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Identification of miniature plasmids in psychrophilic Arctic bacteria of the genus Variovorax
Get PDFThe Svalbard archipelago (Spitsbergen Island) is the northernmost landmass in the European Arctic and has a variety of small- and medium-sized glaciers. The plasmidome of eleven psychrophilic strains of Variovorax spp. isolated from the ice surface of Hans and Werenskiold Glaciers of Spitsbergen Island, was defined. This analysis revealed the presence of six plasmids whose nucleotide sequences have been determined. Four of them, exhibiting high reciprocal sequence similarity, possess unique structures, since their genomes lack any recognized genes. These miniature replicons, not exceeding 1 kb in size, include pHW69V1 (746 bp), which is the smallest autonomous replicon so far identified in free-living bacteria. The miniature plasmids share no similarity with known sequences present in the databases. In silico and experimental analyses identified conserved DNA regions essential for the initiation of replication of these replicons
On the ability of CuAβ1-x peptides to form ternary complexes: Neurotransmitter glutamate is a competitor while not a ternary partner
Get PDFIn the light of conflicting reports on the ability of copper(II) complexes of amyloid beta (Aβ) peptides to form ternary complexes with small molecules co-present in the biological milieu, we performed a study of coordination equilibria in the system containing Cu(II) ions, the Aβ1-16 peptide, glutamic acid and 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid, HEPES) buffer. Using potentiometry, isothermal titration calorimetry (ITC), UV-visible spectroscopy and EPR, we concluded that glutamic acid was not able to form such a ternary complex, but can efficiently compete for the Cu(II) ion with the Aβ peptide at Glu concentrations relevant for the synaptic cleft. We also found that the literature constants for Cu(II) complexes with Glu were overestimated, but this effect was partially compensated by the formation of a ternary Cu(Glu)(HEPES) complex. Our results indicate that small molecules co-present with Cu(II) ions and Aβ peptides in the synaptic cleft are not very likely to enhance Cu(II)/Aβ interactions, but instead should be considered as a Cu(II) buffering system that may help prevent these interactions and participate in Cu(II) clearance from the synaptic cleft
A European Spectrum of Pharmacogenomic Biomarkers: Implications for Clinical Pharmacogenomics
Get PDFPharmacogenomics aims to correlate inter-individual differences of drug efficacy and/or toxicity with the underlying genetic composition, particularly in genes encoding for protein factors and enzymes involved in drug metabolism and transport. In several European populations, particularly in countries with lower income, information related to the prevalence of pharmacogenomic biomarkers is incomplete or lacking. Here, we have implemented the microattribution approach to assess the pharmacogenomic biomarkers allelic spectrum in 18 European populations, mostly from developing European countries, by analyzing 1,931 pharmacogenomics biomarkers in 231 genes. Our data show significant inter-population pharmacogenomic biomarker allele frequency differences, particularly in 7 clinically actionable pharmacogenomic biomarkers in 7 European populations, affecting drug efficacy and/or toxicity of 51 medication treatment modalities. These data also reflect on the differences observed in the prevalence of high-risk genotypes in these populations, as far as common markers in the CYP2C9, CYP2C19, CYP3A5, VKORC1, SLCO1B1 and TPMT pharmacogenes are concerned. Also, our data demonstrate notable differences in predicted genotype-based warfarin dosing among these populations. Our findings can be exploited not only to develop guidelines for medical prioritization, but most importantly to facilitate integration of pharmacogenomics and to support pre-emptive pharmacogenomic testing. This may subsequently contribute towards significant cost-savings in the overall healthcare expenditure in the participating countries, where pharmacogenomics implementation proves to be cost-effective
Colonization of germ-free mice with a mixture of three lactobacillus strains enhances the integrity of gut mucosa and ameliorates allergic sensitization
Get PDFIncreasing numbers of clinical trials and animal experiments have shown that probiotic bacteria are promising tools for allergy prevention. Here, we analyzed the immunomodulatory properties of three selected lactobacillus strains and the impact of their mixture on allergic sensitization to Bet v 1 using a gnotobiotic mouse model. We showed that Lactobacillus (L.) rhamnosus LOCK0900, L. rhamnosus LOCK0908 and L. casei LOCK0919 are recognized via Toll-like receptor 2 (TLR2) and nucleotide-binding oligomerization domain-containing protein 2 (NOD2) receptors and stimulate bone marrow-derived dendritic cells to produce cytokines in species- and strain-dependent manners. Colonization of germ-free (GF) mice with a mixture of all three strains (Lmix) improved the intestinal barrier by strengthening the apical junctional complexes of enterocytes and restoring the structures of microfilaments extending into the terminal web. Mice colonized with Lmix and sensitized to the Bet v 1 allergen showed significantly lower levels of allergen-specific IgE, IgG1 and IgG2a and an elevated total IgA level in the sera and intestinal lavages as well as an increased transforming growth factor (TGF)-β level compared with the sensitized GF mice. Splenocytes and mesenteric lymph node cells from the Lmix-colonized mice showed the significant upregulation of TGF-β after in vitro stimulation with Bet v 1. Our results show that Lmix colonization improved the gut epithelial barrier and reduced allergic sensitization to Bet v 1. Furthermore, these findings were accompanied by the increased production of circulating and secretory IgA and the regulatory cytokine TGF-β. Thus, this mixture of three lactobacillus strains shows potential for use in the prevention of increased gut permeability and the onset of allergies in humans
Draft Genome Sequence of Shewanella baltica M1 Isolated from Brackish Surface Water of the Gulf of Gdańsk
