Effect of repeated stress on the function of pituitary-adrenal and immune systems in gray rats selected for behavior

Reaction of pituitary-adrenal axis to a 10-day immobilisation stress and a humoral immune response to subsequent injection of sheep red blood cells were investigated in gray rats selected for enhancement of decrease of aggressive behavior towards humans. It was show that pituitary-adrenal axis reaction of aggressive animals to repeated stress did not change during the experiment, while a decrease of stress-induced corticosterone level was observed already on day 5 of stress. Repeated stress led to enhancement of humoral immune response in aggressive rats, whereas it did not bring about any change in tame animals. based on the obtained data, it could be supposed that breeding of gray rats for domesticated behavior led a faster adaptation to repeated stress and the absence of stimulating influence on humoral immune response in tame rats.     

A new method for the construction of translationally coupled operons in a bacterial chromosome

A new method for the construction of translationally coupled operons in a bacterial chromosome was developed on the basis of the recombineering approach. The method includes the in vitro construction of an artificial operon with an efficiently translated proximal cistron, its insertion into the Escherichia coli chromosome, the modification of the operon via Red-driven insertion of a special “Junction” with an excisable selective marker into the intercistronic region of the initial operon, and the excision of the marker. The Junction structure was designed and tested. The Junction consists of three components. The first component is the E. coli rplC-rplD intercistronic region and serves for placing the TAA codon of the proximal gene in the SD sequence (TAAGGAG) of rplD. The second component is the Cm ^R gene flanked by λattL/R sites in such a fashion that the residual λattB site after λInt/Xis-driven excision of the marker does not contain termination codons in frame with ATG of rplD. The third component is the E. coli trpE-trpD intercistronic region which is added so that TGA of trpE acts a termination codon of the new open reading frame (ORF), while the overlapping (TGATG) ATG of trpD is in the position of the initiation codon of the distal gene of the original operon. The general design of the Junction provides the conversion of the original two-cistron operon into a three-cistron operon with translationally coupled genes, where the coupling of the artificial ORF (rplD’-λattB-’trpE) with the proximal gene is due to the rplC-rplD intercistronic region and its coupling with the distal gene is due to trpE-trpD. The strategy was experimentally implemented to construct an artificial operon P_tac-aroG4-serA5, where the expression the distal serA5 gene was optimized owing to translational coupling in a three-cistron operon.     

Recombinant Escherichia coli strains deficient in mixed acid fermentation pathways and capable of rapid aerobic growth on glucose with a reduced crabtree effect

In this study, we constructed and characterized Escherichia coli strains deficient in mixed acid fermentation pathways, which are capable of rapid aerobic growth on glucose without pronounced bacterial Crabtree effect. The main pathways of production of acetic and lactic acids and ethanol in these strains were inactivated by a deletion of the ackA, pta, poxB, ldhA, and adhE genes. The phosphoenolpyruvate-dependent phosphotransferase system of glucose transport and phosphorylation was inactivated in the strains by a deletion of the ptsG gene. The possibility of alternative transport and phosphorylation of the carbohydrate substrate was ensured in recombinants by constitutive expression of the galP and glk genes, which encode the low-affinity H⁺-symporter of D-galactose and glucokinase, respectively. The resulting SGM1.0ΔptsG P_tacgalP and SGM1.0ΔptsG PLglk P_tacgalP strains were capable of rapid aerobic growth in a minimal medium containing 2.0 and 10.0 g/L of glucose and secreted only small amounts of acetic acid and trace amounts of pyruvic acid.     

