Purification and biochemical characterization of recombinant human H-ferritins from <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>

Recombinant human H-ferritins produced from Saccharomyces cerevisiae were purified and their molecular properties were characterized. Electrophoresis of the recombinant H-ferritins showed revealed bands with mobilities similar to those of the H-ferritins produced by Escherichia coli. The pI of H-ferritins from yeast was more basic than that of H-ferritins produced by E. coli. When the cells were treated with tunicamycin, the pI of H-ferritins was driven to a sharp band with the pI range similar to that of those produced by E. coli, implying that the H-ferritins from yeast are glycosylated. The iron uptake of H-ferritins was rapid in the initial stage, with a slight reduction when compared to ferritins from E. coli. Recombinant ferritins are commonly used during synthesis of inorganic nanoparticles in their cores for chemical and industrial purposes. Transmission electron microscopy revealed that the reconstitution yield and size distribution of the core minerals were affected depending on the protein shells. The H-ferritins with higher reconstitution yields (64.4%) showed slightly larger sizes (mean 6.52 nm) with narrower size distribution.     

<Emphasis Type="Italic">Acinetobacter</Emphasis> lipases: molecular biology,biochemical properties and biotechnological potential

Lipases (EC 3.1.1.3) have received increased attention recently, evidenced by the increasing amount of information about lipases in the current literature. The renewed interest in this enzyme class is due primarily to investigations of their role in pathogenesis and their increasing use in biotechnological applications [38]. Also, many microbial lipases are available as commercial products, the majority of which are used in detergents, cosmetic production, food flavoring, and organic synthesis. Lipases are valued biocatalysts because they act under mild conditions, are highly stable in organic solvents, show broad substrate specificity, and usually show high regio- and/or stereo-selectivity in catalysis. A number of lipolytic strains of Acinetobacter have been isolated from a variety of sources and their lipases possess many biochemical properties similar to those that have been developed for biotechnological applications. This review discusses the biology of lipase expression in Acinetobacter, with emphasis on those aspects relevant to potential biotechnology applications.     

Housekeeping genes essential for pantothenate biosynthesis are plasmid-encoded in <Emphasis Type="Italic">Rhizobium etli</Emphasis> and <Emphasis Type="Italic">Rhizobium leguminosarum</Emphasis>

Background  

A traditional concept in bacterial genetics states that housekeeping genes, those involved in basic metabolic functions needed for maintenance of the cell, are encoded in the chromosome, whereas genes required for dealing with challenging environmental conditions are located in plasmids. Exceptions to this rule have emerged from genomic sequence data of bacteria with multipartite genomes. The genome sequence of R. etli CFN42 predicts the presence of panC and panB genes clustered together on the 642 kb plasmid p42f and a second copy of panB on plasmid p42e. They encode putative pantothenate biosynthesis enzymes (pantoate-β-alanine ligase and 3-methyl-2-oxobutanoate hydroxymethyltransferase, respectively). Due to their ubiquitous distribution and relevance in the central metabolism of the cell, these genes are considered part of the core genome; thus, their occurrence in a plasmid is noteworthy. In this study we investigate the contribution of these genes to pantothenate biosynthesis, examine whether their presence in plasmids is a prevalent characteristic of the Rhizobiales with multipartite genomes, and assess the possibility that the panCB genes may have reached plasmids by horizontal gene transfer.     

Phenotypic and biological properties of two antagonist <Emphasis Type="Italic">Bacillus subtilis</Emphasis> strains

Bacillus subtilis KB-1111 and KB-1122 were studied to illustrate their phenotypic and biological properties. Comparison of KB-1111 with KB-1122 in morphology was carried out by microscopy and agar plate assays. Biological assay of the test strains showed that they may possess different physiological pathways from those of reference strain ATCC6501. The assessment of antagonism against the indicator fungi showed that both test strains had broad antifungal characteristics against eight phytopathogenic fungi. Of those fungal species, Magnaporthe grisea P131, Sclerotinia sclerotiorum, and F. oxysporium exhibited high sensitivity to the test strains.     

