Influence of Cholesterol and ��-Sitosterol on the Structure of EYPC Bilayers

The influence of cholesterol and β-sitosterol on egg yolk phosphatidylcholine (EYPC) bilayers is compared. Different interactions of these sterols with EYPC bilayers were observed using X-ray diffraction. Cholesterol was miscible with EYPC in the studied concentration range (0-50 mol%), but crystallization of β-sitosterol in EYPC bilayers was observed at X ≥ 41 mol% as detected by X-ray diffraction. Moreover, the repeat distance (d) of the lamellar phase was similar upon addition of the two sterols up to mole fraction 17%, while for X ≥ 17 mol% it became higher in the presence of β-sitosterol compared to cholesterol. SANS data on suspensions of unilamellar vesicles showed that both cholesterol and β-sitosterol similarly increase the EYPC bilayer thickness. Cholesterol in amounts above 33 mol% decreased the interlamellar water layer thickness, probably due to "stiffening" of the bilayer. This effect was not manifested by β-sitosterol, in particular due to the lower solubility of β-sitosterol in EYPC bilayers. Applying the formalism of partial molecular areas, it is shown that the condensing effect of both sterols on the EYPC area at the lipid-water interface is small, if any. The parameters of ESR spectra of spin labels localized in different regions of the EYPC bilayer did not reveal any differences between the effects of cholesterol and β-sitosterol in the range of full miscibility.     

Is it possible to improve homologous recombination in Chlamydomonas reinhardtii?

Targeted modification of the genome has long been an aim of many geneticists and biotechnologists. Gene targeting is a main molecular tool to examine biological effects of genes in a controlled environment. Effective gene targeting depends on the frequency of homologous recombination that is indispensable for the insertion of foreign DNA into a specific sequence of the genome. The main problem associated with the development of an optimal procedure for gene targeting in a particular organism is the variability of homologous recombination (HR) in different species. Chlamydomonas reinhardtii is an attractive model system for the study of many cellular processes and is also an interesting object for the biotechnology industry. In spite of many advantages of this model system, C. reinhardtii does not readily express heterologous genes and does not allow targeted integration of foreign DNA into its genome easily. This paper compares data obtained from several different experiments designed for improving gene targeting in different organisms and reviews the suitability of particular techniques in C. reinhardtii cells. Presented at the International Symposium Biology and Taxonomy of Green Algae V, Smolenice, June 26–29, 2007, Slovakia.     

Altered Astrocytic Swelling in the Cortex of α-Syntrophin-Negative GFAP/EGFP Mice

Brain edema accompanying ischemic or traumatic brain injuries, originates from a disruption of ionic/neurotransmitter homeostasis that leads to accumulation of K⁺ and glutamate in the extracellular space. Their increased uptake, predominantly provided by astrocytes, is associated with water influx via aquaporin-4 (AQP4). As the removal of perivascular AQP4 via the deletion of α-syntrophin was shown to delay edema formation and K⁺ clearance, we aimed to elucidate the impact of α-syntrophin knockout on volume changes in individual astrocytes in situ evoked by pathological stimuli using three dimensional confocal morphometry and changes in the extracellular space volume fraction (α) in situ and in vivo in the mouse cortex employing the real-time iontophoretic method. RT-qPCR profiling was used to reveal possible differences in the expression of ion channels/transporters that participate in maintaining ionic/neurotransmitter homeostasis. To visualize individual astrocytes in mice lacking α-syntrophin we crossbred GFAP/EGFP mice, in which the astrocytes are labeled by the enhanced green fluorescent protein under the human glial fibrillary acidic protein promoter, with α-syntrophin knockout mice. Three-dimensional confocal morphometry revealed that α-syntrophin deletion results in significantly smaller astrocyte swelling when induced by severe hypoosmotic stress, oxygen glucose deprivation (OGD) or 50 mM K⁺. As for the mild stimuli, such as mild hypoosmotic or hyperosmotic stress or 10 mM K⁺, α-syntrophin deletion had no effect on astrocyte swelling. Similarly, evaluation of relative α changes showed a significantly smaller decrease in α-syntrophin knockout mice only during severe pathological conditions, but not during mild stimuli. In summary, the deletion of α-syntrophin markedly alters astrocyte swelling during severe hypoosmotic stress, OGD or high K⁺.     

Characterization of DNA Repair Deficient Strains of Chlamydomonas reinhardtii Generated by Insertional Mutagenesis

While the mechanisms governing DNA damage response and repair are fundamentally conserved, cross-kingdom comparisons indicate that they differ in many aspects due to differences in life-styles and developmental strategies. In photosynthetic organisms these differences have not been fully explored because gene-discovery approaches are mainly based on homology searches with known DDR/DNA repair proteins. Here we performed a forward genetic screen in the green algae Chlamydomonas reinhardtii to identify genes deficient in DDR/DNA repair. We isolated five insertional mutants that were sensitive to various genotoxic insults and two of them exhibited altered efficiency of transgene integration. To identify genomic regions disrupted in these mutants, we established a novel adaptor-ligation strategy for the efficient recovery of the insertion flanking sites. Four mutants harbored deletions that involved known DNA repair factors, DNA Pol zeta, DNA Pol theta, SAE2/COM1, and two neighbouring genes encoding ERCC1 and RAD17. Deletion in the last mutant spanned two Chlamydomonas-specific genes with unknown function, demonstrating the utility of this approach for discovering novel factors involved in genome maintenance.     

