Structure and variability of nuclear ribosomal genes in the genus Helianthus

Summary The restriction map of the rDNA unit of Helianthus annuus was constructed using EcoRI, BamHI, HindIII, KpnI and SacI restriction enzymes. Variations in this map among 61 ecotypes representing 39 species of the genus Helianthus were analyzed. The sizes of the rDNA unit ranged from 9.8 to 11.0 kbp, due to a length-repeat heterogeneity of the external non-transcribed spacer by increments of 200 base pair segments. Lengthrepeat heterogeneity and restriction polymorphism were found to be characteristic of populations or species of Helianthus. Restriction patterns and thermal melting with probes of a cloned H. annuus ENTS segment allowed us to differentiate species from each other. However, most lines of the cultivated sunflower were found to be identical on the basis of the physical properties of their ribosomal DNA.     

Secondary structure of ShK toxin, a potassium-channel-blocking peptide

Summary Sea anemones possess small K-channel-blocking peptides about the same size as the scorpion K-channel toxins. We have estimated the secondary structure content (33% helix, 26% -sheet) of one of these toxins, ShK toxin, using CD, Raman, and FTIR spectroscopy. A hypothetical 3D structure of the peptide core has been constructed using secondary structure and disulfide-linkage constraints; a single helical segment running from Ala¹⁴ through Leu²⁵ is predicted.     

The role of recruitment in structuring patterns of small-scale spatial variability in intertidal and subtidal algal turfs

Recruitment is often important in structuring patterns of distribution and abundance of algal assemblages. Intertidal and subtidal turfing algal assemblages consistently vary on small spatial scales (tens of centimetres), and this variability may be due to patterns of recruitment varying on similar spatial scales. The validity of this model was evaluated by testing the hypothesis that the numbers and types of taxa recruiting to turfs would vary at small spatial scales within intertidal and within subtidal habitats. Abundances of algal recruits were estimated on sandstone plates that were placed at a number of spatial scales within intertidal and within subtidal habitats (centimetres to tens of metres). Significant differences in entire assemblages were found only between habitats. This was explained by abundances of individual taxa, which generally varied between intertidal and subtidal habitats or between sites within habitats. Only small proportions of the overall spatial variation (dissimilarity) could be explained at the scale of replicate recruitment plates that were centimetres apart. Results indicate that while recruitment may contribute to differences between intertidal and subtidal habitats, it cannot explain the small-scale spatial variability in established turfing algal assemblages within these habitats. There was some evidence to suggest that recruitment may contribute to variability in established turfing algal assemblages but only over longer time scales than examined here.     

Secondary structure of pre-rRNA 41S spacers and its possible role in determining the processing sites

The mechanism of pre-rRNA processing is still unknown. In this paper we discuss a possible role of the secondary structure of the internal transcribed spacer region (ITS-1 and ITS-2) in processing of 41S pre-rRNA. Potential possibilities for double strand structure formation in ITS-1 and ITS-2 and its rearrangement in the course of processing of 41S pre-rRNA with involvement of U3 RNA were analyzed for a number of species. We have concluded that this rearrangement is the necessary stage in 41S pre-rRNA processing when every functionally designated area (for mature 18S, 28S and 5,8S rRNAs, for ITS-1, ITS-2 and 21S pre-rRNA) has to be organized into an individual domain. In these strongly structured domains 5'- and 3'-ends are spatially brought together whereas processing sites are localized between these compactised areas.     

DES-ASP1--angiotensin II: possible role in mediating responses of the renin-angiotensin system.

In dexamethasone-treated dogs, both antiogensin II and its heptapeptide fragment (des-asp1-angiotensin II) stimulated the secretion of the adrenal steroids aldosterone, corticosterone, and cortisol. Further, there was no statistically significant difference in the steroidogenic potency of the two peptides. An interesting incidental finding was the decrease in PRA with both peptides, again with no demonstrable difference in the magnitude of the responses. The present data are consistent with the hypothesis that the heptapeptide mediates responses produced by the renin-angiotensin system in both the adrenals and the kidneys.     

