Phylogenetic analysis and molecular genetic characteristics of West Nile virus lineage 2 isolates circulating in the Russian Federation.

Since its initial detection in Africa, the West Nile virus has disseminated widely across all continents, becoming endemic in numerous countries, including the Russian Federation. A substantial expansion of the West Nile virus range was observed in the European part of the Russian territory in 1999. In light of this epidemiological trend, research endeavours focusing on monitoring West Nile virus circulation activity in endemic regions of the country have gained paramount significance. A substantial dataset has been accrued from 2007 onwards regarding genomic variability and dissemination dynamics across the country throughout the entire monitoring period for the West Nile fever pathogen. The objective of this study was to characterise West Nile virus isolates that have been circulating in the Russian Federation and identify their molecular and genetic characteristics. A phylogenetic analysis of 55 complete genome sequences revealed that the West Nile virus population within the Russian Federation is genetically heterogeneous and is represented by four major clades. One of these clades is currently exhibiting extensive spread into new regions of the country.

1,8-Bis(dimethylamino)naphthyl-2-ketimines: Inside vs outside protonation.

The structure and protonation behaviour of four ortho-arylketimines of 1,8-bis(dimethylamonio)naphthalene with a different number of methoxy groups in an aromatic substituent were investigated in solution by NMR (acetone, DMSO, MeCN), in solid state by X-ray analysis and in the gas phase by DFT calculations. Both mono- and diprotonated species were considered. It has been shown that E-isomers of neutral imines can be stabilised by an intramolecular C=N-H···OMe hydrogen bond with a neighbouring methoxy group. Electron-donating OMe groups dramatically increase the basicity of the imino nitrogen, forcing the latter to abstract a proton from the proton sponge moiety in monoprotonated forms. The participation of the out-inverted and protonated 1-NMe2 group in the Me2N-H···NH=C hydrogen bond is experimentally demonstrated. It was shown that the number and position of OMe groups in the aromatic substituents strongly affects the rate of the internal hindered rotation of the NH2 + fragment in dications.

[Structure and evolution of junctions between inverted repeat and small single copy regions of chloroplast genome in non-core Caryophyllales].

The structure of junction between inverted repeat (IR) and small single copy (SSC) regions of chloroplast genome in the representatives of non-core Caryophyllales is investigated in this work. It was found that for two families - Polygonaceae and Plumbaginaceae - the extension of inverted region is characteristic. This extension is due to the duplication the part of ycf1 gene that is partly located in the small single copy region in plants with typical structure of IR/SSC junctions. Comparison of the position of IR/SSC junctions in different species of Polygonaceae has shown that their exact position is not correlated with the affinity of these species inferred from molecular and morphological data. Possible mechanisms leading to the change in position of IR/SSC junctions observed in this work are discussed.

From birth to christening.

A brief history of comparative studies of nucleic acids for systematic purposes is given. These studies were initiated by a group of Moscow State University scientists headed by A. N. Belozersky. Based mostly on comparative DNA studies, some main dogmas of a new branch of systematics were gradually developed. In Russia, this new branch of systematics is called "genosystematics". Some of the main results obtained by genosystematics since its birth (1957) and up to its "christening" (1974) are described.

Phylogeny of flowering plants by the chloroplast genome sequences: in search of a "lucky gene".

One of the most complicated remaining problems of molecular-phylogenetic analysis is choosing an appropriate genome region. In an ideal case, such a region should have two specific properties: (i) results of analysis using this region should be similar to the results of multigene analysis using the maximal number of regions; (ii) this region should be arranged compactly and be significantly shorter than the multigene set. The second condition is necessary to facilitate sequencing and extension of taxons under analysis, the number of which is also crucial for molecular phylogenetic analysis. Such regions have been revealed for some groups of animals and have been designated as "lucky genes". We have carried out a computational experiment on analysis of 41 complete chloroplast genomes of flowering plants aimed at searching for a "lucky gene" for reconstruction of their phylogeny. It is shown that the phylogenetic tree inferred from a combination of translated nucleotide sequences of genes encoding subunits of plastid RNA polymerase is closest to the tree constructed using all protein coding sites of the chloroplast genome. The only node for which a contradiction is observed is unstable according to the different type analyses. For all the other genes or their combinations, the coincidence is significantly worse. The RNA polymerase genes are compactly arranged in the genome and are fourfold shorter than the total length of protein coding genes used for phylogenetic analysis. The combination of all necessary features makes this group of genes main candidates for the role of "lucky gene" in studying phylogeny of flowering plants.

