Source: https://chemweb.com/articles/SV10541/0008000010
Timestamp: 2019-04-22 02:36:15+00:00

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Mechanisms of stress resistance and gene regulation in the radioresistant bacterium Deinococcus radiodurans by A. A. Agapov; A. V. Kulbachinskiy (1201-1216).
The bacterium Deinococcus radiodurans reveals extraordinary resistance to ionizing radiation, oxidative stress, desiccation, and other damaging conditions. In this review, we consider the main molecular mechanisms underlying such resistance, including the action of specific DNA repair and antioxidation systems, and transcription regulation during the anti-stress response.
Molecular mechanisms of latent inflammation in metabolic syndrome. Possible role of sirtuins and peroxisome proliferator-activated receptor type γ by I. S. Stafeev; M. Y. Menshikov; Z. I. Tsokolaeva; M. V. Shestakova; Ye. V. Parfyonova (1217-1226).
The problem of metabolic syndrome is one of the most important in medicine today. The main hazard of metabolic syndrome is development of latent inflammation in adipose tissue, which promotes atherosclerosis, non-alcoholic fatty liver disease, myocarditis, and a number of other illnesses. Therefore, understanding of molecular mechanisms of latent inflammation in adipose tissue is very important for treatment of metabolic syndrome. Three main components that arise during hypertrophy and hyperplasia of adipocytes underlie such inflammation: endoplasmic reticulum stress, oxidative stress, and hypoxia. Each of these components mediates activation in different ways of the key factor of inflammation–NF-κB. For metabolic syndrome therapy, it is suggested to influence a number of inflammatory signaling components by activating other cell factors to suppress development of inflammation. Such potential factors are peroxisome proliferator-activated receptors type γ that suppress transcription factor NF-κB through direct contact or via kinase of a NF-κB inhibitor (IKK), and also the antiinflammatory transcription factor AP-1. Other possible targets are type 3 NAD+-dependent histone deacetylases (sirtuins). There are mutually antagonistic relationships between NF-κB and sirtuin type 1 that prevent development of inflammation in metabolic syndrome. Moreover, sirtuin type 1 inhibits the antiinflammatory transcription factor AP-1. Study of the influence of these factors on the relationship between macrophages and adipocytes, macrophages, and adipose tissue-derived stromal cells can help to understand mechanisms of signaling and development of latent inflammation in metabolic syndrome.
Competitive agonists and antagonists of steroid nuclear receptors: Evolution of the concept or its reversal by O. V. Smirnova (1227-1234).
Guidance receptors in the nervous and cardiovascular systems by K. A. Rubina; V. A. Tkachuk (1235-1253).
Blood vessels and nervous fibers grow in parallel, for they express similar receptors for chemokine substances. Recently, much attention is being given to studying guidance receptors and their ligands besides the growth factors, cytokines, and chemokines necessary to form structures in the nervous and vascular systems. Such guidance molecules determine trajectory for growing axons and vessels. Guidance molecules include Ephrins and their receptors, Neuropilins and Plexins as receptors for Semaphorins, Robos as receptors for Slit-proteins, and UNC5B receptors binding Netrins. Apart from these receptors and their ligands, urokinase and its receptor (uPAR) and T-cadherin are also classified as guidance molecules. The urokinase system mediates local proteolysis at the leading edge of cells, thereby providing directed migration. T-cadherin is a repellent molecule that regulates the direction of growing axons and blood vessels. Guidance receptors also play an important role in the diseases of the nervous and cardiovascular systems.
Association of high light-inducible HliA/HliB stress proteins with photosystem 1 trimers and monomers of the cyanobacterium Synechocystis PCC 6803 by D. V. Akulinkina; Yu. V. Bolychevtseva; I. V. Elanskaya; N. V. Karapetyan; N. P. Yurina (1254-1261).
