Enhanced flux through the methylerythritol 4-phosphate pathway in Arabidopsis plants overexpressing deoxyxylulose 5-phosphate reductoisomerase

The methylerythritol 4-phosphate (MEP) pathway synthesizes the precursors for an astonishing diversity of plastid isoprenoids, including the major photosynthetic pigments chlorophylls and carotenoids. Since the identification of the first two enzymes of the pathway, deoxyxylulose 5-phoshate (DXP) synthase (DXS) and DXP reductoisomerase (DXR), they both were proposed as potential control points. Increased DXS activity has been shown to up-regulate the production of plastid isoprenoids in all systems tested, but the relative contribution of DXR to the supply of isoprenoid precursors is less clear. In this work, we have generated transgenic Arabidopsis thaliana plants with altered DXS and DXR enzyme levels, as estimated from their resistance to clomazone and fosmidomycin, respectively. The down-regulation of DXR resulted in variegation, reduced pigmentation and defects in chloroplast development, whereas DXR-overexpressing lines showed an increased accumulation of MEP- derived plastid isoprenoids such as chlorophylls, carotenoids, and taxadiene in transgenic plants engineered to produce this non-native isoprenoid. Changes in DXR levels in transgenic plants did not result in changes in␣DXS gene expression or enzyme accumulation, confirming that the observed effects on plastid isoprenoid levels in DXR-overexpressing lines were not an indirect consequence of altering DXS levels. The results indicate that the biosynthesis of MEP (the first committed intermediate of the pathway) limits the production of downstream isoprenoids in Arabidopsis chloroplasts, supporting a role for DXR in the control of the metabolic flux through the MEP pathway.     

Inferring the effects of demography and selection on Drosophila melanogaster populations from a chromosome-wide scan of DNA variation

Identifying regions of the Drosophila melanogaster genome that have been recent targets of positive Darwinian selection will provide evidence for adaptations that have helped this species to colonize temperate habitats. We have begun a search for such genomic regions by analyzing multiple loci (about 250) dispersed across the X chromosome in a putatively ancestral population from East Africa and a derived European population. For both populations we found evidence for past changes in population size. We estimated that a major bottleneck associated with the colonization of Europe occurred about 3,500-16,000 years ago. We also found that while this bottleneck can account for most of the reduction in variation observed in the European sample, there is a deficit of polymorphism in some genomic regions that cannot be explained by demography alone.     

The effect of aminoglycoside antibiotics on the adventitious regeneration from apricot leaves and selection of <Emphasis Type="Italic">npt</Emphasis>II-transformed leaf tissues

The effect of different concentrations of the aminoglycoside antibiotics, geneticin, paromomycin and streptomycin on adventitious regeneration from leaf explants of apricot was tested to design an alternative procedure for selecting transgenic shoots. Streptomycin and paromomycin reduced shoot regeneration percentage with increasing concentration of antibiotics. Almost a complete inhibition of regeneration was reached when 20M paromomycin was used, although up to 40M streptomycin was necessary to completely inhibit regeneration. Geneticin had a very toxic effect on apricot leaves and regeneration was inhibited at almost all concentrations tested. Addition of kanamycin hastened the development of adventitious buds although silver thiosulfate and not kanamycin was responsible for the observed increase in the consistency of the results from independent experiments. Kanamycin and paromomycin at the concentrations tested improved selection of transformed cells and resulted in a larger number of gfp-expressing regions. Paromomycin at 40 and 25.7M kanamycin improved proliferation of transformed tissues as compared with the other antibiotics used and non-selected controls.     

Shell recruitment in the Mediterranean hermit crab Clibanarius erythropus

Gastropod shells are vital for the majority of hermit crab species, being essential for their survival, growth, protection, and reproduction. Given their importance, shells are acquired and transferred between crabs through several modalities. We conducted observations and experiments at the Asinara Island (Sardinia, Italy) to investigate the efficacy of the different behavioral tactics adopted by the hermit crab Clibanarius erythropus to acquire shells, such as: (1) locomotion and activity at different tidal phases; (2) attendance at shell-supplying sites (simulated predation sites with five different odors: live and dead gastropods, live and dead crabs, predator); and (3) interactions with conspecifics in aggregations on simulated gastropod predation sites. In each tidal phase, locomotion was slow (0.7 cm min^− 1) and, as a consequence, the probability of encountering empty shells and conspecifics was low. Simulated gastropod predation sites quickly attracted a larger number of hermit crabs than the other sites tested. Aggregations seemed to function as shell exchange markets, as previously suggested for other species: the first attendant took the experimental shell and a chain of shell exchanges among conspecifics followed. Our results show that, in C. erythropus, aggregation is the most efficient tactic for the acquisition of new shells, whereas in other species, such as Pagurus longicarpus, it is associated with exploitation ability due to the intense locomotion. The interspecific plasticity in hermit crabs' behavior is confirmed.     

