Clinical heterogeneity in the tricho-dento-osseous syndrome

The tricho-dento-osseous syndrome (TDO syndrome) involves morphologic abnormalities of hair, teeth, and skeleton. Clinical findings of the TDO syndrome are excessively curly (fuzzy) hair, enamel hypoplasia, and skeletal findings of a generalized pattern of osseous sclerosis. We report an autosomal dominant syndrome with similar hair and teeth morphology, but with a skeletal dysplasia consisting of sclerosis and thickening of the calvarium with long bones that show subtle undertubulation but no sclerosis.     

Sensitive high-performance liquid chromatographic assay for endralazine and two of its metabolites in human plasma

Endralazine (I) is a new antihypertensive which is chemically and pharmacologically related to hydralazine and dihydralazine. A sensitive high-performance liquid chromatographic-fluorescence assay for the drug and two of its metabolites [methyltriazoloendralazine (VII) and hydroxymethyltriazoloendralazine (VIII)] in human plasma was developed. After conversion of I and its internal standard to triazolopyridopyridazine derivatives the latter and metabolites were separated by high-performance liquid chromatography and detected using their fluorescence. The limits of detection of the assay were 1 nmol/l for I and VII and 0.1 nmol/l for VIII. Intra-assay coefficients of variation were 2.5–5.1% for I (range 1000–10 nmol/l), 4.2–4.5% for VII (range 100–5 nmol/l) and 3.4–5.7% for VIII (range 100–1 nmol/l). Following oral administration of 5 and 10 mg of I to two normal volunteers (slow acetylators) peak plasma levels of I occurred between 0.75 and 1 h after the dose, and declined in a biexponential fashion. The terminal half-life ranged from 2.8–3.7 h. These results contrast with those obtained for hydralazine in plasma where in vitro and in vivo half-lives were 30 min.     

Isolation and characterization of rat and human glyceraldehyde-3-phosphate dehydrogenase cDNAs: genomic complexity and molecular evolution of the gene.

Full length cDNAs encoding the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from rat and man have been isolated and sequenced. Many GAPDH gene-related sequences have been found in both genomes based on genomic blot hybridization analysis. Only one functional gene product is known. Results from genomic library screenings suggest that there are 300-400 copies of these sequences in the rat genome and approximately 100 in the human genome. Some of these related sequences have been shown to be processed pseudogenes. We have isolated several rat cDNA clones corresponding to these pseudogenes indicating that some pseudogenes are transcribed. Rat and human cDNAs are 89% homologous in the coding region, and 76% homologous in the first 100 base pairs of the 3'-noncoding region. Comparison of these two cDNA sequences with those of the chicken, Drosophila and yeast genes allows the analysis of the evolution of the GAPDH genes in detail.     

Protein tyrosine phosphatase-1B dephosphorylation of the insulin receptor occurs in a perinuclear endosome compartment in human embryonic kidney 293 cells

Protein tyrosine phosphatase-1B (PTP-1B) is a negative regulator of insulin signaling. It is thought to carry out this role by interacting with and dephosphorylating the activated insulin receptor (IR). However, little is known regarding the nature of the cellular interaction between these proteins, especially because the IR is localized to the plasma membrane and PTP-1B to the endoplasmic reticulum. Using confocal microscopy and fluorescence resonance energy transfer (FRET), the interaction between PTP-1B and the IR was examined in co-transfected human embryonic kidney 293 cells. Biological activities were not significantly affected for either PTP-1B or the IR with the fusion of W1B-green fluorescent protein (GFP) to the N terminus of PTP-1B (W1B-PTP-1B) or the fusion of Topaz-GFP to the C terminus of the IR (Topaz-IR). FRET between W1B and Topaz was monitored in cells transfected with either wild type PTP-1B (W1B-PTP-1B) or the substrate-trapping form PTP-1B(D181A) (W1B-PTP-1B(D181A)) and Topaz-IR. Co-expression of W1B-PTP-1B with Topaz-IR resulted in distribution of Topaz-IR to the plasma membrane, but no FRET was obtained upon insulin treatment. In contrast, co-expression of W1B-PTP-1B(D181A) with Topaz-IR caused an increase in cytosolic Topaz-IR fluorescence and, in some cells, a significant basal FRET signal, suggesting that PTP-1B is interacting with the IR during its synthesis. Stimulation of these cells with insulin resulted in a rapid induction of FRET that increased over time and was localized to a perinuclear spot. Co-expression of Topaz-IR with a GFP-labeled RhoB endosomal marker and treatment of the cells with insulin identified a perinuclear endosome compartment as the site of localization. Furthermore, the insulin-induced FRET could be prevented by the treatment of the cells with a specific PTP-1B inhibitor. These results suggest that PTP-1B appears not only to interact with and dephosphorylate the insulin-stimulated IR in a perinuclear endosome compartment but is also involved in maintaining the IR in a dephosphorylated state during its biosynthesis.     

Host cell preference and variable transmission strategies in malaria parasites

Malaria and other haemosporin parasites must undergo a round of sexual reproduction in their insect vector in order to produce stages that can be transmitted to vertebrate hosts. Consequently, it is crucial that parasites produce the sex ratio (proportion of male sexual stages) that will maximize the number of fertilization and thus, transmission to new vertebrate hosts. There is some evidence to show that, consistent with evolutionary theory, the sex ratios of malaria parasites are negatively correlated to their inbreeding rate. However, recent theory has shown that when fertilization success is compromised, parasites should respond by increasing their investment in sexual stages or by producing a less female biased ration than predicted by their inbreeding rate alone. Here, we show that two species of rodent malaria, Plasmodium chabaudi and Plasmodium vinckei petteri, adopt different strategies in response to host anaemia, a factor though to compromise transmission success: P. chabaudi increases investment in sexual stages, whereas P. vinckei produces a less female biased sex ratio. We suggest that these different transmission strategies may be due to marked differences in host cell preference.     