Hlip (high light-inducible proteins) are important for protection of the photosynthetic apparatus of cyanobacteria from light stress. However, the interaction of these proteins with chlorophyll–protein complexes of thylakoids remains unclear. The association of HliA/HliB stress proteins with photosystem 1 (PS1) complexes of the cyanobacterium Synechocystis PCC 6803 was studied to understand their function. Western blotting demonstrated that stress-induced HliA/HliB proteins are associated with PS1 trimers in wild-type cells grown under moderate light condition (40 µmol photons/m2 per sec). The content of these proteins increased 1.7-fold after light stress (150 µmol photons/m2 per sec) for 1 h. In the absence of PS1 trimers (ΔpsaL mutant), the HliA/HliB proteins are associated with PS1 monomers and the PS2 complex. HliA/HliB proteins are associated with PS1 monomers but not with PS1 trimers in Synechocystis PS2-deficient mutant grown at 5 µmol photons/m2 per sec; the content of Hli proteins associated with PS1 monomers increased 1.2-fold after light stress. The HliA/HliB proteins were not detected in wild-type cells of cyanobacteria grown in glucose-supplemented medium at 5 µmol photons/m2 per sec, but light stress induces the synthesis of stress proteins associated with PS1 trimers. Thus, for the first time, the association of HliA/HliB proteins not only with PS1 trimers, but also with PS1 monomers is shown, which suggests a universal role of these proteins in the protection of the photosynthetic apparatus from excess light.
Regulation of cyclooxygenase 2 expression by agonists of PPAR nuclear receptors in the model of endotoxin tolerance in astrocytes by A. A. Astakhova; D. V. Chistyakov; E. V. Pankevich; M. G. Sergeeva (1262-1270).
Endotoxin tolerance (ET) represents a state of an altered immune response induced by multiple stimulations of a cell, a tissue, or an organism with lipopolysaccharide. Characteristics of ET include downregulation of induction of proinflammatory genes (TNFα, IL6, and others) and enhancement of induction of antiinflammatory genes (IL10, TGFβ). ET generally has protective functions; nevertheless, it might result in a state of innate immune deficiency and cause negative outcomes. A current issue is the search for the mechanisms controlling the level of inflammation in the course of endotoxin tolerance. In this work, we investigated the change in cyclooxygenase 2 (Cox2) expression in the model of endotoxin tolerance in astrocytes and analyzed the possibility of regulating this process applying nuclear receptor PPAR agonists. Our results indicate that: 1) endotoxin tolerance can be induced in astrocytes and results in TNFα and Cox2 mRNA induction decrease upon secondary stimulation; 2) tolerance is revealed on the level of TNFα release and Cox2 protein expression; 3) PPAR agonists GW7647, L-165041, and rosiglitazone control Cox2 mRNA expression levels under conditions of endotoxin tolerance. In particular, rosiglitazone (a PPARγ agonist) induces Cox2 mRNA expression, while GW7647 (a PPARα agonist) and L-165041 (a PPARβ agonist) suppress the expression. Our results demonstrate that Cox2 can be upand downregulated during endotoxin tolerance in astrocytes, and PPAR agonists might be effective for controlling this target under conditions of multiple proinflammatory stimulations of brain tissues with endotoxin.
Purification and characterization of recombinant Deinococcus radiodurans RNA Polymerase by D. M. Esyunina; A. V. Kulbachinskiy (1271-1279).
The radioresistant bacterium Deinococcus radiodurans is one of the most interesting models for studies of cell stress resistance. Analysis of the mechanisms of gene expression in D. radiodurans revealed some specific features of the transcription apparatus that might play a role in cell resistance to DNA-damaging conditions. In particular, RNA polymerase from D. radiodurans forms unstable promoter complexes and during transcription elongation has a much higher rate of RNA cleavage than RNA polymerase from Escherichia coli. Analysis of the structure and functions of D. radiodurans RNA polymerase is complicated due to the absence of convenient genetic systems for making mutations in the RNA polymerase genes and difficulties with enzyme purification. In this work, we developed a system for expression of D. radiodurans RNA polymerase in E. coli cells. We obtained an expression vector encoding all core RNA polymerase subunits and defined optimal conditions for the expression and purification of the RNA polymerase. It was found that D. radiodurans RNA polymerase has much higher rates of RNA cleavage than E. coli RNA polymerase under a wide range of conditions, including variations in the concentration of catalytic magnesium ions and pH values of the reaction buffer. The expression system can be used for further studies of the RNA cleavage reaction and the mechanisms of transcription regulation in D. radiodurans, including analysis of mutant RNA polymerase variants.