[Genosystematics: from E. Chargaff and A. N. Belozersky up to date].

A review of history of genosystematics (macromolecular systematics) from E. Chargaff and A. N. Belozersky up to date. The role of A.N. Belozersky and his collaborators in the development of this new branch of systematics is analyzed. Genosystematics was the source of valuable information clarifying some aspects of biological evolution. Its methods were successfully employed in microorganisms--(e.g., discovery of archaebacteria) and in eucaryote systematics (origin of plastids, falcification of "molecular clock" hypothesis, substantial changes in higher plants phylogenetics, etc.). However, attempts to employ some fragmentary and unreliable data obtained by genosystematics for modifying the existing phylogenetic schemes and systems of organisms failed. Nowadays genosystematics is like a newborn child suffering from children's diseases well-known to "classical" systematics. It is rather far from final conclusions describing the evolution of genotypes. Some of its recent achievments, e.g., elaboration of the concept of PhyloCode, allow to believe that this science is able to suggest revolutionary changes in Linnean systematics.

[On interrelation between genetic systematics and genomics]. / O vzaimosviazi genosistematiki i genomiki.

Review papers describing recent achievements of genomics usually do not pay attention to direct interrelation between genomics and genosystematics (DNA-systematics). Genomics on general is based in complete DNA sequencing of genomes. Initial aim of genosystematics was the same. Absence of historical perspective in review papers devoted to genomics decreases its value. In case it is done deliberately it becomes the problem of scientific ethics. It is postulated that genomics is a natural stage of genosystematics (DNA-systematics) development. Russian scientists were among the founders of these branches of biology.

[Genomics and genosystematics]. / Genomika i genosistematika.

A survey of publications dealing with comparative analysis of genomes shows that modern genomics has naturally evolved from gene systematics--an area whose formation and development was greatly influenced by the scientific school of A.N. Belozersky.

[Novel triterpene glycosides from the sponge Ulosa sp]. / Novye triterpenovye glikozidy iz gubki Ulosa sp.

New triterpene glycosides, ulososides C, (20S,22S,23R,24S)-3 beta,22, 23-trihydroxy-3-O-(beta-D-glucopyranosyl)-32-nor-24-methyllanost- 8(9)-ene-30-oic acid, D, (20S,22S,23R,24S)-3 beta,22, 23-trihydroxy-3-O-(beta-D-N-acetyl-glucosaminopyranosyl)-32-nor- 24-methyllanost-8(9)-ene-30-oic acid, and E, (20S,22S,23R,24S)-3 beta,22, 23-trihydroxy-3-O-(beta-D-glucuronopyranosyl-(1-->2)-alpha-D- arabinopyranosyl-32-nor-24-methyllanost-8(9)-ene-30-oic acid, were isolated from an Ulosa sp. sponge. Their structures were determined by spectral methods and chemical transformations. Specific features of their structures are discussed.

Triterpene glycosides from the far eastern sea cucumber Pentamera calcigera II: disulfated glycosides.

Three new triterpene glycosides, calcigerosides D(1) (1), D(2) (2), and E (3), have been isolated from the sea cucumber Pentamera calcigera. Their structures have been deduced from extensive spectral analysis (NMR and MS) and chemical evidence. All the compounds are disulfated pentaosides differing in aglycon structure and position of sulfate group, which were determined by the measurement of NT(1) values in the cases of glycosides 1 and 2. Glycoside 1 is a nonholostane derivative, that is, it lacks an 18(20)-lactone, which is very rare among the sea cucumber glycosides.

