family. However, one exception to this pattern was the duplication of -globin in simian primates, after which a few sites were identified to be under positive selection. Overall, our results suggest that following gene duplications, paralogous copies of -globin genes evolved under a nonepisodic process of functional divergence.[Reviewing Editor: Martin Kreitman]     

Evidence for the deficiency of β-glucosidase-activating factor in fibroblasts of patients with I-cell disease

Summary Reduced activity of -glucosidase was shown in the cultured skin fibroblasts of four patients with I-cell disease when the enzyme was tested without the use of detergents. In the presence of taurocholate and triton X100 -glucosidase activity was normal. This suggested a deficiency of a -glucosidase-activating factor in I-cell fibroblasts rather than of the enzyme itself. The deficiency of -glucosidase activity was corrected to some extent by mixing cell lysates, and more effectively by cocultivation and fusion of I-cell disease and Gaucher fibroblasts. These results present evidence for the presence of a -glucosidase-activating factor in normal and Gaucher fibroblasts. In fibroblasts of patients with I-cell disease this activator is probably deficient, as is the case for most lysosomal enzymes.     

Integration of heterologous genes into the chromosome of Saccharomyces cerevisiae using a delta sequence of yeast retrotransposon Ty

Summary Distribution of a delta () sequence of the Ty element on a chromosome of the yeast Saccharomyces cerevisiae was analysed by pulsed-field gel electrophoresis. More than 100 copies of the sequence were non-randomly distributed on the chromosome. Using the sequence as a recombination site, mouse -amylase and human -endorphin genes were introduced into the chromosomal DNA. The integration occurred on a particular chromosome in each case and the copy number was estimated as three to five. It was also found that single- or multi-copy integration occured at a single or multiple sites on the particular chromosome. The integrants secreted -amylase and -endorphin by three-to fivefold compared with single-copy integrants. This type of integration was mitotically stable over a period of 50 generations under non-selective conditions. Offprint requests to: A. Sakai     

Adrenergic influences on the permeability of rabbit submandibular salivary glands to blood-borne horseradish peroxidase

Synopsis Horseradish peroxidase (HRP) administered close-arterially, has been found to enter rabbit submandibular saliva elicited by parasympathetic nerve stimulation. Adrenalin, superimposed on parasympathetic nerve stimulation, increased the passage of HRP into the saliva. Use of - and -adrenoceptor agonists, either separately or together, and use of - or -adrenoceptor antagonists together with adrenalin indicate that both - and -receptor stimulation is necessary for this increase in glandular permeability to occur. Histochemical assessment showed that HRP had permeated the interstitial spaces of the gland and entered the spaces between adjacent parenchymal cells. However, in unstimulated glands it had only reached the lumina of striated ducts, but after adrenalin administration, peroxidase was also observed within acinar lumina. This work indicates that the predominant pathway taken by the HRP was via intercellular spaces and it is suggested that the permeability between junctional complexes of parenchymal cells is capable of being modifiedin vivo.     

Monster zooids inCryptosula pallasiana (bryozoa,cheilostomata ascophora)

In the ascophoran bryozoanCryptosula pallasiana (Moll) many examples of regenerative growth and of abnormal zooids have been observed. Most of the different forms of monsters are caused by fusion of zooids or zooid buds respectively. Zooids may often fuse laterally but in a few cases also distally. Some monster zooids have two polypides and combined opercula caused by various degrees of fusion of the apertures. Lack of sufficient space may lead to the formation of dwarf zooids or remainder cystids.     

A Maximum Likelihood Method for Detecting Functional Divergence at Individual Codon Sites, with Application to Gene Family Evolution

The tailoring of existing genetic systems to new uses is called genetic co-option. Mechanisms of genetic co-option have been difficult to study because of difficulties in identifying functionally important changes. One way to study genetic co-option in protein-coding genes is to identify those amino acid sites that have experienced changes in selective pressure following a genetic co-option event. In this paper we present a maximum likelihood method useful for measuring divergent selective pressures and identifying the amino acid sites affected by divergent selection. The method is based on a codon model of evolution and uses the nonsynonymous-to-synonymous rate ratio () as a measure of selection on the protein, with =1, <1, and >1 indicating neutral evolution, purifying selection, and positive selection, respectively. The model allows variation in among sites, with a fraction of sites evolving under divergent selective pressures. Divergent selection is indicated by different s between clades, such as between paralogous clades of a gene family. We applied the codon model to duplication followed by functional divergence of (i) the and globin genes and (ii) the eosinophil cationic protein (ECP) and eosinophil-derived neurotoxin (EDN) genes. In both cases likelihood ratio tests suggested the presence of sites evolving under divergent selective pressures. Results of the and globin analysis suggested that divergent selective pressures might be a consequence of a weakened relationship between fetal hemoglobin and 2,3-diphosphoglycerate. We suggest that empirical Bayesian identification of sites evolving under divergent selective pressures, combined with structural and functional information, can provide a valuable framework for identifying and studying mechanisms of genetic co-option. Limitations of the new method are discussed.     