Temperature dependence of light-induced absorbance changes associated with chlorophyll photooxidation in manganese-depleted core complexes of photosystem II by A. A. Zabelin; V. A. Shkuropatova; A. Ya. Shkuropatov; V. A. Shuvalov (1279-1287).
Mid-infrared (4500-1150 cm–1) absorbance changes induced by continuous illumination of Mn-depleted core complexes of photosystem II (PSII) from spinach in the presence of exogenous electron acceptors (potassium ferricyanide and silicomolybdate) were studied by FTIR difference spectroscopy in the temperature range 100–265 K. The FTIR difference spectrum for photooxidation of the chlorophyll dimer P680 was determined from the set of signals associated with oxidation of secondary electron donors (β-carotene, chlorophyll) and reduction of the primary quinone QA. On the basis of analysis of the temperature dependence of the P 680 + /P680 FTIR spectrum, it was concluded that frequencies of 131-keto-C=O stretching modes of neutral chlorophyll molecules PD1 and PD2, which constitute P680, are similar to each other, being located at ∼1700 cm–1. This together with considerable difference between the stretching mode frequencies of keto groups of P D1 + and P+ D2 cations (1724 and 1709 cm–1, respectively) is in agreement with a literature model (Okubo et al. (2007) Biochemistry, 46, 4390–4397) suggesting that the positive charge in the P 680 + dimer is mainly localized on one of the two chlorophyll molecules. A partial delocalization of the charge between the PD1 and PD2 molecules in P 680 + is supported by the presence of a characteristic electronic intervalence band at ∼3000 cm–1. It is shown that a bleaching band at 1680 cm–1 in the P 680 + /P680 FTIR spectrum does not belong to P680. A possible origin of this band is discussed, taking into account the temperature dependence (100–265 K) of light-induced absorbance changes of PSII core complexes in the visible spectral region from 620 to 720 nm.
Phosphorylation regulates interaction of 210-kDa myosin light chain kinase N-terminal domain with actin cytoskeleton by E. L. Vilitkevich; A. Y. Khapchaev; D. S. Kudryashov; A. V. Nikashin; J. P. Schavocky; T. J. Lukas; D. M. Watterson; V. P. Shirinsky (1288-1297).
High molecular weight myosin light chain kinase (MLCK210) is a multifunctional protein involved in myosin II activation and integration of cytoskeletal components in cells. MLCK210 possesses actin-binding regions both in the central part of the molecule and in its N-terminal tail domain. In HeLa cells, mitotic protein kinase Aurora B was suggested to phosphorylate MLCK210 N-terminal tail at serine residues (Dulyaninova, N. G., and Bresnick, A. R. (2004) Exp. Cell Res., 299, 303–314), but the functional significance of the phosphorylation was not established. We report here that in vitro, the N-terminal actin-binding domain of MLCK210 is located within residues 27-157 (N27-157, avian MLCK210 sequence) and is phosphorylated by cAMP-dependent protein kinase (PKA) and Aurora B at serine residues 140/149 leading to a decrease in N27-157 binding to actin. The same residues are phosphorylated in a PKA-dependent manner in transfected HeLa cells. Further, in transfected cells, phosphomimetic mutants of N27-157 showed reduced association with the detergent-stable cytoskeleton, whereas in vitro, the single S149D mutation reduced N27-157 association with F-actin to a similar extent as that achieved by N27-157 phosphorylation. Altogether, our results indicate that phosphorylation of MLCK210 at distinct serine residues, mainly at S149, attenuates the interaction of MLCK210 N-terminus with the actin cytoskeleton and might serve to regulate MLCK210 microfilament cross-linking activity in cells.
Cytochrome c–cardiolipin complex in a nonpolar environment by A. S. Vikulina; A. V. Alekseev; E. V. Proskurnina; Yu. A. Vladimirov (1298-1302).