Presence and Diversity of <Emphasis Type="Italic">Streptomyces</Emphasis> in <Emphasis Type="Italic">Dendroctonus</Emphasis> and Sympatric Bark Beetle Galleries Across North America

Recent studies have revealed several examples of intimate associations between insects and Actinobacteria, including the Southern Pine Beetle Dendroctonus frontalis and the Spruce Beetle Dendroctonus rufipennis. Here, we surveyed Streptomyces Actinobacteria co-occurring with 10 species of Dendroctonus bark beetles across the United States, using both phylogenetic and community ecology approaches. From these 10 species, and 19 other scolytine beetles that occur in the same trees, we obtained 154 Streptomyces-like isolates and generated 16S sequences from 134 of those. Confirmed 16S sequences of Streptomyces were binned into 36 distinct strains using a threshold of 0.2% sequence divergence. The 16S rDNA phylogeny of all isolates does not correlate with the distribution of strains among beetle species, localities, or parts of the beetles or their galleries. However, we identified three Streptomyces strains occurring repeatedly on Dendroctonus beetles and in their galleries. Identity of these isolates was corroborated using a house-keeping gene sequence (efTu). These strains are not confined to a certain species of beetle, locality, or part of the beetle or their galleries. However, their role as residents in the woodboring insect niche is supported by the repeated association of their 16S and efTu from across the continent, and also having been reported in studies of other subcortical insects.     

Tubulin polymerization promoting protein (TPPP) ortholog from <Emphasis Type="Italic">Suberites domuncula</Emphasis> and comparative analysis of TPPP/p25 gene family

Sponges are one of the oldest metazoan phyla that are, due to their highly conservative nature, often referred to as the living fossils of multicellular animals. As such, they are a very important model for evolutionary, developmental and functional studies of Metazoa. Tubulin polymerization promoting proteins (TPPPs) are defined by the presence of p25-alpha domain (Pfam05517). Their functional characteristics resemble those of microtubule-associated proteins. Presence of TPPP homologous genes has been postulated in all eukaryotes with ciliated cells and their primary function has been proposed as some basic cilia-connected function. We present here the genomic structure and the corresponding cDNA sequence of one poriferan TPPP homolog (SdTPPP) isolated from the marine sponge Suberites domuncula; and a comparative analysis of TPPP homolog sequences and genomic structures from other Eukaryotes. Our results confirm the radiation of one TPPP homolog into three distinct genes in the Vertebrate lineage, but the origin of different sequences and their phylogenetic relationships show to be influenced by alternative protein isoforms, independent gene duplications, modularity of the p25-alpha domain and possible adaptational requirements to environmental conditions.     

Identification and Functional Analysis of Phosphoproteins Regulated by Auxin in <Emphasis Type="Italic">Arabidopsis</Emphasis> Roots

The phytohormone auxin modulates diverse aspects of plant growth and development. Protein phosphorylation is believed to play a key role in regulating auxin-mediated responses. To determine the phosphoproteins affected by auxin in Arabidopsis, we used phospho-specific antibodies to analyze their profiles on two-dimensional gels, then identified them by mass spectrometry. We found two phosphoproteins, enolase and the beta subunit of succinyl-CoA synthetase (SCS-beta), and noted that their phosphorylation was increased by auxin. To investigate their importance in auxin-mediated processes, we characterized Arabidopsis knockout mutants of the two genes. A homozygous null mutation in the gene for SCS-beta conferred embryo lethality. The enolase knockout mutants showed defects in root development similar to those of auxin-related mutants such as alf3 and xbat32. Therefore, we suggest that enolase is involved in auxin-regulated processes.     

Dopamine for “Wanting” and Opioids for “Liking”: A Comparison of Obese Adults With and Without Binge Eating

Obesity research suffers from an overinclusion paradigm whereby all participants with a BMI beyond a certain cutoff value (e.g., 30) are typically combined in a single group and compared to those of normal weight. There has been little attempt to identify meaningful subgroups defined by their salient biobehavioral differences. In order to address this limitation, we examined genetic and psychological indicators of hedonic eating in obese adults with (n = 66) and without (n = 70) binge eating disorder (BED). Our analyses focused on dopamine (DA) and opioid genetic markers because of their conjoint association with the functioning of brain reward mechanisms. We targeted three functional polymorphisms related to the D2 receptor (DRD2) gene, as well as the functional A118G polymorphism of the mu‐opioid receptor (OPRM1) gene. We found that significantly more obese controls had the “loss‐of‐function” A1 allele of Taq1A compared to their BED counterparts, whereas the “gain‐of‐function” G allele of A118G occurred with greater frequency in the BED group. A significant gene–gene combination χ² analysis also indicated that of those participants with the gain‐gain genotype (G⁺ and A1), 80% were in the BED group whereas only 35% with the loss‐loss genotype (G^? and A1⁺) were in this group. Finally, BED subjects had significantly higher scores on a self‐report measure of hedonic eating. Our findings suggest that BED is a biologically based subtype of obesity and that the proneness to binge eating may be influenced by a hyper‐reactivity to the hedonic properties of food—a predisposition that is easily exploited in our current environment with its highly visible and easily accessible surfeit of sweet and fatty foods.     

