Modeling heterochromatin regions in transgenic mice

Transgenic mice carrying bovine satellite DNA IV were obtained. The size of the transgene integrated into the mouse genome was approximately 390 kb (about 100 transgene copies) as determined by a semiquantitative PCR. Restriction analysis with isoschizomeric restrictases HpaII and MspI, showed that the alien DNA was methylated. In the genome of a transgenic founder male, two integration sites for satellite DNA IV were revealed by in situ hybridization and in situ PCR. These sites are situated on two different chromosomes: in pericentromeric heterochromatin and within a chromosomal arm. In transgenic mice, de novo formation of heterochromatin regions (C-block and the CMA3 disk within the centromeric heterochromatin of another chromosome) was revealed by C-banding and staining with chromomycin A3. This formation is not characteristic of mice, because their chromosomes normally contain no interstitial C-blocks or sequences intensely stained by chromomycin A3.     

Phospholipid and cholesterol levels in the tumor cell plasma membranes with different sensitivity to doxorubicin

The investigation is aimed to study qualitative and quantitative composition of phospholipids, cholesterol content and lipids unsaturation index in plasma membranes of Guerin's carcinoma cells sensitive or resistant to doxorubicin. The comparison of infrared spectra and phospholipids unsaturation index showed that the unsaturation level of fatty acids in plasma membrane from resistant cells was lower than that from sensitive carcinoma cells. 31P-NMR spectroscopy of plasma membranes phospholipids shows the increase of phosphatidylserine and sphingomyeline content in plasma membrane isolated from resistant tumor as compared with sensitive tumor. The levels of phosphatidylcholine and phosphatidylethanolamine were equal in drug-resistant and drug-sensitive carcinoma strains. Changes in plasma membrane from resistant cells result in elevation of plasma membrane microviscosity and phosphatidylserine level increase can suggest the activation of P-glycoprotein-mediated efflux of doxorubicin.     

An oligonucleotide primer system for amplification of the ribulose-1,5-bisphosphate carboxylase/oxygenase genes of bacteria of various taxonomic groups

Based on the analysis of GenBank nucleotide sequences of the cbbL and cbbM genes, coding for the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBPC), the key enzyme of the Calvin cycle, a primer system was designed that allows about 800-bp-long fragments of these genes to be PCR-ampliflied in various photo- and chemotrophic bacteria. The efficiency of the designed primer system in detection of RuBPC genes was demonstrated in PCR with DNA of taxonomically diverse bacteria possessing RuBPC genes with a known primary structure. Nucleotide sequences of RuBPC gene fragments of bacteria belonging to the genera Acidithiobacillus. Ectothiorhodospira, Magnetospirillum, Methylocapsa, Thioalkalispira, Rhodobacter, and Rhodospirillum were determined to be deposited with GenBank and to be translated into amino acid sequences and subjected to phylogenetic analysis.     

The role of antioxidant systems in the response of Escherichia coli to acetamidophenol and some antibiotics

The role of glutathione and other antioxidant systems in the response of Escherichia coli to acetamidophenol (paracetamol), rifampicin, and chloramphenicol was studied. The exposure of aerobically growing E. coli cells to acetamidophenol diminished the intracellular level of glutathione by 40% and the reduced-to-oxidized glutathione ratio in the cells by 50%, while it enhanced the expression of the antioxidant genes soxS and sodA by 2.7 and 1.8 times, respectively. Glutathione-deficient cells were more susceptible to acetamidophenol than were normal cells. All this suggests that acetamidophenol induces a mild oxidative stress in E. coli cells. The oxidative stress induced by rifampicin was still less pronounced, whereas chloramphenicol-treated E.coli cells exhibited no signs of oxidative stress at all.     