Triterpene glycosides from the Far-Eastern sea cucumber Pentamera calcigera. 1. Monosulfated glycosides and cytotoxicity of their unsulfated derivatives.

Three new monosulfated triterpene glycosides, calcigerosides B (2), C(1) (3), and C(2) (4), along with the known cucumarioside G(2) (1), have been isolated from the sea cucumber Pentamera calcigera. Their structures have been deduced from extensive spectral analysis (NMR and MS) and chemical evidence. Compounds 2-4 present a novel pentasacharide chain never reported before in sea cucumber triterpene glycosides. The desulfated derivatives of calcigerosides B, C(1), and C(2) (5, 7, and 9, respectively) showed moderate cytotoxicity (IC(50) = 5 microg/mL) against a selection of four human and mouse tumor cell lines.

Molecular data from the chloroplast rpoC1 gene suggest a deep and distinct dichotomy of contemporary spermatophytes into two monophyla: gymnosperms (including Gnetales) and angiosperms.

Partial sequences of the rpoC1 gene from two species of angiosperms and three species of gymnosperms (8330 base pairs) were determined and compared. The data obtained support the hypothesis that angiosperms and gymnosperms are monophyletic and none of the recent groups of the latter is sister to angiosperms.

Sequences of rDNA internal transcribed spacers from the chloroplast DNA of 26 bryophytes: properties and phylogenetic utility.

We determined the sequence of the region of the chloroplast DNA inverted repeat spanning from the 3'-terminus of the 23S rRNA gene to the 5'-terminus of the tRNA[Arg](ACG) gene (about 700 bp) from 25 bryophytes and from the charophycean alga Chara australis. Phylogenetic analysis of these sequences using the neighbor-joining method suggests an early dichotomy of bryophytes and their paraphyly relative to the tracheophyte lineage. A monophyly of liverworts (Marchantiidae plus Jungermanniidae), a deep divergence of Metzgeriales among Jungermanniidae and a close affinity of the two subclasses of mosses, Sphagnidae and Andreaeidae, are evident. The branching pattern observed is consistent with the phylogenetic distribution of several prominent indels observed in the alignment.

Aortic endothelial cells regulate proliferation of human monocytes in vitro via a mechanism synergistic with macrophage colony-stimulating factor. Convergence at the cyclin E/p27(Kip1) regulatory checkpoint.

Monocyte-derived macrophages (Mphis) are pivotal participants in the pathogenesis of atherosclerosis. Evidence from both animal and human plaques indicates that local proliferation may contribute to accumulation of lesion Mphis, and the major Mphi growth factor, macrophage colony stimulating factor (MCSF), is present in atherosclerotic plaques. However, most in vitro studies have failed to demonstrate that human monocytes/Mphis possess significant proliferative capacity. We now report that, although human monocytes cultured in isolation showed only limited MCSF-induced proliferation, monocytes cocultured with aortic endothelial cells at identical MCSF concentrations underwent enhanced (up to 40-fold) and prolonged (21 d) proliferation. In contrast with monocytes in isolation, this was optimal at low seeding densities, required endothelial cell contact, and could not be reproduced by coculture with smooth muscle cells. Intimal Mphi isolated from human aortas likewise showed endothelial cell contact-dependent, MCSF-induced proliferation. Consistent with a two-signal mechanism governing Mphi proliferation, the cell cycle regulatory protein, cyclin E, was rapidly upregulated by endothelial cell contact in an MCSFindependent fashion, but MCSF was required for successful downregulation of the cell cycle inhibitory protein p27(Kip1) before cell cycling. Thus endothelial cells and MCSF differentially and synergistically regulate two Mphi genes critical for progression through the cell cycle.