Comparative ontogenic analysis of <Emphasis Type="Italic">Cyclamen coum</Emphasis> Mill. and <Emphasis Type="Italic">Cyclamen kuznetzovii</Emphasis> Kotov et Czernov

Many issues of ontogeny, morphogenesis, and taxonomy of the Cyclamen genus plants remain poorly studied thus far. Data available on the species native to Crimea and the Caucasus are also contradictory. Here we studied the C. coum populations growing around the Maikop city in the Caucasus and those of C. kuznetzovii in Crimea in order to describe a particular ontogeny of these two taxa and to draw preliminary conclusions concerning their taxonomic location.     

Transfection of Feminizing <Emphasis Type="Italic">Wolbachia</Emphasis> Endosymbionts of the Butterfly, <Emphasis Type="Italic">Eurema hecabe</Emphasis>, into the Cell Culture and Various Immature Stages of the Silkmoth, <Emphasis Type="Italic">Bombyx mori</Emphasis>

Wolbachia are maternally inherited endosymbiotic bacteria of invertebrates that can manipulate the reproductive systems of their arthropod hosts in a variety of ways. To establish a useful model system for investigating the mechanism of Wolbachia-induced host feminization, we conducted the following series of experiments: (1) feminizing Wolbachia of the butterfly, Eurema hecabe, were transferred into cell cultures of the silkmoth, Bombyx mori, and (2) the transfected Wolbachia in cell cultures were inoculated into B. mori at four immature stages. Wolbachia were successfully transfected into the cell cultures and stably maintained for more than 1 year (>30 passages). However, none of the inoculated insects produced mature oocytes that were Wolbachia-positive. This finding was consistent with the fact that Wolbachia was not detected in individuals in subsequent generations. In contrast, Wolbachia were detected at relatively high frequencies (60–80% of individuals) in the somatic tissues of inoculated insects. Real-time quantitative polymerase chain reaction revealed that the Wolbachia densities in the cultured cells were approximately tenfold higher than those in the native host E. hecabe. Among B. mori individuals inoculated at various developmental stages, those inoculated at early stages exhibited higher Wolbachia densities at the adult stage. The Wolbachia densities in individuals inoculated at the second-instar stage were comparable to those in intact E. hecabe. These results suggest that infection and/or proliferation of Wolbachia in germline cells are actively hindered by regulation in B. mori but feasible in somatic cells and that the Wolbachia densities in somatic tissues are regulated by the living host insects.     

Relationship between reproductive resource allocation and resource capacity in the matriphagous spider, <Emphasis Type="Italic">Chiracanthium japonicum </Emphasis>(Araneae: Clubionidae)

Females of the Japanese foliage spider, Chiracanthium japonicum, are eaten by their offspring at the end of the maternal care period. To examine the patterns of allocation of maternal investment to their offspring associated with female resource capacity, the amounts of female body reserves accumulated before oviposition, reproductive components at the egg-production phase and those at the matriphagy phase were measured using an artificial breeding nest. Regardless of size, female bodies were completely consumed by the offspring, and larger females, i.e. those having larger reserves, produced a larger number of offspring, but not larger offspring. Furthermore, the proportion of reserves allocated to egg production was not affected by the total amount of the reserves, which indicated that the females systematically divided the resources for individual offspring between egg yolk and food for the growth and survival of the offspring. These results suggest that C. japonicum females adjust egg production to their own resource capacity so that they can achieve an investment per individual offspring which is not dependent on resource capacity. Electronic Publication     

Evolution of seed dispersal in North American <Emphasis Type="Italic">Ephedra</Emphasis>

Related plants often produce seeds that are dispersed in very different ways, raising questions of how and why plants undergo adaptive shifts in key aspects of their reproductive ecology. Here we analyze the evolution of seed dispersal syndromes in an ancient group of plants. Ephedra (Gymnospermae; Gnetales; Ephedraceae) is a genus containing ≈50 species in semiarid ecosystems worldwide and with three distinct types of cones. We collected mature cones and seeds of ten species of Ephedra in southwestern United States and measured nine morphological traits for each species. Principal component analysis and other data characterized three types of Ephedra cones and seeds. Species with dry, winged cone bracts are dispersed by wind (i.e., E. torreyana and E. trifurca), those with succulent, brightly-colored cone bracts are dispersed by frugivorous birds (i.e., E. antisyphilitica), and those with small, dry cone bracts and large seeds are dispersed by seed-caching rodents (e.g., E. viridis and E. californica). Two species (E. funerea and E. nevadensis) have cone and seed morphologies intermediate between two seed dispersal syndromes. Seed and cones traits were mapped onto two recent phylogenies to help reveal the evolutionary history of seed dispersal syndromes. Bird dispersal is thought to be the ancestral form of seed dispersal in ephedras as it is common in the Old World where Ephedra originated, but the three North American species dispersed by birds are not monophyletic. The two wind dispersed species in North America also do not cluster together, suggesting separate origins. Seed dispersal by seed-caching rodents is common in North America and appears to have evolved several times, but this syndrome is absent form other continents. The evolutionary history of Ephedra in North America suggests that the means of seed dispersal has been malleable. Evolutionary shifts were likely linked to changes in ecological conditions.     

