Inhibiton by sulfanilamide of sporulation in Saccharomyces cerevisiae.

The antimetabolite sulfanilamide inhibits sporulation in Saccharomyces cerevisiae strain AP1. Cells exposed to sulfanilamide at various times during the sporulation process become progressively insensitive to the drug, although accumulation of sulfanilamide by the cells increases with time. Vegetative growth of AP1 is practically unaffected by sulfanilamide; pregrowth of the cells in the presence of the drug does not prevent sporulation. Thus, inhibition is confined to the meiotic phase of the cell cycle. Sensitivity to sulfanilamide is independent of pH. Increasing the time cells are exposed to sulfanilamide results in a progressive reduction of ascus formation; however, the inhibition is reversible since sporulation can occur in cells exposed to the drug for greater than 24 h. The drug arrests the cells at a point before commitment to sporulation, since yeast cells exposed to sulfanilamide for 12 h do not complete the sporulation process when returnedto vegetative medium, but resume mitotic growth instead. Meiotic nuclear division is largely prevented by sulfanilamide, and synthesis of RNA and protein is severely retarded. DNA synthesis is inhibited up to 50%; glycogen synthesis is approximately 90% inhibited. Other yeast strains showed varying sensitivity to sulfanilamide; homothallic strains were generally less affected.     

Evaluation of the usefulness of a new direct immunofluorescence assay (ScanVIT‐Legionella™) for monitoring hospital water systems contaminated with Legionella spp.

Aims: To compare the efficiency of the ScanVIT‐Legionella^? test (Vermicon, Munich, Germany) vs a conventional culture method for the quantification of Legionella spp. in hospital water samples in daily hospital practice. Methods and Results: The detection of Legionella spp. takes place on a cultivated filter brought into contact with dye‐marked gene probes. The results are analysed under fluorescence microscopy. Bacteria that light up green belong to the genus Legionella; those that light up both green and red belong to the species Legionella pneumophila. Our results showed that the ScanVIT test has a sensitivity of 90%; agreement between the two methods was 82%. In the 48 samples that tested positive with both methods, the Legionella concentration detected by the culture method was consistently higher. A statistically significant difference between the results obtained with the two test methods emerged at the Wilcoxon test (P < 0·001). Conclusion: The ScanVIT test may be recommended for investigating the presence of Legionella by qualitative testing. Significance and Impact of the Study: Given the simplicity of colony identification by fluorescence, the ScanVIT test can be used in laboratories where staffs are not experienced in identifying typical colonies of Legionella.     

Physio-pathological roles of transglutaminase-catalyzed reactions

Transglutaminases (TGs) are a large family of related and ubiquitous enzymes that catalyze post-translational modifications of proteins. The main activity of these enzymes is the cross-linking of a glutaminyl residue of a protein/peptide substrate to a lysyl residue of a protein/peptide co-substrate. In addition to lysyl residues, other second nucleophilic co-substrates may include monoamines or polyamines (to form mono- or bi-substituted /crosslinked adducts) or -OH groups (to form ester linkages). In the absence of co-substrates, the nucleophile may be water, resulting in the net deamidation of the glutaminyl residue. The TG enzymes are also capable of catalyzing other reactions important for cell viability. The distribution and the physiological roles of TG enzymes have been widely studied in numerous cell types and tissues and their roles in several diseases have begun to be identified. "Tissue" TG (TG2), a member of the TG family of enzymes, has definitely been shown to be involved in the molecular mechanisms responsible for a very widespread human pathology: i.e. celiac disease (CD). TG activity has also been hypothesized to be directly involved in the pathogenetic mechanisms responsible for several other human diseases, including neurodegenerative diseases, which are often associated with CD. Neurodegenerative diseases, such as Alzheimer's disease, Parkinson's disease, supranuclear palsy, Huntington's disease and other recently identified polyglutamine diseases, are characterized, in part, by aberrant cerebral TG activity and by increased cross-linked proteins in affected brains. In this review, we discuss the physio-pathological role of TG-catalyzed reactions, with particular interest in the molecular mechanisms that could involve these enzymes in the physio-pathological processes responsible for human neurodegenerative diseases.     

