[PAIRWISE CROSS-SPECIES TRANSCRIPTOME ANALYSIS OF POLYPLOIDY-ASSOCIATED EXPRESSION CHANGES OF DEVELOPMENTAL GENE MODULES].

Design and development of highly sensitive method bioinformatics are important for investigation of casual relationships between epigenetic changes and gene activity. Cell polyploidy may trigger such changes. However, maintaining the balance of gene dosage, polyploidy may provide only a rather weak effect on their expression. Currently, there is no comprehensive and concordant data in regard to ploidy-associated transcriptomic changes. To find out how polypoidy affects gene activity, we have developed an integrative bioinformatic method of pairwise cross-species transcriptome analysis of mammalian tissues with various polyploidy degrees. The main benefit of this approach is its ability to separate species- and tissue-specific noises of evolutionary conserved effects. We demonstrat the application of the method for the analysis of gene modules and protein interactions networks coordinating programs of development, differentiation and pluripotency. The analysis was performed with transcriptomes of polyploid and diploid organs (human and mouse heart and liver). Our data indicate that ploidy-induced genes enrich Gene Ontology (GO) biological processes and KEGG pathways related to development, morphogenesis and stem cells biology (including Hippo, Pi3K, WNT, Hedgehog and TGF-ß pathways) with higher degree than ploidy-inhibited genes. Thas, our data are the first to show that polyploidy may induce and coordinate developmental modules.

[The relation of the phenomenon of nuclear achromasia to cell activation for proliferation]. / Sviaz' fenomena iadernoi akhromazii s aktivatsiei kletok k proliferatsii.

As it has been shown earlier, the cells of experimental tumours as well as some normal cells of the thymus, spleen and testes show the phenomenon of nuclear achromasia (low affinity for ionic dyes) in cytological preparations. Possible connection between the phenomenon and activation for proliferation was the subject of the study presented. A reliable increase in the number of cells displaying the nuclear achromasia when staining with methylene blue after treatment with phytohaemagglutinin for 30 min as well as appearance of such cells in rat liver regenerates 1 h after partial hepatectomy was substantiated to be a result of the cell activation as tested by the enhanced acridine orange intercalation into their DNA.

[Effect of single-stranded breaks on the ultrastructural organization and cytochemistry of the chromatin in tumor cells]. / Vliianie odnonitevykh razryvov na ul'trastrukturnuiu organizatsiiu i tsitokhimiiu khromatina opukholevykh kletok.

Single-stranded DNA breaks in Zajdela's ascitic hepatoma and transformed hamster fibroblasts were caused by treating alive cells with 1% dimethylsulfoxide for 2 h or 100 micrograms/ml bleomycin for 5 min and tested by alkaline and neutral DNA elutions. Electron microscopy of thin sections revealed decompaction of the loosened approximately 25 nm globules within diffuse chromatin into thin fibrillar mesh while supranucleosomal structure of the compact chromatin remained untouched. The chromatin enhanced its affinity for cationic dyes and contrast agents. It is concluded that the diffuse chromatin possesses torsional stress of DNA superhelicity and its loosened subunits represent a form for its organization. They probably correspond to the functionally active (dynamic) nucleosomes which display destruction under DNA domain relaxation caused by one-strand breaks.

[Conformational changes in the chromatin of tumor cells and the phenomenon of nuclear achromasia]. / Konformatsionnye izmeneniia khromatina opukholevykh kletok i fenomen iadernoi akhromazii.

Tumour cell nuclei display achromasia in smears relative to methylene blue and to other low-affinitive ionic dyes. In the course of the smear air-drying the tumour DNA undergoes conformational changes as evidenced by enhanced ethidium and monomer acridine orange uptake and increased sensitivity to DNAse I hydrolysis. Very mild treatment with bleomycin prevents nuclear achromasia with methylene blue in tumour cells. It is concluded that the phenomenon of nuclear achromasia of tumour cells is due to the properties of secondary structure of their DNA.

[Analysis of cellular nucleoproteins by the nucleoprotein-celite chromatography method. III. Two types of DNA-nuclear matrix interaction]. / Analiz kletochnykh nukleoproteidov metodom nukleoproteid-tselit-khromatografii. III. Dva tipa vzaimodeistvii DNK s iadernym matriksom.

