Simplicity-correlated size growth of the nuclear 28S ribosomal RNA D3 expansion segment in the crustacean order isopoda

The expansion segments within the eukaryote nuclear 23S-like ribosomal RNA molecule are now well characterized in many diverse organisms. A different base compositional bias, a higher propensity for size variability, and an increased evolutionary rate distinguish these regions from the universally conserved core regions of the molecule. In addition, some expansion segments of higher eukaryotes exhibit significant sequence simplicity which is hypothesized to occur by slippage-mediated mutational processes. We describe the discovery of extreme size variation of the D3 expansion segment in the crustacean order Isopoda. Among 11 species D3 varies in size from 180 to 518 nucleotides but maintains a homologous secondary structure. The D3 size is significantly positively correlated to relative simplicity factor (RSF), indicating that growth is most likely by insertion of simple sequences. D3 size and RSF correlate approximately with a morphology-based phylogeny, and within oniscideans RSF increases as more recent divergences occur. The D3 ofArmadillidium vulgare, with an RSF of 1.87, is the highest value recorded for any known expansion segment. Regions of high sequence simplicity in nuclear ribosomal RNA were previously only known from the higher vertebrate lineage. Here we demonstrate that this phenomenon occurs in a more extreme condition within a monophyletic invertebrate lineage. The extreme size changes identified could indicate that expansion segments are an extraneous element in the functioning ribosome.     

Fast effects of aldosterone on electrolytes in human lymphocytes are mediated by the sodium-proton-exchanger of the cell membrane

An in vitro effect of aldosterone on intracellular sodium, potassium and calcium concentrations and cell volume was shown in human mononuclear leukocytes (HML). In the present paper the effect of aldosterone on propionate-induced swelling of HML and, thereby, the Na+/H+-exchanger of the cell membrane was investigated. Aldosterone in physiological concentrations stimulated the cell swelling from 3 min on significantly. This effect had to be attributed to an increased activity of the Na+/H+-exchanger since amiloride blocked it almost completely. The inhibitor of protein synthesis actinomycin D prevented this action, cortisol had an intermediate effect. These data are the first to identify the Na+/H+-exchanger as a primary target of mineralocorticoid action in HML.     

Comparative analysis of the site-specific N-glycosylation of human lactoferrin produced in maize and tobacco plants.

We have compared the site-by-site N-glycosylation status of human lactoferrin (Lf) produced in maize, a monocotyledon, and in tobacco, used as a model dicotyledon. Maize and tobacco plants were stably transformed and recombinant Lf was purified from both seeds and leaves. N-glycopeptides were generated by trypsin digestion of recombinant Lf and purified by reverse-phase HPLC. The N-glycosylation pattern of each site was determined by mass spectrometry. Our results indicated that the N-glycosylation patterns of recombinant Lf produced in maize and tobacco share common structural features. In particular, both N-glycosylation sites of each recombinant Lf are mainly substituted by typical plant paucimannose-type N-glycans, with beta1,2-xylose and alpha1,3-linked fucose at the proximal N-acetylglucosamine. However, tobacco Lf shows a significant amount of processed N-glycans with one or two beta1,2GlcNAc linked to the trimannose core, which are weakly expressed in maize Lf. Finally, no Lewisa epitope was observed on tobacco Lf.     

A kinetic method for determination of free vanadium(IV) and (V) at trace level concentrations

A kinetic method based on alkaline phosphatase has been developed to measure free trace levels of vanadium(IV) and (V). The method involves measuring the rate of the alkaline phosphatase-catalyzed hydrolysis of p-nitrophenyl phosphate with (Vi) and without (Vo) a competitive inhibitor in the assay. Michaelis-Menten kinetics for a competitive inhibitor was used to express the relationship between Vo/Vi and the inhibitor concentration. Measuring both Vo and Vi thus yields a Vo/Vi ratio that allows calculation of the competitive inhibitor concentration. Determination of free vanadium in complex fluids can be accomplished by comparing the ratio of rates of p-nitrophenyl phosphate hydrolysis with and without a sequestering agent to the ratios of rates measured on addition of a known vanadium concentration. Free vanadium(V) can conveniently be measured from 10(-7) to 10(-5) M and free vanadium(IV) can be measured at 10(-8) M and above. The error limits on the vanadium determinations range from +/- 3 to +/- 12% of the concentration under investigation depending on the conditions under which the assay was conducted.     

Influence of nanofibers on growth and gene expression of human tendon derived fibroblast

Background  

Rotator cuff tears are a common and frequent lesion especially in older patients. The mechanisms of tendon repair are not fully understood. Common therapy options for tendon repair include mini-open or arthroscopic surgery. The use of growth factors in experimental studies is mentioned in the literature. Nanofiber scaffolds, which provide several criteria for the healing process, might be a suitable therapy option for operative treatment. The aim of this study was to explore the effects of nanofiber scaffolds on human tendon derived fibroblasts (TDF's), as well as the gene expression and matrix deposition of these fibroblasts.     