Methodological problems in evolutionary biology III. Selection and levels of organization

Apparently factual disagreement on the level(s) at which selection operates often results from different interpretations of the term selection. Attempts to resolve terminological problems must come to grips with a dilemma: a narrow interpretation of selection may lead to a restricted view on evolution; a broader, less precise, definition may wrongly suggest that selection is the centre of a unified, integrated theory of evolution. Different concepts of selection, therefore, should carefully be kept apart.     

Socio-Ecological Webs and Sites of Sociality:Levins' Strategy of Model Building Revisited

This essay extends Levins' 1966 analysis of modelbuilding in ecology and evolutionary biology. Amodel, as the product of modeling, might bevalued according to its correspondence to reality. Yet Levins' emphasis on provisionality and changeredirects attention to the processes ofmodeling, through which scientists select and generatetheir problems, define their categories, collect theirdata, compare competing models, and present theirfindings. I identify several points where decisionsare required that are not determined by nature. Thisinvites examination of the social considerationsmodelers are reacting to at the sites of sociality.Modelers must weave socio-ecological webs so thatthe models can be seen to represent their subjectmatter at the same time as the modelers secure thesupport of colleagues, collaborators and institutions,and enjoin others to act upon their conclusions. Notonly do theory justification and theory generationmerge, but the joint project becomes simultaneouslyphilosophical and sociological.     

Biochemical characterization of the β-chain variant haptoglobin Marburg

Summary -chains were isolated from two individuals heterozygous for the -chain mutant haptoglobin Marburg. Total amino acid composition and tryptic peptides were compared with -chains from common haptoglobin types. ^Mb chain preparations are characterized by the presence of -chains with an atypical electrophoretic migration rate and by at least three, possibly four additional peptides in their tryptic digests. It is probable that haptoglobin Marburg is the result of an mutational event other than a single base substitution.Supported by US-PHS grant AM 11796 and by US-PHS grant HD 03321 and aided by a grant from the Deutsche Forschungsgemeinschaft.     

Cytochemical investigations on the localization of 5′-nucleotidase in the rat hippocampus with special reference to synaptic regions

Summary The fine structural localization of the 5-nucleotidase was investigated in the CA₃ region of the rat hippocampus. The attention was focussed on the occurrence of the enzyme in the synaptic region. The 5-nucleotidase activity was demonstrated at the surface membranes of axons and dendrites. Prominent portions of enzyme activity were detectable in the nuclei and the nuclear envelope, whereas the cytoplasmic organelles were nearly devoid of reaction product. In synapses five types of 5-nucleotidase localization were revealed. A participation of the enzyme in the process of neurotransmission is discussed.The research reported in this paper was supported by the Ministerium für Wissenschaft und Technik der DDR     

Cell wall regeneration by protoplasts of Chlamydomonas

Protoplasts from Chlamydomonas smithii prepared by the action of C. reinhardii gamete autolysine have been studied with respect to cell wall regeneration. Natural protoplasts within sporangia were also investigated for purposes of comparison. In both cases a new cell wall is completed within 2–3 h of the onset of regeneration. The first visible stages of wall regeneration are to be seen after 40–60 min as a fine fringe outside of the plasmalemma. The development of the typical central triplet follows within the next 1 h. Cell wall regeneration is reversibly inhibited by cycloheximide (10g ml^-1) and reversibly disturbed by concanavalin A (50 g ml^-1). Actinomycin D at concentration over 100g ml^-1 also inhibit but the inhibition is irreversible and peculiar membrane effects are observed. Chelators (ethylenediamine tetraacetic acid; ethyleneglycol-bis-aminoethyl ether) and 2-deoxyglucose slightly retard or have no effect on cell wall regeneration.Abbreviations EDTA ethylenediamine tetraacetic acid - EGTA ethyleneglycol-bis(aminoethyl ether) - N,N tetraacetic acid     