There are two types of DNA-nuclear matrix interactions in animal cells as revealed by the release of DNA from isolated nuclei by three successive gradients: NaCl, LiCl-urea and temperature. Nuclei were treated with dissociating agents while being adsorbed on the Celite columns. "Weak" DNA-matrix interactions which dissociate in 1.5 M LiCl-3 M urea at 2 degrees appear to be sensitive to ethidium bromide and resistant to exogeneous nucleases (DNAase I, DNAase II and micrococcal nuclease), to DNA-damaging agents, including alkylators and gamma-irradiation, and also to psoralen-induced cross-links. "Strong" DNA-matrix interactions proved to be very different. They dissociate in 4 M LiCl-8 M urea at approximately 90 degrees, are very sensitive to DNAase I and other nucleases, slightly sensitive to chemicals and irradiation at doses stimulating single-stranded DNA breaks, but resistant to ethidium bromide. DNA strand separation seems to be necessary prerequisite for DNA release from its "strong" complex with nuclear matrix. A model for the topological link between DNA and the nuclear matrix involved in DNA replication complex is discussed.

[Method of contrast enhancement of DNA-containing structures on ultrathin sections]. / Metodika spetsificheskogo usileniia kontrasta DNK-soderzhashchikh struktur na ul'tratonkikh srezakh.

Thin epoxy sections of formaldehyde-fixed tissue underwent depurinizing hydrolysis with 5N HCl at room temperature for 1 hour, the treatment with an aldehyde blocking agent, 10% sodium bisulfite, at 60 degrees C during 90 min, and staining with 2% aqueous uranyl acetate. Introduction of bisulfite considerably enhanced the contrast of chromatin. Staining of DNA in these conditions is summed up of preferential binding of uranyl cations with phosphates of DNA and with bisulfite anions specifically bound to deoxyribose.

[Improved method of staining semithin sections with toluidine blue]. / Uluchshennaia metodika okraski polutonkikh srezov toluidinovym sinim.

Treatment of semi-thin epoxy sections with 0.1 N Hcl prior to staining with toluidine blue (pH approximately 9.0) ensures contrastic and reproducible image of tissues fixed for electron microscopy.

[Current state of the problem of electron-cytochemical detection of DNA]. / Sovremennoe sostoianie problemy elektronno-tsitokhimicheskogo vyiavleniia DNK.

Direct and indirect methods for electron microscopic detection of DNA are reviewed. Indirect methods based on DNA removal or preferential contrasting of RNP-structures make it possible to define DNA localization only in outline. By means of direct methods it is possible to reveal all three components of DNA-phosphoricacid groups, pseudoaldehyde groups of desoxyribose, nitrogen bases. Within the framework of this classification the reactions suggested so far and the results obtained with them have been discussed. The main difficulty--lack of specificity, low contrast, poor reproduction, damage of ultrastructures.

[Modification of the method for staining DNA with basic dyes]. / Midifikatsiia metoda okraski na DNK s osnovnymi krasiteliami

The author studied the mechanisms and the applicability in histochemistry of the sodium bisulfate treatment with subsequent toluidine and methylene blue staining after Felgen's hydrolysis. Bisulfite treatment proved to increase the reaction intensity 11/2-fold; the stain is bound stoichiometrically. Toludidine blue results in a metachromatic and anisotropic staining of the cell nuclei. The method is recommended as a sensitive test for DNA in cytochemical investigations and for the study of dichroism of the DNA-containing structures.

[Sequential demonstration of nucleic acids and protein fractions in the same cells]. / Posledovatel'noe vyiavlenie nukleinovykh kislot i belkovykh fraktsii v odnikh i tekh zhe kletkakh

Some combinations are suggested to reveal cell components in the following sequence: 1) RNA--DNA--total proteins; 2) RNA--DNA--histones; 3) RNA--DNA--histones--total proteins; 4) RNA--DNA--acidic proteins. RNA is stained with pyronin Y (in the mixture with methyl green). DNA--by the Feulgen procedure (with TCA hydrolysis), histones and total proteins--by Alfert--Geschwind's method and acidic proteins--according to the technique of Smetana and Bush. The details of the whole procedure, adopted for formol-fixed smears are described.

[Feulgen reaction conducted on unfixed smears]. / Provedenie reaktsii Fel'gena na nefiksirovannykh mazkakh.

After hydrolysis with 5N HCl at room temperature Feulgen's reaction either with Schiff's reagent or with some basic dyes instead of it can be carried out on the air-dried smears without any chemical fixation. Both reactions give quantitative results.