Viral Glycoproteins Accumulate in Newly Formed Annulate Lamellae following Infection of Lymphoid Cells by Human Herpesvirus 6

Ultrastructural analysis of HSB-2 T-lymphoid cells and human cord blood mononuclear cells infected with human herpesvirus 6 revealed the presence, in the cell cytoplasm, of annulate lamellae (AL), which were absent in uninfected cells. Time course analysis of the appearance of AL following viral infection showed that no AL were visible within the first 72 h postinfection and that their formation correlated with the expression of the late viral glycoprotein gp116. The requirement of active viral replication for AL neoformation was further confirmed by experiments using inactivated virus or performed in presence of the viral DNA polymerase inhibitor phosphonoacetic acid. Both conventional electron microscopic examination and immunogold fracture labeling with anti-endoplasmic reticulum antibodies indicated a close relationship of AL with the endoplasmic reticulum and nuclear membranes. However, when the freeze-fractured cells were immunogold labeled with an anti-gp116 monoclonal antibody, AL membranes were densely labeled, whereas nuclear membranes and endoplasmic reticulum cisternae appeared virtually unlabeled, showing that viral envelope glycoproteins selectively accumulate in AL. In addition, gold labeling with Helix pomatia lectin and wheat germ agglutinin indicated that AL cisternae, similar to cis-Golgi membranes, contain intermediate, but not terminal, forms of glycoconjugates. Taken together, these results suggest that in this cell-virus system, AL function as a viral glycoprotein storage compartment and as a putative site of O-glycosylation.     

A Psychological Exploration of Engagement in Geek Culture

Geek culture is a subculture of enthusiasts that is traditionally associated with obscure media (Japanese animation, science fiction, video games, etc.). However, geek culture is becoming increasingly mainstream; for example, in the past year alone, Dragon*Con, a major Geek convention in Atlanta, Georgia, attracted an attendance of over 57,000 members. The present article uses an individual differences approach to examine three theoretical accounts of geek culture. Seven studies (N = 2354) develop the Geek Culture Engagement Scale (GCES) to quantify geek engagement and assess its relationships to theoretically relevant personality and individual differences variables. These studies present evidence that individuals may engage in geek culture in order to maintain narcissistic self-views (the great fantasy migration hypothesis), to fulfill belongingness needs (the belongingness hypothesis), and to satisfy needs for creative expression (the need for engagement hypothesis). Geek engagement is found to be associated with elevated grandiose narcissism, extraversion, openness to experience, depression, and subjective well-being across multiple samples. These data lay the groundwork for further exploration of geek culture as well as provide a foundation for examining other forms of subculture participation.     

Optimizing Human Synovial Fluid Preparation for Two-Dimensional Gel Electrophoresis

Background

Prenatal screening for Down Syndrome (DS) would benefit from an increased number of biomarkers to improve sensitivity and specificity. Improving sensitivity and specificity would decrease the need for potentially risky invasive diagnostic procedures.

Results

We have performed an in depth two-dimensional difference gel electrophoresis (2D DIGE) study to identify potential biomarkers. We have used maternal plasma samples obtained from first and second trimesters from mothers carrying DS affected fetuses compared with mothers carrying normal fetuses. Plasma samples were albumin/IgG depleted and expanded pH ranges of pH 4.5 - 5.5, pH 5.3 - 6.5 and pH 6 - 9 were used for two-dimensional gel electrophoresis (2DE). We found no differentially expressed proteins in the first trimester between the two groups. Significant up-regulation of ceruloplasmin, inter-alpha-trypsin inhibitor heavy chain H4, complement proteins C1s subcomponent, C4-A, C5, and C9 and kininogen 1 were detected in the second trimester in maternal plasma samples where a DS affected fetus was being carried. However, ceruloplasmin could not be confirmed as being consistently up-regulated in DS affected pregnancies by Western blotting.

