The multiple T-maze in vivo testing of the neuroprotective effect of humanin analogues

Humanin (HN) and its analogues have been shown to protect cells against death induced by various Alzheimer's disease (AD) genes and amyloid-beta-peptides in vitro; the analogues [Gly(14)]-HN and colivelin have also been shown to be potent in reversing learning and memory impairment induced by scopolamine or quinuclidinyl benzilate (QNB) in mice or rats in vivo using the Y-maze or multiple T-maze tests. This paper describes the activity of new peptides of the HN family, after i.p. administration, on QNB-induced impairment of spatial memory in the multiple T-maze test in rats. The following peptides have been studied: HN analogues truncated either on the C- or N-terminus, or analogues having a tert-Leu in place of Leu in the central part of the molecule, the active HN core PAGASRLLLLTGEIDLP (RG-PAGA) and its analogues having three or five leucines instead of four, and finally the recently described hybrid peptide colivelin (i.e. a peptide having the activity-dependent neurotrophic factor SALLRSIPA attached to the N-terminus of the active RG-PAGA) and its des-Leu- and plus-Leu-analogues. While the truncated analogues and most of the tert-Leu containing analogues were devoid of activity, the analogues of the RG-PAGA were active, i.e. they reversed the impairment of spatial memory irrespective of the number of Leu present in their sequence. The highest activity was shown by colivelin and its des-Leu-analogue. These results demonstrate the potential of HN analogues in the modulation of the cholinergic system, which plays an important role in the cognitive deficits associated with AD and other neurodegenerative diseases.     

Crystal structure of Rsr, an ortholog of the antigenic Ro protein, links conformational flexibility to RNA binding activity

Ro ribonucleoproteins are a class of antigenic ribonucleoproteins associated with rheumatic autoimmune diseases like systemic lupus erythematosus and Sj?grens syndrome in humans. Ro ribonucleoproteins are mostly composed of the 60-kDa Ro protein and small cytoplasmic RNAs, called Y RNAs, of unknown function. In eukaryotes, where Ro has been found to associate with damaged or mutant RNAs, it has been suggested that Ro may play a role in RNA quality control. In the radiation-resistant bacterium Deinococcus radiodurans and some eukaryotes, Ro has also been implicated in cell survival following UV damage. Here we present the first high resolution structure of a prokaryotic Ro ortholog, Rsr from D. radiodurans. The structure has been solved to 1.9 A resolution and shows distinct differences when compared with the eukaryotic apo- and RNA-bound Ro structures. Rsr is composed of two domains: a helical RNA binding domain and a mixed "von Willebrand factor A-like" domain containing a divalent metal binding site. Although the individual domains of Rsr are similar to the eukaryotic Ro, significantly large differences are seen at the interface of the two domains. Since this interface communicates with the conserved central cavity of Ro, which is implicated in RNA binding, changes at this interface could potentially influence RNA binding by Ro. Although the apo-Rsr protein is monomeric, Rsr binds Y RNA to form multimers of approximately 12 molecules of a 1:1 Rsr-Y RNA complex. Rsr binds D. radiodurans Y RNA with low nanomolar affinity, comparable with previously characterized eukaryotic Ro orthologs.     

Risk Factors for Contamination of Hotel Water Distribution Systems by Legionella Species

The Legionella colonization frequency at 385 Greek hotel hot and cold water distribution systems was 20.8%. Legionella contamination was associated with the presence of an oil heater (odds ratio [OR] = 2.04, 95% confidence interval [CI] = 1.12 to 3.70), with the sample temperature (OR = 0.26, 95% CI = 0.1 to 0.5), with seasonal operation (OR = 3.23, 95% CI = 1.52 to 6.87), and with the presence of an independent disinfection system (OR = 0.30, 95% CI = 0.15 to 0.62). The same water temperatures, free-chlorine levels, and pHs differently affect the survival of various Legionella spp.     

Anti-inflammatory and antioxidant activity of coumarins designed as potential fluorescent zinc sensors

A series of coumarin analogs, designed and synthesised as potential fluorescent zinc probes were evaluated for their biological activity as anti-inflammatory and antioxidant agents. The effect of the synthesised compounds on inflammation, using the carrageenin-induced rat paw oedema model, was studied. In general, the compounds were found to be potent anti-inflammatory agents (26.5-64%). Compound 5 was found to interact significantly with 1,1-diphenyl-2-picryl-hydrazyl stable free radical (DPPH) whereas the remainder were inactive in this assay. The compounds inhibit in general the soybean lipoxygenase and scavenge superoxide anion radicals. The anti-inflammatory activity seems to be connected with their reducing activity. Their RM values were determined as an expression of their lipophilicity. Theoretical calculations of their lipophilicity as clog P were performed indicating that only a poor relationship exists between their lipophilicity and anti-inflammatory activity.     

Towards understanding of the complex structure of growing yeast populations

In a growing Saccharomyces cerevisiae population, cell size is finely modulated according to both the chronological and genealogical ages. This generates the complex heterogeneous structure typical of budding yeast populations. In recent years, there has been a growing interest in developing mathematical models capable of faithfully describing population dynamics at the single cell level. A multistaged morphologically structured model has been lately proposed based on the population balance theory. The model was able to describe the dynamics of the generation of a heterogeneous growing yeast population starting from a sub-population of daughter unbudded cells. In this work, which aims at validating the model, the simulated experiment was performed by following the release of a homogeneous population of daughter unbudded cells. A biparametric flow cytometric approach allowed us to analyse the time course joint distribution of DNA and protein contents at the single cell level; this gave insights into the coupling between growth and cell cycle progression that generated the final population structure. The comparison between experimental and simulated size distributions revealed a strong agreement for some unexpected features as well. Therefore, the model can be considered as validated and extendable to more complex situations.     