Plant regeneration via somatic embryogenesis in Styrian pumpkin: cytological and biochemical investigations

Somatic embryo formation was induced from cotyledon explants of Styrian pumpkin (Cucurbita pepo L. subsp. pepo var. styriaca Greb.) by using a solid MS medium supplemented with 16.11M NAA and 4.44M BA or 26.85M NAA and 13.32M BA. The callus proliferation was more efficient on medium supplemented with 26.85M NAA and 13.32M BA. In contrast, the embryogenic response was higher on medium with lower concentrations of growth regulators (16.11M NAA and 4.44M BA). The time needed for embryo induction did not depend on medium composition. Embryos in globular stage were transferred to three different maturation media, containing 2.89M GA₃ in combination with 0.54M NAA, 11.42M IAA and growth regulator-free medium. The germination rate was the highest when embryos were cultured on medium with 11.42M IAA. Plantlets grown on this medium achieved maturity suitable for transplantation into soil within 9 to 10weeks. The regenerated plants were successfully transferred into field and developed fertile flowers and set fruits. Biochemical analysis showed significant lower total glutathione levels among in vitro grown plantlets compared to seedlings grown in soil. When the plantlets were transferred into soil, they reached a normal size within a month and the glutathione concentration was comparable to seed-derived plants at the same developmental stage. Transmission electron microscopy was used to investigate possible differences in the ultrastructure of cells from callus cultures, and leaf cells of regenerated and seed-derived plants. Differences in the ultrastructure were found within chloroplasts which contained only single thylakoids, large starch grains and small plastoglobuli in callus cells in comparison to leaf cells, which possessed a well developed thylakoid system, small starch grains and large plastoglobuli.     

The “averaging fallacy” and the levels of selection

This paper compares two well-known arguments in the units of selection literature, one due to , the other due to . Both arguments concern the legitimacy of averaging fitness values across contexts and making inferences about the level of selection on that basis. The first three sections of the paper shows that the two arguments are incompatible if taken at face value, their apparent similarity notwithstanding. If we accept Sober and Lewontin's criterion for when averaging genic fitnesses across diploid genotypes is illegitmate, we cannot accept Sober and Wilson's criterion for when averaging individual fitnesses across groups is illegitimate, and vice versa. The final section suggests a possible way of reconciling the two arguments, by invoking an ambiguity in the concept of genic selection.     

Study of RecA-Independent Homologous Recombination and a Chromosomal Rearrangement in the Escherichia coli Strain Carrying an Extended Heterozygous Tandem Duplication

A heterozygous tandem duplication in the Escherichia coli deo operon region deoAdeoB::Tn5/ deoCdeoDthr::Tn9 with the total length approximately 150 kb, which was obtained in the conjugational mating in the HfrH strain, was examined. By means of digestion with the NotI enzyme, pulsed-field gel electrophoresis, and the conjugational transfer of the duplication in the F^– strain, the chromosomal rearrangement, which occurred in the duplication region upon its stabilization in the bacterial genome, was studied. In a more stable strain, two new NotI sites were shown to appear in the chromosomal region located close to the duplication, which might have resulted from the transposition of the IS50 sequence from Tn5. The data were also obtained indicating the possibility of secondary transposition of the chromosomal segment between the two new NotI sites (approximately 30 kb) in the region located near the duplication. With the use of rec ⁺ and recA strains, two types of haploid and diploid segregants generated by the duplication were studied: DeoD⁺ (the deoD⁺ allele is not expressed in the original duplication due to the polar effect of the deoB::Tn5 insertion) and DeoC DeoD. The segregation of DeoD⁺ clones was shown to be RecA-dependent, whereas the DeoC DeoD segregants selected on the medium that contained thymine at a low concentration (i.e., under conditions of thymine starvation) appeared at a rather high frequency. However, the relative frequency of haploid clones, which have lost the duplication, strongly decreased in the recA genome among segregants of both types.     

