De novo assembly and analysis of the transcriptome of the Siberian wood frog Rana amurensis.

The Siberian wood frog Rana amurensis Boulenger, 1886 is the most hypoxia-tolerant amphibian. It can survive for several months in an almost complete absence of oxygen. Little is known about the mechanisms of this remarkable resilience, in part because studies of amphibian genomes are impeded by their large size. To make the Siberian wood frog more amenable for genetic analysis, we performed transcriptome sequencing and de novo assembly for the R. amurensis brain under hypoxia and normoxia, as well as for the normoxic heart. In order to build a de novo transcriptome assembly of R. amurensis, we utilized 125-bp paired-end reads obtained from the brain under normoxia and hypoxia conditions, and from the heart under normoxia. In the transcriptome assembled from about 100,000,000 reads, 81.5 % of transcripts were annotated as complete, 5.3 % as fragmented, and 13.2 % as missing. We detected 59,078 known transcripts that clustered into 22,251 genes; 11,482 of them were assigned to specific GO categories. Among them, we found 6696 genes involved in protein binding, 3531 genes involved in catalytic activity, and 576 genes associated with transporter activity. A search for genes encoding receptors of the most important neurotransmitters, which may participate in the response to hypoxia, resulted in a set of expressed receptors of dopamine, serotonin, GABA, glutamate, acetylcholine, and norepinephrine. Unexpectedly, no transcripts for histamine receptors were found. The data obtained in this study create a valuable resource for studying the mechanisms of hypoxia tolerance in the Siberian wood frog, as well as for amphibian studies in general.

[Structure of Neuroglobin from Cold-Water Sponge Halisarca dujardinii].

The iron-containing protein neuroglobin (Ngb) involved in the transport of oxygen is generally considered the precursor of all animal globins. In this report, we studied the structure of Ngb of the cold-water sponge Halisarca dujardinii. In sponges, the oldest multicellular organisms, the Ngb gene contains three introns. In contrast to human Ngb, its promoter contains a TATA-box, rather than CG-rich motifs. In sponges, Ngb consists of 169 amino acids showing rather low similarity with its mammalian orthologues. It lacks Glu and Arg residues in positions required for prevention of hypoxia-related apoptosis. Nevertheless, Ngb contains both proximal and distal conserved heme-biding histidines. The primary structure of H. dujardinii neuroglobin predicted by sequencing was confirmed by mass-spectrometry analysis of recombinant Ngb expressed in E. coli. The high level of Ngb expression in sponge tissues suggests its possible involvement in the gas metabolism and presumably in other key metabolic processes in H. dujardinii.

PTPN11 Knockdown Prevents Changes in the Expression of Genes Controlling Cell Cycle, Chemotherapy Resistance, and Oncogene-Induced Senescence in Human Thyroid Cells Overexpressing BRAF V600E Oncogenic Protein.

The MAPK (RAS/BRAF/MEK/ERK) signaling pathway is a kinase cascade involved in the regulation of cell proliferation, differentiation, and survival in response to external stimuli. The V600E mutation in the BRAF gene has been detected in various tumors, resulting in a 500-fold increase in BRAF kinase activity. However, monotherapy with selective BRAF V600E inhibitors often leads to reactivation of MAPK signaling cascade and emergence of drug resistance. Therefore, new targets are being developed for the inhibition of components of the aberrantly activated cascade. It was recently discovered that resistance to BRAF V600E inhibitors may be associated with the activity of the tyrosine phosphatase SHP-2 encoded by the PTPN11 gene. In this paper, we analyzed transcriptional effects of PTPN11 gene knockdown and selective suppression of BRAF V600E in a model of thyroid follicular epithelium. We found that the siRNA-mediated knockdown of PTPN11 after vemurafenib treatment prevented an increase in the expression CCNA1 and NOTCH4 genes involved in the formation of drug resistance of tumors. On the other hand, downregulation of PTPN11 expression blocked the transcriptional activation of genes (p21, p15, p16, RB1, and IGFBP7) involved in cell cycle regulation and oncogene-induced senescence in response to BRAF V600E expression. Therefore, it can be assumed that SHP-2 participates not only in emergence of drug resistance in cancer cells, but also in oncogene-induced cell senescence.