Developmental defects by antisense-mediated inactivation of micro-RNAs 2 and 13 in Drosophila and the identification of putative target genes

Micro-RNAs are a class of small non-coding regulatory RNAs that impair translation by imperfect base pairing to mRNAs. For analysis of their cellular function we injected different miRNA-specific DNA antisense oligonucleotides in Drosophila embryos. In four cases we observed severe interference with normal development, one had a moderate impact and six oligonucleotides did not cause detectable phenotypes. We further used the miR-13a DNA antisense oligonucleotide as a PCR primer on a cDNA library template. In this experimental way we identified nine Drosophila genes, which are characterised by 3' untranslated region motifs that allow imperfect duplex formation with miR-13 or related miRNAs. These genes, which include Sos and Myd88, represent putative targets for miRNA regulation. Mutagenesis of the target motif of two genes followed by transfection in Drosophila Schneider 2 (S2) cells and subsequent reporter gene analysis confirmed the hypothesis that the binding potential of miR-13 is inversely correlated with gene expression.     

Human-mediated introgression of exotic chukar (Alectoris chukar, Galliformes) genes from East Asia into native Mediterranean partridges

Mediterranean red-legged (Alectoris rufa) and rock (Alectoris graeca) partridge populations are affected by genetic pollution. The chukar partridge (Alectoris chukar), a species only partly native to Europe, is the most frequently introgressive taxon detected in the genome of hybrid partridges. Both theoretical (evolutionary) and practical (resources management) matters spur to get insight into the geographic origin of the A. chukar hybridizing swarm. The phenotypic A. rufa populations colonizing the easternmost part of the distribution range of this species, the islands of Elba (Italy) and Corsica (France), were investigated. The analysis of both mitochondrial (mtDNA: Cytochrome-b gene plus Control Region: 2,250 characters) and nuclear (Short Tandem Repeats, STR; Random Amplified Polymorphic DNA, RAPD) genomes of 25 wild (Elba) and 20 captive (Corsica) partridges, disclosed spread introgression of chukar origin also in these populations. All mtDNA haplotypes of Elba and Corsica partridges along with those we obtained from other A. rufa (total, n = 111: Italy, Spain, France) and A. graeca (n = 6, Italy), were compared with the mtDNA haplotypes of chukars (n = 205) sampled in 20 countries. It was found that the A. chukar genes detected in red-legged (n = 43) and rock partridges (n = 4) of Spain, France and Italy as well as in either introduced (Italy) or native (Greece, Turkey) chukars (n = 35) were all from East Asia. Hence, a well-defined geographic origin of the exotic chukar genes polluting the genome of native Mediterranean A. rufa and A. graeca (inter-specific level) as well as A. chukar (intra-specific level), was demonstrated.     

A Drosophila hsp70 gene contains long,antiparallel, coupled open reading frames (LAC ORFs) conserved in homologous loci

A clone isolated from a Drosophila auraria heat-shock cDNA library presents two long, antiparallel, coupled (LAC) open reading frames (ORFs). One strand ORF is 1,929 nucleotides long and exhibits great identity (87.5% at the nucleotide level and 94% at the amino acid level) with the hsp70 gene copies of D. melanogaster, while the second strand ORF, in antiparallel in-frame register arrangement, is 1,839 nucleotides long and exhibits 32% identity with a putative, recently identified, NAD⁺-dependent glutamate dehydrogenase (NAD⁺-GDH). The overlap of the two ORFs is 1,824 nucleotides long. Computational analysis shows that this LAC ORF arrangement is conserved in other hsp70 loci in a wide range of organisms, raising questions about possible evolutionary benefits of such a peculiar genomic organization.Correspondence to: Z.G. Scouras     

Reduction of sialic acid O-acetylation in human colonic mucins in the adenoma-carcinoma sequence

The oligo-O-acetylation of sialic acids found in normal colonic mucins is greatly reduced in colorectal cancer. Mucins prepared from cancer tissue in adenocarcinoma showed this reduction, while normal O-acetylation was detected in resection margin and control cases and total mucin sialic acid content was significantly decreased in cancer vs control samples. A reduction of the O-acetyl transferase activity catalysing the O-acetylation reaction was also found. A series of cultured human colorectal cell lines derived from the same premalignant adenomatous line, and representative of the adenoma-carcinoma sequence were examined and revealed a depletion of oligo-O-acetylation in the original diploid premalignant line, re-expression in a further premalignant line and reduction in malignant mucinous and adenocarcinoma cell lines. Reduction of sialic acid O-acetylation appears as an early event in the process of malignant transformation in human colorectal cancer.     

Focal adhesion kinase is not required for integrin function or viability in Drosophila

The mammalian focal adhesion kinase (FAK) family of non-receptor protein-tyrosine kinases has been implicated in controlling a multitude of cellular responses to the engagement of cell-surface integrins and G-protein-coupled receptors. The high level of sequence conservation between the mammalian proteins and the Drosophila homologue of FAK, Fak56, suggested that it would have similar functions. However, we show here that Drosophila Fak56 is not essential for integrin functions in adhesion, migration or signaling in vivo. Furthermore, animals lacking Fak56 are viable and fertile, demonstrating that Fak56 is not essential for other developmental or physiological functions. Despite this, overexpressed Fak56 is a potent inhibitor of integrins binding to the extracellular matrix, suggesting that Fak56 may play a subtle role in the negative regulation of integrin adhesion.