Vol 52, No 11 (2016)

The dependence of expression of miRNAs and their precursors (pre-miRNAs) on the DNA methylation level in HeLa cells 8 days after mitomycin C treatment was studied. A massive parallel DNA sequencing method was applied to analyze miRNA expression. 5-Azacytidine (DNA methylation inhibitor) was added to the medium 6 days after mutagenic agent exposure. The results indicated that the change in expression for some mature miRNAs (39 of 61) was accompanied by the change in the expression of their pre-miRNAs, while there were no significant changes in the expression of pre-miRNA for other mature miRNAs (22 of 61). The aberrant expression was maintained by 8 of 61 mature miRNAs and 6 of 55 pre-miRNAs in the induced HeLa cells after 5-azacytidine treatment. In addition, the expression of more than 90% of miRNAs, which indicated a significant change in expression after mitomycin C treatment, does not depend or depends slightly on the DNA methylation level in HeLa cells without mitomycin C treatment. The results suggest that mitomycin C induces aberrant DNA methylation which affects maintenance of changes in the miRNA expression in cell generations after mutagen treatment.

In meiotic prophase I, chromatin fibrils attached to the lateral elements of the synaptonemal complexes (SC) form loops. SCAR DNA (synaptonemal complex associated regions of DNA) is a family of genomic DNA tightly associated with the SC and located at the chromatin loop basements. Using the hybridization technique, it was demonstrated that localization of SCAR DNA was evolutionarily conserved in the isochore compositional fractions of the three examined genomes of warm-blooded vertebrates—human, chicken, and golden hamster. The introduction of the concept of the comparative loops (CL) of DNA that form of chromatin attach to SC in the isochore compositional fractions provided the calculation of their length. An inverse proportional relationship between the length of CL DNA and the GC level in the isochore compartments of the studied warm-blooded vertebrate genomes was revealed. An exception was the GCpoorest L1 isochore family. For different compositional isochores of the human and chicken genomes, the number of genes in the CL DNA was evaluated. A model of the formation of GC-rich isochores in vertebrate genomes, according to which there was not only an increase in the GC level but also the elimination of functionally insignificant noncoding DNA regions, as well as joining of isochores decreasing in size, was suggested.

The nucleotide sequence of cryptic plasmid (designated as pBL90) detected in the cells of Brevibacterium lactofermentum DSM 1412 was determined. The length of plasmid DNA is 67826 bp. Comparison of the nucleotide sequence of pBL90 with known plasmid sequences showed no long regions of significant homology. Computer analysis of the plasmid DNA revealed 29 open reading frames (ORFs). The amino acid sequences of 15 ORFs (approximately 25% of plasmid length) have a high (>70%) level of identity to proteins from different plasmids of Corynebacterium representatives, including replicative proteins. Unusual in pBL90 is the presence of replicative genes from two different families and types of replication.

Transgenic Nicotiana tabacum L. cv. SR1 plants, characterized by an increase in the level of dsRNA-specific hydrolytic activity after induction by wounding, were obtained. The Solanum lycopersicum anionic peroxidase gene promoter (new for plant genetic engineering) was for the first time used for the induced expression of the target Serratia marcescens RNase III gene. Upon infection with the tobacco mosaic virus (TMV), the transgenic plants of the obtained lines did not differ significantly from the control group in the level of TMV capsid protein accumulation. In general, no delay in the development of the infection symptoms was observed in transgenic plants as compared with the control group. The obtained transgenic plants represent a new model for the study of the biological role of endoribonucleases from the RNase III family, including in molecular mechanisms of resistance to pathogens.

Genetic similarity and relatedness within the set of pear genotypes including autochthonous Circassian cultivars from North Caucasus, European cultivars, accessions of Pyrus caucasica Fed., and modern Russian cultivars were estimated on the basis of analysis of SSR loci. The level of polymorphism for the studied loci varied from 11 to 15 alleles per locus in the set of 29 samples of pears. A higher level of allelic polymorphism of SSR loci was revealed for a set of P. caucasica samples in comparison with modern cultivated cultivars: from 9 to 12 alleles for P. caucasica and from 6 to 8 alleles for modern cultivars. Specific alleles for the mentioned groups of pears were identified. UPGMA clustering revealed two distinct groups: one includes P. caucasica accessions and autochthonous Caucasian cultivars and the other group includes all cultivated European and Russian pear cultivar. The results support the hypothesis of an isolated gene pool formation of autochthonous pear cultivars of the North Caucasus and their probable origin from the wild P. caucasica.