[Molecular biological research on the origin of the angiosperms]. / Molekuliarno-biologicheskie issledovaniia proiskhozhdeniia pokrytosemennykh rastenii.

Nucleotide sequence of 23S-5S chloroplast rDNA spacer region including 4.5S rRNA gene of several dozens of seed plants was determined. The data obtained were used to construct phylogenetic trees and to compare them with the analogous data from literature. Topologies of trees constructed for various types of macromolecules and by different methods demonstrate obvious similarities although they are not identical. Some clue stages of seed plants evolution still remain obscure. Critical analysis of all the available information allows to come to several more or less definite conclusions. All the data say that angiosperms are a monophyletic group which diversified far before their fossils are definitely registered, i.e., before lower Cretaceous. Ancestral angiosperms were not genealogicaly related to modern woody Magnoliales but were represented by "paleoherbs", i. e. herbaceous and semiherbaceous magnoliids and monocots. Monocots originated at the earliest stages of angiosperms evolution and are not, probably, monophyletic. Woody Magnoliales and eudicots with tricolpate pollen seem to appear later in evolution. The conclusion that Gnetales is a sister group to angiosperms does not find enough support in molecular studies. Summing up, it looks as if a long period of existence of angiophytes preceded the pre-Cretaceous angiosperms irradiation. This line of development originated simultaneously with phylogenetic lineages of modern gymnosperms.

Secretion of a novel T-lymphocyte cytokine possessing both chemotactic and growth factor activity by serotonin-stimulated human aortic endothelial cells.

The development of an extralymphatic T-lymphocyte focus of inflammation requires chemoattractant-induced cell migration and growth factor-induced cell proliferation. In a previous study, we identified a novel 13- to 15-kDa T-lymphocyte-specific chemotactic cytokine, endothelial cell-derived lymphocyte chemoattractant activity (ED-LCA), secreted by serotonin-stimulated human aortic endothelial cells. Based on its physicochemical and functional characteristics and antibody inhibition studies, ED-LCA is distinct from previously identified endothelial cell-derived IL-1, IL-6, and IL-8. Because of the association between T-lymphocyte chemotactic and growth factor activity, in the current study, we investigated the effect of ED-LCA on T cell growth by assessing its capacity to induce markers of the passage of T cells from the resting (G0) state into the G1 phase of the cell cycle, such as receptors for IL-2 (IL-2R) and transferrin (TFR), and class II major histocompatibility complex antigens (HLA-DR). Incubation of G0 freshly isolated human T lymphocytes for 48 h with chromatographically resolved, partially purified ED-LCA resulted in a threefold increase in expression of IL-2R, a threefold increase in TFR, and a twofold increase in HLA-DR. Double antibody labeling demonstrated that IL-2R was induced in both CD4+ and CD8+ T cell subsets. Although incubation of human T cells with ED-LCA alone did not induce DNA synthesis, addition of exogenous IL-2 to T cells pulsed with ED-LCA for 24 h caused an increase in DNA synthesis with a stimulation index of 3.5. By up-regulating functional cell surface receptors for IL-2 on T lymphocytes and priming them to respond to exogenous IL-2, ED-LCA is a competence growth factor. By virtue of its effect on T cells, as a chemotactic and competence factor, this endothelial cell-derived mitoattractant could participate with other T-cell growth factors like IL-2 in the generation of an extralymphatic T-lymphocyte inflammatory response.

[The regional morphological characteristics of the endothelium of the human thoracic aorta in perfusion fixation]. / Regionarnye morfologicheskie osobennosti éndoteliia grudnoi chasti aorty cheloveka pri perfuzionnoi fiksatsii.

The ultramicroscopic organization and the endotheliocyte surface relief in ventral portions of the thoracic part of human aorta and in zones of division of blood flow were studied under conditions of early post mortem examinations and perfusion fixation of corpses of 14 humans dead from accidental causes. Zones of entrance into the intercostal aortas are compared with the straight portions of the aorta and are found to be characterized, as compared with the latters, by polymorphism of the endothelium, higher adhesiveness of its surface to blood elements, as well as by the presence of intravitally de-endothelialized portions localized on the ridge of the intimal valve.