Polarity of microtubular structures in manchette-like formations: possible role of the "11th filament".

Bundles of microtubular structures appear in the cytoplasm of spermatids of the African frog Dicroglossus occipitalis. They are observed in the vicinity of axonemes. Natural tubulin polymerization leads to the formation of hooks on microtubular structures. They can be related to experimentally induced tubulin hooks. The direction of curvature of the hooks allows us to define the polarity of the bundles. This is opposite to the polarity of axonemal microtubules: Bundles and axonemes are antiparallel. Under colchicine action, arch-like microtubular structures are shown to open in the same direction as they lock. This enables us to characterize their opening and locking site: It corresponds to the place of the "11th filament" described in microtubular structures such axonemes. The "11th filament" is thus demonstrated to be the most susceptible to natural opening and to the action of colchicine in microtubular structures.     

Secondary structure and orientation of the pore-forming toxin lysenin in a sphingomyelin-containing membrane

Lysenin is a sphingomyelin-recognizing toxin which forms stable oligomers upon membrane binding and causes cell lysis. To get insight into the mechanism of the transition of lysenin from a soluble to a membrane-bound form, surface activity of the protein and its binding to lipid membranes were studied using tensiometric measurements, Fourier-transform infrared spectroscopy (FTIR) and FTIR-linear dichroism. The results showed cooperative adsorption of recombinant lysenin-His at the argon-water interface from the water subphase which suggested self-association of lysenin-His in solution. An assembly of premature oligomers by lysenin-His in solution was confirmed by blue native gel electrophoresis. When a monolayer composed of sphingomyelin and cholesterol was present at the interface, the rate of insertion of lysenin-His into the monolayer was considerably enhanced. Analysis of FTIR spectra of soluble lysenin-His demonstrated that the protein contained 27% β-sheet, 28% aggregated β-strands, 10% α-helix, 23% turns and loops and 12% different kinds of aggregated forms. In membrane-bound lysenin-His the total content of α-helices, turns and loops, and β-structures did not change, however, the 1636cm⁻¹ β-sheet band increased from 18% to 31% at the expense of the 1680cm⁻¹ β-sheet structure. Spectral analysis of the amide I band showed that the α-helical component was oriented with at 41° to the normal to the membrane, indicating that this protein segment could be anchored in the hydrophobic core of the membrane.     

Secondary cell-wall assembly in flax phloem fibres: role of galactans

Non-lignified fibre cells (named gelatinous fibres) are present in tension wood and the stems of fibre crops (such as flax and hemp). These cells develop a very thick S2 layer within the secondary cell wall, which is characterised by (1) cellulose microfibrils largely parallel to the longitudinal axis of the cell, and (2) a high proportion of galactose-containing polymers among the non-cellulosic polysaccharides. In this review, we focus on the role of these polymers in the assembly of gelatinous fibres of flax. At the different stages of fibre development, we analyse in detail data based on sugar composition, linkages of pectic polymers, and immunolocalisation of the β-(1→4)-galactans. These data indicate that high molecular-mass gelatinous galactans accumulate in specialised Golgi-derived vesicles during fibre cell-wall thickening. They consist of RG-I-like polymers with side chains of β-(1→4)-linked galactose. Most of them are short, but there are also long chains containing up to 28 galactosyl residues. At fibre maturity, two types of cross-linked galactans are identified, a C–L structure that resembles the part of soluble galactan with long side chains and a C–S structure with short chains. Different possibilities for soluble galactan to give rise to C–L and C–S are analysed. In addition, we discuss the prospect for the soluble galactan in preventing the newly formed cellulose chains from completing immediate crystallisation. This leads to a hypothesis that firstly the secretion of soluble galactans plays a role in the axial orientation of cellulose microfibrils, and secondly the remodelling and cross-linking of pectic galactans are linked to the dehydration and the assembly of S2 layer.     