Plasticity in courtship and sneaking behaviors depending on tail length in the male guppy, <Emphasis Type="Italic">Poecilia reticulata</Emphasis>

Male guppies Poecilia reticulata exhibit two types of mating behavior, i.e., courtship displays for cooperative copulation and sneaking attempts for forced copulation. The frequencies of the two male mating behaviors are influenced by tail length. Males possessing long tails exhibit courtship displays less frequently and sneaking attempts more frequently than those possessing short tails, even though they have similar total lengths. To examine whether these male behavioral tendencies depending on tail length are genetically controlled or are determined by tail length per se, tail length manipulation was conducted. The tail lengths of males that had previously possessed longer tails were surgically shortened to a greater degree than those of their counterparts that had previously possessed shorter tails. Although the frequencies of the mating behaviors exhibited by the latter males did not apparently change, the former males clearly increased the frequency of courtship displays and decreased that of sneaking attempts following tail shortening. These results indicate that males adjust the frequencies of the two mating behaviors according to their tail length. Since females avoid cooperative mating with males possessing long tails, the change in mating behavioral patterns by males depending on their tail length may increase their mating opportunities.     

Secondary galling: a novel feeding strategy among ‘non‐pollinating’ fig wasps from Ficus curtipes

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The symptom difference induced by <Emphasis Type="Italic">Tobacco mosaic virus</Emphasis> and <Emphasis Type="Italic">Tomato mosaic virus</Emphasis> in tobacco plants containing the <Emphasis Type="Italic">N</Emphasis> gene is determined by movement protein gene

Tobacco mosaic virus (TMV) and Tomato mosaic virus (ToMV) are two closely related viruses in the genus Tobamovirus, but they induce obviously different sizes of necrotic lesions in tobacco plants containing the N gene. Comparison of the symptoms produced by TMV, ToMV and a chimaeric virus (T/OMP), in which the TMV movement protein (MP) gene was replaced by the ToMV MP gene, showed T/OMP caused necrotic lesions that were similar in size to those of ToMV in tobacco plants containing the N gene. The coat protein and MP of the three viruses accumulated in planta with similar levels, and the replication level of TMV and T/OMP in protoplasts also had no difference. Comparison of the activities of defense-related enzymes (PAL, POD and PPO) induced by the three viruses also showed that the variability of enzyme activity induced by T/OMP was similar to that induced by TMV, but different from that induced by ToMV. The results indicate that the size difference of necrotic lesions induced by TMV and ToMV in tobacco plants containing the N gene results from the functional difference of their MP genes.     

Male–female associations and female olfactory neurogenesis with pair bonding in Mus spicilegus

Mound‐building mice Mus spicilegus exhibit life‐history traits that are unique among the Mus species complex, such as the cooperative mound‐building behaviour that gives the species its common name. In this and other socially coordinated activities, such as those associated with reproduction, these mice should be able to recognize individuals (via discrimination based on kinship, population and species) to mediate their interactions. Our previous studies have provided evidence of population and species recognition in M. spicilegus. The aims of the present study were: (i) to study associations of mice during their reproductive period (in outdoor enclosures), (ii) to investigate whether there is an influence of relatedness of females in these associations, (iii) to determine whether female M. spicilegus are able to make kin vs. non‐kin discriminations, and (iv) to study certain neurobiological correlates of male–female bonds. Stable male–female associations were found in almost all the experimental groups, both those with unrelated and unfamiliar females and those with unfamiliar sisters. Kinship between females did not affect female associations in our enclosure experiment, but in our kin discrimination experiment females did distinguish between unfamiliar sisters and unfamiliar unrelated females. Shared kinship may not encourage cooperative rearing of pups but could enhance cooperation in building mounds where mice over‐winter. Male–female associations could be based on a social bond, as hypothesized from previous laboratory experiments. This was substantiated in this study by increased olfactory bulbar neurogenesis in females that preferred (in two choice tests) their sexual partner 3 weeks after their first mating. Based on these results there is clear evidence to suggest that the mating system of M. spicilegus is social monogamy. © 2005 The Linnean Society of London, Biological Journal of the Linnean Society, 2005, 84 , 323–334.     