Some specific features of vaginal microflora depending on ecological factors

In 75 female residents of Vladikavkaz, aged 18-45 years, who had applied to the gynecological department with complaints of vaginal discharge, the qualitative and quantitative composition of vaginal microflora was studied. All these women were divided into 2 groups: group 1 including 38 women living in ecologically unfavorable districts of the city and group 2 including 37 women living under ecologically favorable conditions. The relationship between the ecological situation of the districts of residence and vaginal microflora in women of the reproductive age was established. Unfavorable exogenous factors were found to lead to the development of vaginal dysbacteriosis: a sharp decrease in the amount of lactoflora or its complete absence accompanied by increased amount of staphylococci, enterococci, Escherichia coli, Proteus, Klebsiella and yeast-like fungi of the genus Candida.     

Crosslinking of mRNA analog, pUUAGUAUUUAUU derivative with a photoactivated group at the guanosine residue, with human 80S ribosomes

Crosslinking of mRNA analog, dodecaribonucleotide pUUAGUAUUUAUU derivative carrying a perfluoroarylazido group at the guanine N7, was studied in model complexes with 80S ribosomes involving tRNA and in binary complex (i.e., in the absence of tRNA). It was shown that, irrespectively of complex formation conditions (13 mM Mg2+, or 4 mM Mg2+ in the presence of polyamines), the mRNA analog in binary complex with 80S ribosomes was crosslinked with sequence 1840-1849 of 18S rRNA, but in the complexes formed with participation of Phe-TPHKPhe (where the G residue carrying the arylazido group occupied position-3 to the first nucleotide of the UUU codon at the P site) the analog was crosslinked with nucleotide 1207. The presence and the nature of tRNA at the E site had no effect on the environment of position-3 of the mRNA analog. Efficient crosslinking of the mRNA analog with tRNA was observed in all studied types of complex. Modified codon GUA, when located at the E site, underwent crosslinking with both cognate valine tRNA and noncognate aspartate tRNA for which the extent of binding at the E site of 80S ribosomes was almost the same and depended little on Mg2+ concentration and the presence of polyamines.     

Studying plant genome variation using molecular markers

The authors' studies on the organization and variation of plant genome with the use of molecular markers are briefly reviewed with special emphasis on random amplified polymorphic DNA (RAPD), inter simple sequence repeat (ISSR), sequence characterized amplified region (SCAR), and cleaved amplified polymorphic sequence (CAPS) markers detected with the use of polymerase chain reaction (PCR). These markers have been demonstrated to be promising for identifying cultivars and determining the purity of genetic strains of pea. Genetic relationships between strains, cultivars, and mutants of pea have been studied. The role of molecular markers in molecular genetic mapping and localizing the genes of commercially important characters of pea has been shown. The possibility of the use of molecular markers for studying somaclonal variation and detecting mutagenic factors in plants during long-term spaceflights is considered. The prospects of using DNA markers for understanding the organization and variability of higher plant genomes are discussed.     

Equilibrium fluctuations in myoglobin and lysozyme

The angular dependencies of inelastic intensities of Rayleigh scattering of Moessbauer radiation were measured for myoglobin and lysozyme (in the hydration range h = 0.05-0.7). The data were fitted within the framework of model, when two types of intraglobular motions were taken into account: individual motions of small side-chain groups and cooperative motions of segments. The best agreement with the experiment at h > 0.05 was obtained when individual motions of small groups together with the cooperative motions of alpha-helices and beta-sheets for lysozyme, and alpha-helices for myoglobin were considered. At further hydration (h = 0.45), mean-square displacements (x2) of both types of motions strongly increase with the increase in hydration degree, while the motions with a large correlation radius (not less than macromolecule radius) remain nearly the same as for h = 0.05. The results of the study of the radial distribution function deduced by Fourier-transform from the diffuse x-ray measurements together with RSMR data allow one to conclude that the water during protein hydration competes with the intramolecular hydrogen bonds, loosens the protein and increases the internal dynamics. Concurrently, water arranges the ordering of macromolecule, which takes the native structure at h = 0.4-0.7. The analysis of auto and cross-correlation functions of bending fluctuations of alpha-helices in the large domain of lysozyme performed by molecular dynamics allows one to come to the final conclusion that it is the difference in the structural organization of myoglobin and lysozyme and not the presence of SS-bonds in lysozyme macromolecule that is responsible for different structural fluctuations in these proteins.