Host-guest Interactions in Intercalates Zr(HPO4)2·2C2H5OH and VOPO4·2C2H5OH

Molecular mechanics simulations using Cerius² modelling environment combined with vibrational spectroscopy (IR and Raman) have been used to study the host-guest interactions in zirconium and vanadyl phosphate intercalated with ethanole. The strategy of investigation is based on the comparison of vibrational spectra for the host compound, intercalate and guest species. This comparison confirmed the rigidity of VOPO₄- and Zr(HPO₄)₂-layers during the intercalation and provided us with the basis for the strategy of modelling. Molecular mechanics simulations revealed the structure of intercalates and enabled to analyse the host-guest interaction energy and bonding geometry. The bilayer arrangement of ethanole molecules in the interlayer space with two differently bonded ethanole molecules has been found in both intercalates. The average interaction energy ethanole-layer for two differently bonded ethanole molecules is : 127.5 and 135.7 kcal·mol^-1 in Zr(HPO₄)₂·2C₂H₅OH, respectively 94.0 and 104.4 kcal·mol^-1 in VOPO₄·2C₂H₅OH. The Coulombic contribution to the ethanole-layer interaction energy is predominant in all cases, but the hydrogen bonding contribution is much higher in Zr(HPO₄)₂·2C₂H₅OH than in VOPO₄·2C₂H₅OH. Present results of modelling enabled the interpretation of vibrational spectra and explanation of small changes in positions and shapes of spectral bands, in infrared and Raman spectra, proceeding from the host structure to intercalates.     

Zur Syntaxonomie desDeschampsio flexuosae-Abietetum Husová 1968

Based on new relevés from the area of the Czech Socialist Republic, the associationDeschampsio flexuosae-Abietetum Husová 1968 is divided into two subassociations:D.f.-A. deschampsietosum flexuosae andD.f.-A. calamagrostietosum. The species composition and variability of the association and both subassociations is given in a synoptic table. The soil properties are also added.     

Meiotic findings in human reciprocal 1;3 translocation

Summary A balanced reciprocal translocation between chromosomes 1 and 3 was found in an infertile man. Meiotic analysis showed a ring quadrivalent in diakinesis and complete spermatogenic breakdown after the first meiotic division.     

A study of the formation of antibodies in new-born animals following transfer of nucleoprotein fractions

1. (1)
   Мы выделяли РНП и ДНП из селезенки через 24 и 48 час. после введения 1 дозы антигена Brucella suis взрослому кролику. Нуклеопротеины, перенесенные 3–5-дневным крольчатам, не вызывали у реципиентов появления антител к Brucella suis.     

The effect of ceramic fibers on the immune system

Male Sprague-Dawley rats were treated by intratracheal instillation with 1 mg/animal of refractory ceramic fibers. Intratracheal exposure to ceramic fibers led to significant changes of immune response. Results of proliferative activity of spleen lymphocytes showed significantly decreased proliferative activity of T-cells in response to mitogens phytohemagglutinin and concanavalin A in animals given ceramic fibers in comparison with control rats. Similarly, T-dependent B-cell response to pokeweed mitogen was significantly suppressed. Spontaneous proliferative activity of lymphocytes in non-stimulated spleen cell cultures did not differ in exposed and control rats. No significant changes were found among groups in percentage of phagocytic blood polymorphonuclear leukocytes and percentage of cells with respiratory burst.     

Low variability of the POLG (CAG)n repeat in north Eurasian populations

We investigated the frequency of different repeat-length alleles of the trinucleotide CAG microsatellite repeat in the coding sequence of the nuclear gene for the catalytic subunit of mitochondrial DNA polymerase gamma (POLG) in 12 ethnic groups from northern Eurasia. The population sample consisted of 1,330 individuals from 3 large geographic areas: Europe, Southwest Asia, and Siberia/East Asia. We found that the 10-repeat allele of the POLG gene is the most frequent in all analyzed populations, with a frequency of 88-96%. The heterozygosity level ranges from 22% in Europe to 13.6% in Southwest Asia with the lowest value of 7.4% in Siberia/East Asia. The present study provides evidence of clinal distribution of POLG gene heterozygosity in North Eurasian populations. In general, we found an extremely low variability of the trinucleotide CAG microsatellite repeat, suggesting that purifying selection acts against deleterious alleles, although low mutability of the repeated region cannot be ruled out.