Daily temporal organization of laying in Japanese quail: variability and heritability

In birds, many behavioral and physiological processes that occur during reproduction show daily rhythms in response to environmental temporal constraints. In this study, the individual daily organization of laying and its genetic determinant in Japanese quail (Coturnix coturnix japonica) were analyzed. For this purpose, the oviposition time of 102 randomly chosen females, maintained in long-day photoperiodic conditions (LD 14h:10h) for 1 mon was observed and extreme phenotypes selected. Laying is characterized by two parameters: oviposition interval and laying hour.

The birds showed a specific time of laying during 24 h. All eggs were laid in the afternoon between 6.5 and 14 h after lights on (HALO). Two laying profiles were determined: 20% of females with an oviposition interval greater than 24 h (24.7±0.2 h) (the “delayed” profile) laid progressively later each day until a pause day. The remaining 80% of the females laid at the same time each day, with few pause days and an oviposition interval close to 24 h (24.0±0.2 h) (the “stable” profile). Among the females, showing this last profile, an intra-individual stability and an inter-individual variability of laying hour was established. Two extreme laying phenotypes were then determined: the “early” phenotype (E) for females laying on average between 7.5 and 9.5 HALO and the “late” phenotype (L) for females laying between 12.5 and 14 HALO.

In order to study the genetic basis of the laying hour, three females of each extreme phenotype were selected and crossed with two different males. The E and L females produced 57 F1E and 42 F1L daughters, respectively. F1 females displayed both laying profiles. However, the proportion of females displaying a “delayed” profile was higher in the L line (50%) than in the E line (29.8%). For the “stable” daughters, artificial selection induced an advance in laying hour of 4.7% for the E line and a delay of 4.7% for the L line. Realized heritability was estimated at 0.5. Moreover, the laying hour of the daughters was correlated positively to that of the mothers (N=61;r=0.45). These results support the notion of heritability of oviposition time in Japanese quail.     

Daily temporal organization of laying in Japanese quail: variability and heritability

In birds, many behavioral and physiological processes that occur during reproduction show daily rhythms in response to environmental temporal constraints. In this study, the individual daily organization of laying and its genetic determinant in Japanese quail (Coturnix coturnix japonica) were analyzed. For this purpose, the oviposition time of 102 randomly chosen females, maintained in long-day photoperiodic conditions (LD 14h:10h) for 1 mon was observed and extreme phenotypes selected. Laying is characterized by two parameters: oviposition interval and laying hour.

The birds showed a specific time of laying during 24 h. All eggs were laid in the afternoon between 6.5 and 14 h after lights on (HALO). Two laying profiles were determined: 20% of females with an oviposition interval greater than 24 h (24.7±0.2 h) (the “delayed” profile) laid progressively later each day until a pause day. The remaining 80% of the females laid at the same time each day, with few pause days and an oviposition interval close to 24 h (24.0±0.2 h) (the “stable” profile). Among the females, showing this last profile, an intra-individual stability and an inter-individual variability of laying hour was established. Two extreme laying phenotypes were then determined: the “early” phenotype (E) for females laying on average between 7.5 and 9.5 HALO and the “late” phenotype (L) for females laying between 12.5 and 14 HALO.

In order to study the genetic basis of the laying hour, three females of each extreme phenotype were selected and crossed with two different males. The E and L females produced 57 F1E and 42 F1L daughters, respectively. F1 females displayed both laying profiles. However, the proportion of females displaying a “delayed” profile was higher in the L line (50%) than in the E line (29.8%). For the “stable” daughters, artificial selection induced an advance in laying hour of 4.7% for the E line and a delay of 4.7% for the L line. Realized heritability was estimated at 0.5. Moreover, the laying hour of the daughters was correlated positively to that of the mothers (N=61;r=0.45). These results support the notion of heritability of oviposition time in Japanese quail.     

Estimation of a Secondary Parameter in a Group Sequential Clinical Trial

In this article, we investigate estimation of a secondary parameter in group sequential tests. We study the model in which the secondary parameter is the mean of the normal distribution in a subgroup of the subjects. The bias of the naive secondary parameter estimator is studied. It is shown that the sampling proportions of the subgroup have a crucial effect on the bias: As the sampling proportion of the subgroup at or just before the stopping time increases, the bias of the naive subgroup parameter estimator increases as well. An unbiased estimator for the subgroup parameter and an unbiased estimator for its variance are derived. Using simulations, we compare the mean squared error of the unbiased estimator to that of the naive estimator, and we show that the differences are negligible. As an example, the methods of estimation are applied to an actual group sequential clinical trial, The Beta-Blocker Heart Attack Trial.     