[ULTRASTRUCTURE OF PARENCHYMA IN THE SYNCYTIAL DIGESTIVE SYSTEM IN TURBELLARIA Convoluta convoluta (Acoela].

The paper presents data on the ultrastructure of parenchyma that is involved in the digestion in turbellaria Convoluta convoluta (n = 15). Unusual connections between the nuclear envelope, endoplasmic reticulum and plasma membrane of parenchymal cells were found for the first time, which may indicate the origin of these cell structures. The double trophic role of zooxanthellae in the organism of Convoluta is described.

[Electron microscopic study of the cytopathogenic action of meningococci in a continuous human amnion cell culture]. / Elektronno-mikroskopicheskoe izuchenie tsitopatogennogo deistviia meningokokkov na perevivaemuiu kul'turu kletok amniona cheloveka.

The electron-microscopic study of the interaction of meningococci with continuous human amnion cell culture F1 has revealed that this process comprises 3 stages. The study has shown that, following the adhesion of meningococci to the surface of cells F1, these cells are invaded by individual coccal forms of meningococci. In response to infection vacuoles appear in the cytoplasm of the cells. Meningococci are either phagocytosed inside these vacuoles, or their release into the intercellular space and the death of the infected by meningococci are observed. When the cells are infected by cytopathogenic strains, the infectious process results in the appearance of degenerative changes in the cells.

[Interactions between Neisseria meningitidis and cell cultures (data of cytomorphologic studies)]. / Vzaimodeistvie Neisseria meningitidis s kul'turoi kletok po tsitomorfologicheskim nabliudeniiam.

The present work shows that the cytopathogenic action of N. meningitidis on continuous human amniotic epithelial cell culture FL begins from their active adhesion and subsequent invasion. The degree of the manifestation of the cytopathic effect depends on the capacity of the infective agent for adhesion and invasion and on its biological properties, as well as on the initial state of the cells. The infection of the cells is accompanied by disturbances in their mitotic activity together with the lesions of their chromosomal apparatus. The cells die either in the state of degenerative mitosis, or as the result of the rupture of the cytoplasm in massively invaded cells. The response of the cells to the invasion of faintly cytopathogenic and noncytopathogenic strains takes the form of nonprofessional phagocytosis.

[Characteristics of nonpathogenic Neisseria and Branhamella catarrhalis based on cytopathogenicity]. / Kharakteristika nepatogennykh neisserii i Branhamella catarrhalis po tsitopatogennosti.

The study of the action of 12 Neisseria species belonging to 112 strains, 6 B. catarrhalis strains and 202 meningococcal strains on the culture of continuous cell line F1 (human amniotic cells) has revealed that nonpathogenic Neisseria are essentially weaker than meningococci in their pathogenicity (expressed in terms of CPD50). Among nonpathogenic Neisseria highly cytopathogenic strains occur in 13.9% of cases, which gives grounds for considering them opportunistic bacteria. Sharply pronounced correlation between the adhesive and pathogenic properties of Neisseria has been observed. The cytopathogenic action of Neisseria is accompanied by the lesion of the chromosomal apparatus of mitotic infected cells.

[The effect of diphtheria toxin on proliferation of a transplantable culture of human amnion cells]. / Vliianie difteriinogo toksina na proliferatsiiu perevivaemoi kul'tury amnioticheskikh kletok cheloveka

Cytoxic effect (CTE) of diphtheria toxin (DT) is preceded by disturbances of proliferation process, this being expressed in reduction of the proliferation index (PI) and of mitotic index (MI) before the appearance of the CTE. The effect of the toxin on the proliferation and mitoses of the cell culture is expressed after the incubation period of about 3 hours. The extent and the rate of reduction of the indices directly depended on the dose of the toxin whose low doses at the early periods stimulated the proliferation. The maximum of all the manifestations of the toxin action coincided with the period of increased proliferation in control. On the basis of investigations carried out a hypothesis was put forward that the CTE of diphtheria toxin developed with the entrance of the cells into mitosis.