Conclusions

Despite the in depth 2DE approach used in this study the results underline the deficiencies of gel-based proteomics for detection of plasma biomarkers. Gel-free approaches may be more productive to increase the number of plasma biomarkers for DS for non-invasive prenatal screening and diagnosis.     

Mirna expression profiles identify drivers in colorectal and pancreatic cancers

Background and Aim

Altered expression of microRNAs (miRNAs) hallmarks many cancer types. The study of the associations of miRNA expression profile and cancer phenotype could help identify the links between deregulation of miRNA expression and oncogenic pathways.

Methods

Expression profiling of 866 human miRNAs in 19 colorectal and 17 pancreatic cancers and in matched adjacent normal tissues was investigated. Classical paired t-test and random forest analyses were applied to identify miRNAs associated with tissue-specific tumors. Network analysis based on a computational approach to mine associations between cancer types and miRNAs was performed.

Results

The merge between the two statistical methods used to intersect the miRNAs differentially expressed in colon and pancreatic cancers allowed the identification of cancer-specific miRNA alterations. By miRNA-network analysis, tissue-specific patterns of miRNA deregulation were traced: the driving miRNAs were miR-195, miR-1280, miR-140-3p and miR-1246 in colorectal tumors, and miR-103, miR-23a and miR-15b in pancreatic cancers.

Conclusion

MiRNA expression profiles may identify cancer-specific signatures and potentially useful biomarkers for the diagnosis of tissue specific cancers. miRNA-network analysis help identify altered miRNA regulatory networks that could play a role in tumor pathogenesis.     

Tetrahydroisoquinoline derivatives as melatonin MT2 receptor antagonists

A series of tetrahydroisoquinolines has yielded potent MT(2) receptor antagonists, which are selective versus the MT(1) receptor.     

Extreme difference in rate of mitochondrial and nuclear DNA evolution in a large ectotherm, Galápagos tortoises

We sequenced approximately 4.5 kb of mtDNA from 161 individuals representing 11 named taxa of giant Galápagos tortoises (Geochelone nigra) and about 4 kb of non-coding nuclear DNA from fewer individuals of these same 11 taxa. In comparing mtDNA and nucDNA divergences, only silent substitutions (introns, ITS, mtDNA control region, and synonymous substitutions in coding sequences) were considered. mtDNA divergence was about 30 times greater than that for nucDNA. This rate discrepancy for mtDNA and nucDNA is the greatest yet documented and is particularly surprising for large ectothermic animals that are thought to have relatively low rates of mtDNA evolution. This observation may be due to the somewhat unusual reproductive biology and biogeographic history of these organisms. The implication is that the ratio of effective population size of nucDNA/mtDNA is much greater than the usually assumed four. The nearly neutral theory of molecular evolution predicts this would lead to a greater difference between rates of evolution.     

Protective role of the complement regulatory protein human CD-55 in cardiac xenograft: a descriptive study and a revision of the literature

The limited and inadequate availability of organs from human donors has resulted in the utilisation of xenografts as an alternative tool. Nevertheless, hyperacute rejection (HAR) following xenograft determines the loss of the transplanted organ. The "primum movens" is the activation of the complement pathway mediated by the binding of natural xenogenic antibodies to the endothelium of the graft, followed by the lysis of the endothelial cells with subsequent oedema, thrombosis and necrosis of the transplanted organ. In this work we describe morphological and biomolecular observations of isolated human-decay accelerating factor (h-DAF, CD55) transgenic pig hearts, after perfusion for four hours with human blood. H-DAF is a membrane glycoprotein inhibiting the complement activation in humans. We describe considerably reduced damages in transgenic hearts, compared to controls. The cardiac function resulted preserved. Our data are in agreement with what was already shown by other groups using different experimental models. In conclusion, we encourage the use of new sources of transgenic animals, pointing out the importance of morphological analysis in evaluation of xenograft.