An ecological approach to biosystem thermodynamics

The general attributes of ecosystems are examined and a naturally occurring reference ecosystem is established, comparable with the isolated system of classical thermodynamics. Such an autonomous system with a stable, periodic input of energy is shown to assume certain structural characteristics that have an identifiable thermodynamic basis. Individual species tend to assume a state of least dissipation; this is most clearly evident in the dominant species (the species with the best integration of energy acquisition and conservation). It is concluded that ecosystem structure results from the antagonistic interaction of two nearly equal forces. These forces have their origin in the Principle of Most Action (least dissipation or least entropy production) and the universal Principle of Least Action. Most action is contingent on the equipartitioning of the energy available, through uniform interaction of similar individuals. The trend to Least action is contingent on increased dissipation attained through increasing diversity and increasing complexity. These principles exhibit a basic asymmetry. Given the operation of these opposing principles over evolutionary time, it is argued that ecosystems originated in the vicinity of thermodynamic equilibrium through the resonant amplification of reversible fluctuations. On account of the basic asymmetry the system was able to evolve away from thermodynamic equilibrium provided that it remained within the vicinity of ergodynamic equilibrium (equilibrium maintained by internal work, where the opposing forces are equal and opposite).At the highest level of generalization there appear to be three principles operating: i) maximum association of free-energy and materials; ii) energy conservation (deceleration of the energy flow) through symmetric interaction and increased homogeneity; and iii) the principle of least action which induces acceleration of the energy flow through asymmetrical interaction. The opposition and asymmetry of the two forces give rise to natural selection and evolution.     

Somatic hybrid plants produced by electrofusion between dihaploid potatoes: BF15 (H1), Aminca (H6) and Cardinal (H3)

In order to regenerate somatic hybrids, mesophyll protoplasts from a dihaploid potato, BF15 (H1), were electrofused with those from two other dihaploid clones, Aminca (H6) and Cardinal (H3). Determination of the ploidy level by flow cytometry showed that 10% of plants regenerated from the fusion experiment with BF15 + Aminca were diploids, 14% triploids, 63% tetraploids and very few were mixoploids or had a higher ploidy level. Using morphological markers and vigour in plant growth, we were able to recover a total of 24 somatic hybrid plants, respectively 20 and 4 hybrids (accounting for 12% and 13% of regenerants) from the fusions BF15 + Aminca and BF15 + Cardinal. Most of the somatic hybrids were at the expected tetraploid level (2n=4x=48). The hybrid nature was confirmed by examining isoenzyme patterns for malate dehydrogenase (MDH) and isocitrate dehydrogenase (ICD).     

Multiple forms of inducible drug-metabolizing enzymes: A reasonable mechanism by which any organism can cope with adversity

Summary All organisms possess a number of genetically regulated mechanisms in order to cope with rapid adverse changes in the environment. The two systems which appear to respond to a seemingly endless array of chemical specificities are the immune response and the induction of drug-metabolizing enzymes. Similarities and differences between the immunoglobulin and the cytochrome P-450-mediated monooxygenase systems are described. DNA insertion sequences, plasmid transposons, maize controlling elements, gene duplication, intervening sequences, and high-frequency intergenic recombination are all discussed as possible methods by which organisms can adapt quickly to a new selective pressure. If the regulation of P-450 induction resembles in any way the other methods by which pro- and eukaryotes cope genetically with numerous forms of environmental adversity, therefore, it is very likely that mammalian tissues contain hundreds, if not thousands, of inducible forms of P-450.Portions of this overview were presented at the Symposium on The Effects of Drugs on Enzyme Induction Related to the Metabolism of Drugs and Carcinogenic Compounds, 14th Symposium Medicum Hoechst, County Mayo, Ireland, May, 1978 (1), and the Symposium on Isolated Drug-Metabolizing Enzymes, Mainz, Germany, July, 1978 (2). The author is Chief of the Developmental Pharmacology Branch, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20205.     

Mitotic cycle time and somatic selection after neutron irradiation of germinating tomato seeds

Mitotic cycle time was estimated by means of a natural method in control and irradiated material of a dwarf tomato line in order to express the intensity of somatic selection in terms of this biological factor. In the natural method the mitotic index is estimated in relation to the number of hours after germination. The time interval between successive peaks in the mitotic index is considered to be an indication of mitotic cycle time. In control material mitotic cycle time was also estimated by the continuous colchicine method and compared with the natural method.The investigations reported here were performed at the Genetics Department, Catholic University, Nijmegen, Director Prof. Dr. S. J. Geerts and the Institute for the Application of Atomic Energy in Agriculture, Wageningen, Director Dr. D. de Zeeuw.