To study the phylogenetic relationships, evolutionary history, and molecular systematics of firs (genus Abies), the phylogenetic reconstruction, based on nuclear multilocus markers—amplified fragment length polymorphism (AFLP)—was conducted. Using seven combinations of selective primers, 84 samples of 39 taxa were genotyped for 553 polymorphic AFLP loci. A comparison with our earlier chloroplast and mitochondrial phylogenies of the genus (in 2014) shows that the nuclear phylogeny generally is more congruent to the chloroplast tree. Most of the clades resolved by the chloroplast phylogeny were supported also in the AFLP tree. Employing the nuclear DNA-based tree, we revealed the presence of new groups and the differences in the topology of several clades. AFLP confirmed the monophyly of Asian species of section Balsamea and their sister position in relation to the American group of species of this section. As shown by the tree of chloroplast DNA, Asian species of section Balsamea do not form a monophyletic group, but belong to the clade comprising the majority of Asian species. Phylogenetically mitochondrial DNA data to a large extent are not congruent to the nuclear and chloroplast DNA trees, and are more in line with geographical distribution of species. Conflicts between nuclear and cytoplasmic phylogeny were analyzed. Taking them into account, we consider the hypothesis of a hybrid origin of particular groups of firs, including ancient hybridization in section Balsamea. A comparison of molecular data with traditional taxonomy of the genus is discussed.

Somatic mitotic and meiotic chromosomes at the pachytene and at the metaphase I of the males of the viviparous lizard, Zootoca vivipara (Lichtenstein, 1823), from northwestern Russia, belonging to the Russian form of Z. v. vivipara, are examined. The spreading of synaptonemal complexes (SC) of their chromosomes are obtained and analyzed for the first time. Eighteen SC are observed, including SC of the Z₁Z₁ (pairs 5 or 6) and the Z₂Z₂ (pair 13) sex chromosomes. Characteristics of SC are compared with the number and the shape of bivalents and with those of the karyotype structure. In the studied Russian form of Z. v. vivipara, the length ratios of bivalents correlate with that of mitotic chromosomes (2n = 36); however, some specificity in the morphology of SC of the Z₁Z₁ sex chromosomes is reported in this article.

The increase in diabetes was noted at the turn of the 21st century. Patients with type 2 diabetes (T2DM) make up the majority of patients. Diabetes is a multifactorial disease. It arises from adverse effects of environmental factors on the body of genetically susceptible peoples. According to modern concepts, T2DM is a polygenic disease. Each of the involved genes contributes to the risk of developing of this disease. In our study, the association between polymorphic genetic markers rs7756992, rs9465871, rs7754840, and rs10946398 in the CDKAL1 gene and rs1111875 in the HHEX/IDE locus and T2DM in the Russian population were studied. Four hundred forty patients with type 2 diabetes and 264 healthy individuals without any signs of the disease were examined. The comparative analysis of distribution of genotypes and allele frequencies points to an association between polymorphic genetic markers rs7756992, rs9465871, and rs10946398 in the CDKAL1 gene and this disease. For the other polymorphic genetic markers (rs7754840 in the CDKAL1 gene and rs1111875 in the HHEX/IDE locus), no statistically significant associations are found. On the basis of these data, we can conclude that the CDKAL1 gene is associated with development of T2DM. For the HHEX/IDE locus, such an association is absent.

The value of chromatin diminution (CD) in different species of freshwater cyclopoid copepods can differ significantly. The biological and evolutionary roles of these differences remain unclear. To expand the knowledge on CD distribution and magnitude in this group of copepods, a quick method for its evaluation was required. This study proposes a simple approach for CD assessment in copepods using quantitative realtime PCR (qPCR). The magnitude of changes in the genome size was assessed by comparing fluorescence curves of qPCR fragments of target genes for pre- and post-diminution materials. The method was tested on four cyclopoid copepods species. In Cyclops kolensis, CD was assessed as 95.3 ± 1.2; in Acanthocyclops vernalis it was assessed at 94.6 ± 0.8%; at C. insignis, it was 82.3 ± 5.2%; and for the first time, CD was found in Megacyclops viridis at 91.1 ± 2.6%. The advantages of our approach are its rapidity, simplicity and minimal requirements of materials studied.

Increasing evidence suggests that noncoding RNA transcribed from the enhancers play an important role in the regulation of gene transcription. Insulators are the regulatory elements that limit the activity of enhancers and form independent transcriptional domains. Using a transgenic lines, we show that the Fab-7 insulator of the bithorax complex and the MDG4 (gypsy) insulator are able to disrupt weak transcription derived from the enhancer regulating the white gene expression in the eyes. The ability of insulators to disrupt weak transcription may play a role in the enhancer-blocking activity.

The insulin/insulin-like growth factor signaling pathway is involved in the regulation of the synthesis of insect gonadotropic hormones, juvenile (JH) and 20-hydroxyecdysone (20E). We carried out the immunohistochemical analysis of the insulin receptor (InR) expression in the corpus allatum (the JH-producing gland) and in the ovarian follicular cells (a site for the synthesis of 20E precursor, ecdysone) in the process of sexual maturation of D. melanogaster females and examined the influence of exogenous JH on the InR expression in these tissues. For the first time, it was demonstrated that InR was expressed in follicular cells and that its expression in corpus allatum and follicular cells of Drosophila females was stage-specific, i.e., the expression intensity in young females greatly exceeded that in mature individuals. We also found a negative feedback loop in the regulation of JH levels by the insulin signaling pathway in Drosophila adults: the experimental increase in the JH titers in young females dramatically reduced the InR expression intensity in corpus allatum and follicular cells.