A model for the development of the tandem repeat units in the EBV ori-P region and a discussion of their possible function

Summary This paper presents an analysis of the repeat units of the ori-P region of the Epstein-Barr virus (EBV) genome. These repeat units are well-conserved palindromes. The pattern of these repeats, their lengths, phases, and the distribution of the relatively few substitutions are explained by a scenario that gives a reasonable course for the evolutionary development of the pattern. The scenario suggests a model for the production of an initiating 3/2 palindrome from a moderately lengthy sequence. The palindromic units are then multiplied in judicious combinations by mechanisms of unequal crossing-over events associated with some point substitutions and a few instances of slippage replication. The potential secondary structures of the two separated tandem palindromic repeat regions in ori-P are contrasted. Possible modes of binding of Epstein-Barr nuclear antigen (EBNA) 1 protein to these hairpins are discussed. A number of possibilities for the origin and development of the ori-P region in relation to viral and cellular function are considered.     

Isolation of a putative cell adhesion mediating lectin from teratocarcinoma stem cells and its possible role in differentiation

We have identified a cell surface teratocarcinoma stem cell lectin with a fucan/mannan specificity by the use of an erythrocyte rosetting assay as well as hemagglutination assay. We have also described experiments that suggest that the lectin is involved in mediating divalent cation-independent adhesion of the stem cells. This molecule has been purified from stem cell conditioned medium and identified as a polypeptide of 56 000 apparent molecular weight. An antibody has been raised to this 56K polypeptide (using material eluted from an SDS-polyacrylamide gel as the immunogen) and its specificity determined by protein blot analysis. In addition, we have recently observed that only carbohydrates recognized by the lectin interfere with in vitro embryoid body formation by the stem cells, suggesting that the lectin may be involved in differentiation.     

Bombesin: a possible role in wound repair

During tissue regeneration and wound healing of the skin, migration, proliferation and differentiation of keratinocytes are important processes. Here we assessed the effect of a neuropeptide, bombesin, on keratinocytes during regeneration from scratch wounding. Bombesin purified from amphibian skin, is homologous of mammalian gastrin-releasing peptide and is active in mammals. Its pharmacological effects mediate various physiological activities: hypertensive action, stimulating action on gastric secretion, hyperglycemic effect or increased insulin secretion. In vitro it shows a hyperproliferative effect on different experimental models and is involved in skin repair. The aim of this study was to elucidate the effect of Bombesin in an in vitro experimental model on a mechanically injured human keratinocyte monolayer. We evaluated different mediators involved in wound repair such as IL-8, TGFbeta, IL-1, COX-2, VEGF and Toll-like receptors 2 and 4 (TLR2 and TLR4). We also studied the effects of bombesin on cell proliferation and motility and its direct effect on wound repair by observing the wound closure after mechanical injury. The involvement of the bombesin receptors neuromedin receptor (NMBR) and gastrin-releasing peptide receptor (GRP-R) was also evaluated. Our data suggest that bombesin may have an important role in skin repair by regulating the expression of healing markers. It enhanced the expression of IL-8, TGFbeta, COX-2 and VEGF. It also enhanced the expression of TLR2, while TLR4 was not expressed. Bombesin also increased cell growth and migration. In addition, we showed that NMBR was more involved in our experimental model compared to GRP-R.     