Get PDFHere, we present the 5.168-Mbp draft genome sequence of Shewanella baltica M1, the first Shewanella strain from the Gulf of Gdańsk to have its genome sequenced and annotated. The availability of the genome sequence of strain M1 will promote further global analyses of bacterial stress responses in the unique Gulf of Gdan´ sk ecosystem
Phosphatidylinositol-3-phosphate regulates response of cells to proteotoxic stress
Get PDFHuman Nedd4 ubiquitin ligase, or its variants, inhibit yeast cell growth by disturbing the actin cytoskeleton organization and dynamics, and lead to an increase in levels of ubiquitinated proteins. In a screen for multicopy suppressors which rescue growth of yeast cells producing Nedd4 ligase with an inactive WW4 domain (Nedd4w4), we identified a fragment of ATG2 gene encoding part of the Atg2 core autophagy protein. Expression of the Atg2-C1 fragment (aa 1074-1447) improved growth, actin cytoskeleton organization, but did not significantly change the levels of ubiquitinated proteins in these cells. The GFP-Atg2-C1 protein in Nedd4w4-producing cells primarily localized to a single defined structure adjacent to the vacuole, surrounded by an actin filament ring, containing Hsp42 and Hsp104 chaperones. This localization was not affected in several atg deletion mutants, suggesting that it might be distinct from the phagophore assembly site (PAS). However, deletion of ATG18 encoding a phosphatidylinositol-3-phosphate (PI3P)-binding protein affected the morphology of the GFP-Atg2-C1 structure while deletion of ATG14 encoding a subunit of PI3 kinase suppressed toxicity of Nedd4w4 independently of GFP-Atg2-C1. Further analysis of the Atg2-C1 revealed that it contains an APT1 domain of previously uncharacterized function. Most importantly, we showed that this domain is able to bind phosphatidylinositol phosphates, especially PI3P, which is abundant in the PAS and endosomes. Together our results suggest that human Nedd4 ubiquitinates proteins in yeast and causes proteotoxic stress and, with some Atg proteins, leads to formation of a perivacuolar structure, which may be involved in sequestration, aggregation or degradation of proteins
Effect of surfactants on surface activity and rheological properties of type I collagen at air/water interface
Get PDFWe describe the effect of three synthetic surfactants (anionic - sodium dodecyl sulfate (SDS), cationic - cetyltrimethylammonium bromide (CTAB) and nonionic - Triton X-100 (TX-100)) on surface properties of the type I calf skin collagen at the air/water interface in acidic solutions (pH 1.8). The protein concentration was fixed at 5×10-6molL-1 and the surfactant concentration was varied in the range 5×10-6molL-1-1×10-4molL-1, producing the protein/surfactant mixtures with molar ratios of 1:1, 1:2, 1:5, 1:10 and 1:20. An Axisymmetric Drop Shape Analysis (ADSA) method was used to determine the dynamic surface tension and surface dilatational moduli of the mixed adsorption layers. Two spectroscopic techniques: UV-vis spectroscopy and fluorimetry allowed us to determine the effect of the surfactants on the protein structure. The thermodynamic characteristic of the mixtures was studied using isothermal titration calorimetry (ITC) and differential scanning calorimetry (DSC). Modification of the collagen structure by SDS at low surfactant/protein ratios has a positive effect on the mixture's surface activity with only minor deterioration of the rheological properties of the adsorbed layers. The collagen/CTAB mixtures do not show that pronounced improvement in surface activity, while rheological properties are significantly deteriorated. The mixtures with non-ionic TX-100 do not show any synergistic effects in surface activity
Biodiversity of bacteriophages: morphological and biological properties of a large group of phages isolated from urban sewage
Get PDFA large scale analysis presented in this article focuses on biological and physiological variety of bacteriophages. A collection of 83 bacteriophages, isolated from urban sewage and able to propagate in cells of different bacterial hosts, has been obtained (60 infecting Escherichia coli, 10 infecting Pseudomonas aeruginosa, 4 infecting Salmonella enterica, 3 infecting Staphylococcus sciuri, and 6 infecting Enterococcus faecalis). High biological diversity of the collection is indicated by its characteristics, both morphological (electron microscopic analyses) and biological (host range, plaque size and morphology, growth at various temperatures, thermal inactivation, sensitivity to low and high pH, sensitivity to osmotic stress, survivability upon treatment with organic solvents and detergents), and further supported by hierarchical cluster analysis. By the end of the research no larger collection of phages from a single environmental source investigated by these means had been found. The finding was confirmed by whole genome analysis of 7 selected bacteriophages. Moreover, particular bacteriophages revealed unusual biological features, like the ability to form plaques at low temperature (4 °C), resist high temperature (62 °C or 95 °C) or survive in the presence of an organic solvents (ethanol, acetone, DMSO, chloroform) or detergent (SDS, CTAB, sarkosyl) making them potentially interesting in the context of biotechnological applications
Cytoplasmic RNA decay pathways - enzymes and mechanisms
Get PDFRNA decay plays a crucial role in post-transcriptional regulation of gene expression. Work conducted over the last decades has defined the major mRNA decay pathways, as well as enzymes and their cofactors responsible for these processes. In contrast, our knowledge of the mechanisms of degradation of non-protein coding RNA species is more fragmentary. This review is focused on the cytoplasmic pathways of mRNA and ncRNA degradation in eukaryotes. The major 3’ to 5’ and 5’ to 3’ mRNA decay pathways are described with emphasis on the mechanisms of their activation by the deprotection of RNA ends. More recently discovered 3’-end modifications such as uridylation, and their relevance to cytoplasmic mRNA decay in various model organisms, are also discussed. Finally, we provide up-to-date findings concerning various pathways of non-coding RNA decay in the cytoplasm