[Signal-conducting and low molecular weight GTP-binding proteins from the lung and endothelium: localization in membranes and cytosol, interaction with F-actin]. / Signal-provodiashchie i nizkomolekuliarnye GTP-sviazyvaiushchie belki legkikh i éndoteliia: lokalizatsiia v membranakh i tsitozole, vzaimodeistvie s F-aktinom.

The following proteins have been identified in mammalian lung and endothelium, using [32P]ADP-ribosylation by bacterial ADP-ribosyltransferase, immuno- and [alpha-32P]GTP-blottings: 41 kDa Gi1 alpha, 40 kDa Gi2 alpha, 41 kDa Gi3 alpha, 40 kDa and 45 kDa subunits of GS alpha, 36 kDa beta 1 and 35 kDa beta 2 subunits of signal-transmitting GTP-binding proteins (G-proteins), the 19-26 kDa low molecular weight GTP-binding proteins (SMG-proteins) ras, rho, rac, G25K (Gp), as well as ARF and SMG proteins binding with a high affinity to [alpha-32P]GTP. These G- and SMG-proteins are contained in various proportions in membrane and cytosol fractions of lung and endothelium cells. Subunits Gi2 alpha and GS alpha (but not beta 1 or SMG-proteins) my partially (approximately 1%) dissociate from the membrane by the action of the GTP analogs GTP[S] or Gpp(NH)p in the presence of magnesium ions. Extraction with low ionic strength buffer solutions in the presence of EDTA is accompanied by the release of G-actin sensitive to whooping cough toxin Gi2 alpha and beta i subunits. The functionally coupled into a alpha beta gamma heterodimer Gi-protein subunits (predominantly Gi2 alpha and beta i) present in the cytosol fraction as well as the SMG-proteins revealed by [alpha-32P]GTP-blotting (but not the SMG-proteins sensitive to the botulinic C3 exoenzyme, rho/rac, or ARF, may interact with F-actin. Approximately 20% of these proteins are associated with the Triton X-100 insoluble (cytoskeletal) fraction of the endothelium. A conclusion is drawn that interactions of G- and SMG-proteins with actin filaments may be the reason for the formation of "multidisperse" structure in a cell.

Phenotype related changes of intimal smooth muscle cells from human aorta in primary culture.

To study the functional characteristics of smooth muscle cell (SMC) phenotypes, we have investigated myosin expression, cell proliferation, collagen production and low-density lipoprotein (LDL) receptor activity in intimal SMCs of normal human aorta during their growth in primary culture. By staining with rabbit antibodies to smooth muscle myosin (ASMM) 3 cell types could be distinguished in culture: homogeneously stained cells, cells with discontinuous myosin fibrils and myosin-negative cells. The ratio of cell types greatly changed with culture growth: on days 5, 7 and 14 it was 82:1:17%, 70:5:25% and 10:30:60%, respectively. After 5-6 days of culture intimal SMCs began to proliferate and DNA-synthesizing nuclei were seen 1.5-4.3 times more frequently in myosin-negative cells than in cells with homogeneous myosin distribution. At that time the number of cells reacted with monoclonal antibody (MAb) to an epitope shared collagen types I and III started to increase. By double immunofluorescence staining it was shown that the cultured cells containing both ASMM and MAb markers were found 2.0-4.8 times more rarely than MAb-positive staining in myosin-negative cells. During the first 5 days in culture LDL binding and uptake were diminished in intimal cells with intercellular lipid inclusions independently of their myosin staining pattern, but their activity increased with culture growth. Thus, SMCs from human aortic intima change their phenotype on days 6 and 7 in primary culture as manifested by alteration of myosin expression, increased cell proliferation, collagen production and LDL receptor activity. Changes in myosin expression, however, are not an essential prerequisite for cell proliferation and collagen production.