Optimal risky choice in humans: effects of amount of variability

The present study investigated whether the predictions of an optimal risk-sensitive foraging model (the energy-budget rule) would extend to humans’ choices between high- and low-variance monetary response options. In Experiment 1, participants were presented with repeated choices between two options that had the same mean values but different variances across positive- and negative-budget conditions. In Experiment 2, a within-subject comparison was conducted to investigate choice under positive- and negative-budget conditions when the options were either (a) a fixed option and a high-variance option, or (b) a low-variance option and a high-variance option. Session-wide choices were analyzed in relation to the predictions of the energy-budget rule and sequential choices were analyzed with dynamic optimization modeling. When both options were variable choice was generally consistent with predictions of the energy-budget rule and was more risk prone under negative-budget than positive-budget conditions. Sequential choices were sensitive to local budget conditions, but choices were less consistent with optimality when both options were variable, possibly because of the greater similarity in expected earnings for optimal and nonoptimal choices in these conditions. Overall, the results provide further evidence that the energy-budget rule may have broad applicability and that it can extend to human risky choice between multiple variable response options.     

On the possible role of egg mimics in speciation

Sexually active male haplochromine cichlid fishes possess pronounced yellow ovoid spots on the anal fm, which mimic eggs of the female and have therefore been called egg dummies (Wickler, 1962b).It is thought that divergence in egg dummy characteristics can considerably reduce gene flow and in this way may trigger off reproductive isolation. Two ways in which egg dummy divergence can develop are described. Both mechanisms may have been operating, at the same time and in the same area, in different species, or possibly even in a single species.Firstly, egg dummy divergence can develop in species with accurately mimetic egg dummies if selection pressures on egg characteristics (size, shape) differ in temporarily isolated demes. As soon as egg characteristics diverge, egg dummy characteristics follow. This type of egg dummy divergence leads to reproductive isolation by mimicry. Mimetic isolation is proposed as a term to describe the mechanism.Secondly, egg dummy divergence can develop if selection pressures on the egg dummy characteristics are different in two temporarily isolated demes. Upon secondary contact, this divergence may work as a sexual isolation mechanism, since female mate choice is important in haplochromines. Egg dummy divergence is likely to develop rapidly and, if this happens repeatedly, explosive speciation may result. Apparently, explosive speciation has occurred in haplochromines. Neither of the two models has been properly tested. Some preliminary evidence was found that the second model is valid, but so far there is no evidence in favour of the first model.     

Melatonin and its possible role in mediating seasonal metabolic changes of Antarctic krill, Euphausia superba

Melatonin, the chief secretory product of the vertebrate pineal gland is suspected to be a ubiquitous molecule principally involved in the transduction of photoperiodic information. Besides vertebrates, melatonin has been detected throughout phylogeny in numerous non-vertebrate taxa. In the present study, the occurrence of melatonin in Antarctic krill Euphausia superba and its possible role in mediating seasonal metabolic changes was evaluated. Melatonin was quantified by enzyme linked immunosorbent assay (ELISA) in high performance liquid chromatography (HPLC) purified extracts of eyestalks and hemolymph of krill sampled in the Lazarev Sea during the Antarctic winter and summer. In addition, oxygen uptake rates and the activities of the metabolic enzyme malate dehydrogenase (MDH) were recorded to assess the metabolic status of krill. Validation of melatonin measurements was carried out on the basis of three different extraction methods with parallel determination of melatonin by ELISA in crude extracts and in HPLC purified extracts, and after derivatization of melatonin under alkaline conditions in the presence of hydrogen peroxide. A significantly higher respiration rate and MDH activity was found in summer krill than in winter krill indicating that krill was in a state of reduced metabolic activity during winter. However, neither during winter nor during summer there were detectable melatonin concentrations in the visual system or hemolymph of krill. Based on these results, we question a mediating role of melatonin in the control of seasonal metabolic changes in Antarctic krill.     

Mobilization of maternal mRNA in amphibian eggs with special reference to the possible role of membraneous supramolecular structures

Current knowledge of the mechanism of mobilization of maternal mRNA is summarized herein and a working hypothesis has been constructed to explain the mechanism on the assumption that the mRNA enters the cytoplasm in association with the cytoplasmic membraneous structures and is then stored in the structures until liberation and relocation at the step of oocyte maturation. An extensive turnover of poly(A) sequences as well as the occurrence of repetitive sequences in the maternal mRNA may be relevant to mRNA activation.