Human herpesvirus 8 (HHV-8/KSHV) and hematologic malignancies

Human herpesvirus 8 (HHV-8), also defined Kaposi's sarcoma (KS)-associated herpesvirus, was identified by Chang and colleagues in 1994 using purely molecular techniques, before any serological evidence or virus isolation in cell culture could be achieved. HHV-8 is unique among herpesviruses because its prevalence in the general population is low and because it possesses the richest weaponry of viral oncogenes and tumor-promoting factors ever described. Eleven HHV-8-specific genes are homologs of cellular genes, which were hijacked from the host during a long parallel evolution, and at least five of such genes show both in vitro and in vivo transforming ability. HHV-8 is the causative agent of KS, but it has also been associated with different hematologic malignancies, including primary effusion lymphoma (PEL), multicentric Castelman's disease (MCD), MCD-related immunoblastic/plasmablastic lymphoma and various atypical lymphoproliferative disorders. Although low-level silent infection was detected in bone marrow stromal cells from patients with multiple myeloma, a role of HHV-8 in this disease is unlikely. As seen with KS, the incidence of HHV-8-associated lymphoproliferative disorders is increased in the setting of human immunodeficiency virus infection.     

Assembly of truncated HCV core antigen into virus-like particles in Escherichia coli

Core protein is one of the most conserved and immunogenic of the hepatitis C virus proteins. Several pieces of experimental evidence suggest its ability for formation of virus like particles alone or in association with other viral proteins in mammalian or yeast cells with great similarity to those detected in patient sera and liver extract. In this work we report an Escherichia coli-derived truncated hepatitis C core protein that is able to aggregate. SDS-PAGE and size exclusion chromatography patterns bring to mind the aggregation of monomers of recombinant protein Co.120. The Co.120 protein migrated with buoyant density of 1.28 g/cm(3) when analyzed using CsCl density gradient centrifugation. Spherical structures with an average diameter of 30 nm were observed using electron microscopy. We report here that VLPs are generated when the first 120 aa of HCV core protein are expressed in E. coli.     

New exopolysaccharides produced by Aureobasidium pullulans grown on glucosamine

The polysaccharides produced by Aureobasidium pullulans, grown using glucosamine as the carbon source, were investigated by means of methylation analysis, affinity chromatography and NMR spectroscopy. The results indicated that, besides a small amount of pullulan, this micro-organism was capable of producing-in low yields-mixtures of at least two different complex polysaccharides containing mainly mannose and galactose. (1)H NMR spectra of two fractions obtained by lectin affinity chromatography indicated that one polymer was constituted exclusively of mannose residues while the other contained both galactofuranosyl and mannopyranosyl residues.     

Acidophilic character of yeast PID261/BUD32, a putative ancestor of eukaryotic protein kinases

Yeast piD261/Bud32 and its homologues are present in eukaryotes and in archaea but not in bacteria and are believed to make up a primordial branch of the eukaryotic protein kinase superfamily. Here, we show that, at variance with the majority of Ser/Thr protein kinases which recognize phosphoacceptor sites specified by basic and/or proline residues, piD261 phosphorylates in vitro a number of acidic proteins and peptides, and it recognizes seryl residues specified by carboxylic side chains. These data suggest that recognition of acidic sites might have been a primordial trait of protein kinases, which was modified during evolution to cope with the increasing complexity of protein phosphorylation in eukaryotes.     

Comparison of Two Different Actigraphs with Polysomnography in Healthy Young Subjects

The present study aimed to compare two commercially available actigraphs, with a concurrent polysomnographic (PSG) recording. Twelve healthy volunteers (six women; age range 19–28 yrs) simultaneously wore the Basic Mini‐Motionlogger® and Actiwatch® for seven overnight polysomnographic recordings. Comparisons of the following sleep measures were focused on: sleep onset latency (SOL), total sleep time, wake after sleep onset, and sleep efficiency. Both devices underestimated SOL in comparison to PSG, but they had similar performance compared to PSG for the other sleep measures. A limit of the study is that the results can be only generalized to healthy young subjects.     

Phosphorylation of NHERF1 S279 and S301 differentially regulates breast cancer cell phenotype and metastatic organotropism

Metastatic cancer cells are highly plastic for the expression of different tumor phenotype hallmarks and organotropism. This plasticity is highly regulated but the dynamics of the signaling processes orchestrating the shift from one cell phenotype and metastatic organ pattern to another are still largely unknown. The scaffolding protein NHERF1 has been shown to regulate the expression of different neoplastic phenotypes through its PDZ domains, which forms the mechanistic basis for metastatic organotropism. This reprogramming activity was postulated to be dependent on its differential phosphorylation patterns. Here, we show that NHERF1 phosphorylation on S279/S301 dictates several tumor phenotypes such as in vivo invasion, NHE1-mediated matrix digestion, growth and vasculogenic mimicry. Remarkably, injecting mice with cells having differential NHERF1 expression and phosphorylation drove a shift from the predominantly lung colonization (WT NHERF1) to predominately bone colonization (double S279A/S301A mutant), indicating that NHERF1 phosphorylation also acts as a signaling switch in metastatic organotropism.