Slow clearance of Plasmodium vivax with chloroquine amongst children younger than six months of age in the Brazilian Amazon

Plasmodium vivax is the most widespread parasite causing malaria, being especially prevalent in the Americas and Southeast Asia. Children are one of the most affected populations, especially in highly endemic areas. However, there are few studies evaluating the therapeutic response of infants with vivax malaria. This study retrospectively evaluated the parasitaemia clearance in children diagnosed with vivax malaria during the first five days of exclusive treatment with chloroquine (CQ). Infants aged less than six months old had a significantly slower parasitaemia clearance time compared to the group of infants and children between six months and 12 years old (Kaplan-Meier survival analysis; Wilcoxon test; p = 0.004). The impaired clearance of parasitaemia in younger children with vivax malaria is shown for the first time in Latin America. It is speculated that CQ pharmacokinetics in young children with vivax malaria is distinct, but this specific population may also allow the detection of CQ-resistant parasites during follow-up, due to the lack of previous immunity.     

Preparative chromatography for obtaining pure samples of rosaniline,magenta II,and new fuchsine

A simple low pressure liquid chromatographic method is reported that can separate the basic fuchsine homologues, rosaniline, magenta II and new fuchsine from an impure commercial dye. The chromatographic purity of the separated dyes is > 90%. All homologues were obtained in multi-milligram amounts per chromatographic run; precise yields depend on the composition of the starting material and potentially may be greater. This is a useful preparative procedure for generating chromatographically pure samples of basic fuchsine homologues, especially those that cannot be obtained in pure form by direct synthesis.     

Complete Genome Sequence of the Aerobic Facultative Methanotroph Methylocella silvestris BL2

Methylocella silvestris BL2 is an aerobic methanotroph originally isolated from an acidic forest soil in Germany. It is the first fully authenticated facultative methanotroph. It grows not only on methane and other one-carbon (C₁) substrates, but also on some compounds containing carbon-carbon bonds, such as acetate, pyruvate, propane, and succinate. Here we report the full genome sequence of this bacterium.Methylocella spp. are abundant in acidic soils and wetlands and help attenuate methane emissions from these habitats (2). They are unique in several ways compared to all other known aerobic methanotrophs. Notably, they lack extensive internal membrane systems and also appear to lack the particulate methane monooxygenase (pMMO) enzyme found in all other methanotrophs (6). Instead, they use only a soluble methane monooxygenase (sMMO) for methane oxidation. In addition, Methylocella spp. are not limited like other methanotrophs to growing on one-carbon (C₁) compounds but also utilize a number of multicarbon compounds (3). The genome of Methylocella silvestris BL2 (4) was sequenced, assembled, and annotated by the Joint Genome Institute (U.S. Department of Energy; http://www.jgi.doe.gov/sequencing/strategy.html). A total of 38,459 reads (∼6× coverage), including 32,993 paired-end shotgun Sanger reads, 5,040 Roche 454 reads, and 580 finishing reads were included in the final assembly. Three lanes of Solexa data were used to polish the project.The genome size is 4.3 Mbp. The G+C content is 63%. In total, 3,917 candidate genes were predicted and 99 pseudogenes were found. Functionality was assigned to 67.9% of the genes, while 30.9% of the genes could not be assigned any known function. Based on BLASTP searches against the KEGG (Kyoto Encyclopedia of Genes and Genomes) database, 3,413 out of 3,917 (87.1%) candidate genes have significant similarity to genes from Proteobacteria. Only 11 and 14 genes have best hits to genes from Archaea and Eukarya, respectively. All tRNA-encoding regions were identified, and two identical rRNA operons were found.The absence of any pmoCAB genes encoding a pMMO enzyme that is present in all other genera of methanotrophs is now conclusively verified by the genome sequence. A complete operon encoding sMMO (mmoXYBZDC) was verified, as was a complete operon encoding methanol dehydrogenase (mxaFJGIRSACKLDEH) and all genes necessary for fixation of methane-derived carbon via the serine cycle. Genes encoding key enzymes in both the tetrahydrofolate and the tetrahydromethanopterin-mediated formaldehyde oxidation pathways were found.M. silvestris can grow on two-carbon compounds, particularly acetate. Acetate kinase- and phosphotransacetylase-encoding genes are present, allowing acetate to be fed into the tricarboxylic acid (TCA) cycle. Genes encoding glyoxylate bypass enzymes (i.e., isocitrate lyase and malate synthase) have been identified. This pathway is essential for bacteria when growing on two-carbon compounds (1). The bacterium can also grow on C₃ and C₄ compounds, and a full gene set encoding enzymes of the TCA cycle is present, including genes encoding α-ketoglutarate dehydrogenase, which are lacking in some methanotrophs. Interestingly, a gene cluster encoding di-iron-containing multi-component propane monooxygenase is also present.The genome sequence of M. silvestris is the first genome available for an alphaproteobacterial methanotroph. It joins the gammaproteobacterial methanotroph Methylococcus capsulatus Bath (7) and the verrucomicrobial methanotroph “Methylacidiphilum infernorum” (5). More detailed analyses of the genome as well as comparative analysis with obligate methanotrophs will provide deeper insight into the metabolism of this fascinating bacterium.