Involvement of claudin-7 in HIV infection of CD4(-) cells

Background

The antibacterial activity of host defense peptides (HDP) is largely mediated by permeabilization of bacterial membranes. The lipid membrane of enveloped viruses might also be a target of antimicrobial peptides. Therefore, we screened a panel of naturally occurring HDPs representing different classes for inhibition of early, Env-independent steps in the HIV replication cycle. A lentiviral vector-based screening assay was used to determine the inhibitory effect of HDPs on early steps in the replication cycle and on cell metabolism.

Results

Human LL37 and porcine Protegrin-1 specifically reduced lentiviral vector infectivity, whereas the reduction of luciferase activities observed at high concentrations of the other HDPs is primarily due to modulation of cellular activity and/or cytotoxicity rather than antiviral activity. A retroviral vector was inhibited by LL37 and Protegrin-1 to similar extent, while no specific inhibition of adenoviral vector mediated gene transfer was observed. Specific inhibitory effects of Protegrin-1 were confirmed for wild type HIV-1.

Conclusion

Although Protegrin-1 apparently inhibits an early step in the HIV-replication cycle, cytotoxic effects might limit its use as an antiviral agent unless the specificity for the virus can be improved.     

Tuning of Catalytic Activity by Thermoelectric Materials for Carbon Dioxide Hydrogenation

An innovative use of a thermoelectric material (BiCuSeO) as a support and promoter of catalysis for CO₂ hydrogenation is reported here. It is proposed that the capability of thermoelectric materials to shift the Fermi level and work function of a catalyst lead to an exponential increase of catalytic activity for catalyst particles deposited on its surface. Experimental results show that the CO₂ conversion and CO selectivity are increased significantly by a thermoelectric Seebeck voltage. This suggests that the thermoelectric effect can not only increase the reaction rate but also change chemical equilibrium, which leads to the change of thermodynamic equilibrium for the conversion of CO₂ in its hydrogenation reactions. It is also shown that this thermoelectric promotion of catalysis enables BiCuSeO oxide itself to have a high catalytic activity for CO₂ hydrogenation. The generic nature of the mechanism suggests the possibility that many catalytic chemical reactions can be tuned in situ to achieve much higher reaction rates, or at lower temperatures, or have better desired selectivity through changing the backside temperature of the thermoelectric support.     

Phylogeography and connectivity of molluscan parasites: Perkinsus spp. in Panama and beyond

Panama is a major hub for commercial shipping between two oceans, making it an ideal location to examine parasite biogeography, potential invasions, and the spread of infectious agents. Our goals were to (i) characterise the diversity and genetic connectivity of Perkinsus spp. haplotypes across the Panamanian Isthmus and (ii) combine these data with sequences from around the world to evaluate the current phylogeography and genetic connectivity of these widespread molluscan parasites. We collected 752 bivalves from 12 locations along the coast of Panama including locations around the Bocas del Toro archipelago and the Caribbean and Pacific entrances to the Panama Canal, from December 2012 to February 2013. We used molecular genetic methods to screen for Perkinsus spp. and obtained internal transcribed spacer region (ITS) ribosomal DNA (rDNA) sequences for all positive samples. Our sequence data were used to evaluate regional haplotype diversity and distribution across both coasts of Panama, and were then combined with publicly available sequences to create global haplotype networks. We found 26 ITS haplotypes from four Perkinsus spp. (1–12 haplotypes per species) in Panama. Perkinsus beihaiensis haplotypes had the highest genetic diversity, were the most regionally widespread, and were associated with the greatest number of hosts. On a global scale, network analyses demonstrated that some haplotypes found in Panama were cosmopolitan (Perkinsus chesapeaki, Perkinsus marinus), while others were more geographically restricted (Perkinsus olseni, P. beihaiensis), indicating different levels of genetic connectivity and dispersal. We found some Perkinsus haplotypes were shared across the Isthmus of Panama and several regions around the world, including across ocean basins. We also found that haplotype diversity is currently underestimated and directly related to the number of sequences. Nevertheless, our results demonstrate long-range dispersal and global connectivity for many haplotypes, suggesting that dispersal through shipping probably contributes to these biogeographical patterns.     

Preliminary crystallographic analysis of the breakage-reunion domain of the Escherichia coli DNA gyrase A protein

The 64 x 10(3) Mr N-terminal breakage-reunion domain of the Escherichia coli DNA gyrase A protein was purified from an over-expressing strain. When complexed with the gyrase B protein, this truncated A protein has all of the enzymic properties of the full-length counterpart, although with reduced efficiency in some cases. The 64 x 10(3) Mr protein has been crystallized in several forms, a number of which were too small for crystallographic analysis. However, two forms grew to sufficient size for preliminary X-ray analysis. Both forms were tetragonal with a primitive lattice. One form (type I) had cell dimensions of a = b = 170 A, c = 145 A a space group of either P41212 (P43212) or P42212, and diffracted to 6 A resolution. The type II crystals had cell dimensions of a = b = 177 A, c = 175 A, a space group of P41212 (P43212) or P42212, and diffracted to at least 4.5 A resolution. Both crystal forms apparently contained four subunits (possibly a tetramer) in the asymmetric unit. We are attempting to increase the size and quality of these crystals.