Anaerobic biosynthesis of intermediates of reductive branch of tricarboxylic acids cycle by <Emphasis Type="Italic">Escherichia coli</Emphasis> strains with inactivated <Emphasis Type="Italic">frdAB</Emphasis> and <Emphasis Type="Italic">sdhAB</Emphasis> genes

The genes frdAB and sdhAB, which encode components of fumarate reductase and succinate dehydrogenase, have been deleted in a recombinant E. coli strain with the inactivated pathways of mixed-acid fermentation and a modified system of glucose transport and phosphorylation upon the heterological expression of the pyruvate carboxylase gene. Under anaerobic conditions, the parental strain efficiently converted glucose to succinic acid without synthesizing notable amounts of fumaric or malic acid. Upon individual deletion of the frdAB genes, the mutant strain fermented glucose to succinic acid less efficiently secreting notable amounts of malic and fumaric acids. Individual deletion of the sdhAB genes in the parental strain did not significantly affect the formation of the main fermentation end-product. The combined inactivation of fumarate reductase and succinate dehydrogenase in the constructed strain enhanced the anaerobic conversion of glucose to fumaric and malic acids with the activation of the glyoxylate bypass and decrease in the contribution of the reductive branch of the TCA cycle to the formation of the target products.     

S-Arachidonoyl-2-thioglycerol synthesis and use for fluorimetric and colorimetric assays of monoacylglycerol lipase

We report the first synthesis of 2-thioglycerol and S-arachidonoyl-2-thioglycerol (the thioester analog of the endocannabinoid 2-arachidonoylglycerol) in an eight or nine step procedure with a yield of ～25% and establish the use of this substrate for maleimide-based fluorescent and dithiobis(2-nitrobenzoic acid)-based colorimetric assays of human recombinant monoacylglycerol (MAG) lipase (hMAGL) and human brain membrane MAG hydrolase activity. Inhibitor structure–activity relationships observed here for hMAGL and 2-ATG correlate well (r² = 0.93, n = 9) with earlier findings for mouse brain MAG hydrolase with non-thiol substrates.     

A Phosphodiesterase from Ascites Carcinoma Krebs II Cells Specifically Cleaves the Bond between VPg and RNA of Encephalomyocarditis Virus

The substrate specificity of the unlinking enzyme from ascites carcinoma Krebs II cells has been investigated. The enzyme specifically splits the interpolymeric phosphodiester bond between Kp and the 5´-terminal phosphate group of the uridylic acid residue in the Kp–pUpUpGp complex.     

Gonadal and adrenal endocrine functions in female minks of 2 genotypes in postnatal ontogeny

Radioimmune assay has been made of the content of estradiol and progesterone in the blood (from July to March including the 7th day after mating), as well as on the level of estradiol and progesterone production in young females of the standard and mutant sapphire minks in November. It was shown that within certain periods, estradiol and progesterone content of the blood was significantly higher in the standard animals. Gonadal production of estradiol, as well as progesterone production both by the gonads and adrenals in November, were similar in females of both genotypes. It is suggested that sapphire minks have another pattern of correlation between estradiol content of the blood and gonadal production of estradiol as compared to that in standard animals.     

Heterochromatinic segments in chromosomes of cultured human cells, sensitive and resistant to Coxsackie B viruses

Comparative karyological studies of C-heterochromatin have been made on line J-96 of human cells, which are susceptible to enteroviruses, and on cell line J-41 derived from this culture and possessing highly specific resistance to Coxsackie B viruses. It was shown that the development of specific resistance to Coxsackie B viruses was accompanied by the loss of one of the chromosomes of pairs 1 and 9, and by the dissapearance of two marker chromosomes. There appeared new marker chromosomes with additional C-heterochromatain regions. The data obtained are discussed with respect to a possible interrelationship between these chromosomal alterations and the specific resistance to Coxsakie B viruses.     

Relationship between group G chromosomes, especially chromosome 21, and the sensitivity of human cells to Coxsackie B viruses]

The R-method of differential chromosome staining by length was applied to comparative karyological studies on the culture of J 96 human cells susceptible to enteroviruses, and on the J 41 cell line derived from this culture and possessing high specific resistance to Coxsackie B viruses. Karyotype of the J 41 cell line was shown to be deprived of chromosome G21 (P less than 0.0001). The number of other chromosomes varied from cell to cell, but they are constantly present in the majority of cells of both the J 96 and J 41 cell lines. A conclusion is drawn that chromosome G21 incorporates gene(s) which controls the human cells susceptibility to Coxsackie B viruses.