Role of planktonic and sessile extracellular metabolic byproducts on <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> and <Emphasis Type="Italic">Escherichia coli</Emphasis> intra and interspecies relationships

Bacterial species are found primarily as residents of complex surface-associated communities, known as biofilms. Although these structures prevail in nature, bacteria still exist in planktonic lifestyle and differ from those in morphology, physiology, and metabolism. This study aimed to investigate the influence of physiological states of Pseudomonas aeruginosa and Escherichia coli in cell-to-cell interactions. Filtered supernatants obtained under planktonic and biofilm cultures of each single species were supplemented with tryptic soy broth (TSB) and used as the growth media (conditioned media) to planktonic and sessile growth of both single- and two-species cultures. Planktonic bacterial growth was examined through OD₆₄₀ measurement. One-day-old biofilms were evaluated in terms of biofilm biomass (CV), respiratory activity (XTT), and CFU number. Conditioned media obtained either in biofilm or in planktonic mode of life triggered a synergistic effect on planktonic growth, mainly for E. coli single cultures growing in P. aeruginosa supernatants. Biofilms grown in the presence of P. aeruginosa biofilms-derived metabolites presented less mass and activity. These events highlight that, when developed in biofilm, P. aeruginosa release signals or metabolites able to prejudice single and binary biofilm growth of others species and of their own species. However, products released by their planktonic counterparts did not impair biofilm growth or activity. E. coli, living as planktonic or sessile cultures, released signals and metabolites or removed un-beneficial compounds which promoted the growth and activity of all the species. Our findings revealed that inter and intraspecies behaviors depend on the involved bacteria and their adopted mode of life.     

Cold shock domain proteins in the extremophyte <Emphasis Type="Italic">Thellungiella salsuginea</Emphasis> (salt cress): Gene structure and differential response to cold

Four genes encoding cold shock domain (CSD) proteins have been identified in salt cress Thellungiella salsuginea (halophila), an extremophyte currently recognized as a promising model for studying stress tolerance]. The deduced proteins prove highly homologous to those of Arabidopsis thaliana (up to 95% identity) and are accordingly enumerated TsCSDP1-TsCSDP4; after the N-proximal conserved CSD, they have respectively 6, 2, 7, and 2 zinc finger motifs evenly spaced by Gly-rich stretches. Much lower similarity (∼45%) is observed in the regions upstream of TATA-box promoters of TsCSDP1 vs. AtCSP1, with numerous distinctions in the sets of identifiable cis-regulatory elements. Plasmid expression of TsCSDP1 (like AtCSP1/3) rescues a cold-sensitive csp-lacking mutant of Escherichia coli, confirming that the protein is functional. In leaves of salt cress plants under normal conditions, the mRNA levels for the four TsCSDPs relate as 10: 27: 1: 31. Chilling to 4°C markedly alters the gene expression; the 4-day dynamics are different for all four genes and quite dissimilar from those reported for their Arabidopsis homologous under comparable conditions. Thus, the much greater cold hardiness of Thellungiella vs. Arabidopsis cannot be explained by structural distinctions of its CSDPs, but rather may be due to expedient regulation of their expression at low temperature.     

Locomotion and fine structure of parapodia in <Emphasis Type="Italic">Myzostoma cirriferum</Emphasis> (Myzostomida)

Most myzostomids are ectocommensals of crinoids on which they move freely. Their locomotion is ensured by five pairs of parapodia located laterally below their trunk. Each parapodium in Myzostoma cirriferum is a conical structure that includes a hook-like chaeta, replacement chaetae and an aciculum. Structure and ultrastructure of the myzostomid chaetae are similar to those of polychaetes: they are formed by a chaetoblast, which gives rise to microvilli where chaetal material is assembled on the outer surface. Myzostoma cirriferum walks on its host. It moves the anterior part, the posterior part or the lateral parts forwards but is able to rotate of 180° on itself. Its locomotion entirely depends on parapodial motions and not on trunk movements. Three pairs of muscles are involved in parapodial motions: parapodium flexor and parapodium extensor, aciculum protractor and aciculum retractor, and hook protractor with conjunctor. A functional model is proposed for explaining the global motion of a parapodium in M. cirriferum that may be extended to all ectocommensal myzostomids.