Programmed cell death (apoptosis) plays an important role in the life of multicellular organisms and in the development of socially significant human diseases. Cytochrome c–cardiolipin complex (Cyt-CL) is formed at the very beginning of a cascade of apoptotic reactions. Nevertheless, the structure of the complex and the mechanism of its participation in lipid peroxidation in mitochondrial membranes are not yet understood. In previous work (Vladimirov, Y. A., et al. (2011) Crystallography, 56, 712-719), it was shown that the Cyt-CL complex precipitates in concentrated water solution, the sediment containing orderly nanospheres formed by cytochrome c molecules with changed conformation and surrounded by a cardiolipin monolayer, and they are essentially hydrophobic. In this work, we obtained chloroform and hexane solutions of Cyt-CL with lipid/protein ratio of 77 ± 11. The conditions are described under which the solutions were obtained. Study of the properties of Cyt-CL solutions in hydrophobic media will reveal their structure and the mechanism of their catalytic activity inside the lipid layer of biological membranes.
Isolation and purification of recombinant serine/threonine protein kinases of the strain Bifidobacterium longum B379M and investigation of their activity by M. G. Alekseeva; D. A. Mavletova; N. V. Kolchina; V. Z. Nezametdinova; V. N. Danilenko (1303-1311).
Previously, we identified six serine/threonine protein kinases (STPK) of Bifidobacterium and named them Pkb1–Pkb6. In the present study, we optimized methods for isolation of the six STPK catalytic domains proteins of B. longum B379M: a method for isolation of Pkb3 and Pkb4 in native conditions, a method for isolation of Pkb5 in denaturing conditions, and a method for isolation of Pkb1, Pkb2, and Pkb6 from inclusion bodies. The dialysis conditions for the renaturation of the proteins were optimized. All of the enzymes were isolated in quantities sufficient for study of the protein activity. The proteins were homogeneous according to SDS-PAGE. The autophosphorylation ability of Pkb1, Pkb3, Pkb4, and Pkb6 was investigated for the first time. Autophosphorylation was detected only for the Pkb3 catalytic domain.
Polyphosphates and polyphosphatase activity in the yeast Saccharomyces cerevisiae during overexpression of the DDP1 gene by L. V. Trilisenko; N. A. Andreeva; M. A. Eldarov; M. V. Dumina; T. V. Kulakovskaya (1312-1317).
The effects of overexpression of yeast diphosphoinositol polyphosphate phosphohydrolase (DDP1) having endopolyphosphatase activity on inorganic polyphosphate metabolism in Saccharomyces cerevisiae were studied. The endopolyphosphatase activity in the transformed strain significantly increased compared to the parent strain. This activity was observed with polyphosphates of different chain length, being suppressed by 2 mM tripolyphosphate or ATP. The content of acid-soluble and acid-insoluble polyphosphates under DDP1 overexpression decreased by 9 and 28%, respectively. The average chain length of salt-soluble and alkali-soluble fractions did not change in the overexpressing strain, and that of acid-soluble polyphosphate increased under phosphate excess. At the initial stage of polyphosphate recovery after phosphorus starvation, the chain length of the acid-soluble fraction in transformed cells was lower compared to the recipient strain. This observation suggests the complex nature of DDP1 involvement in the regulation of polyphosphate content and chain length in yeasts.
Matrix metalloproteinases in primary culture of cardiomyocytes by N. B. Bildyug; I. V. Voronkina; L. V. Smagina; N. M. Yudintseva; G. P. Pinaev (1318-1326).
The highly organized contractile apparatus of cardiomyocytes in heart tissue allows for their continuous contractility, whereas extracellular matrix components are synthesized and spatially organized by fibroblasts and endothelial cells. However, reorganization of the cardiomyocyte contractile apparatus occurs upon their 2D cultivation, which is accompanied by transient loss of their contractility and acquired capability of extracellular matrix synthesis (Bildyug, N. B., and Pinaev, G. P. (2013) Tsitologiya, 55, 713-724). In this study, matrix metalloproteinases were investigated at different times of cardiomyocyte 2D cultivation and 3D cultivation in collagen gels. It was found that cardiomyocytes in 2D culture synthesize matrix metalloproteinases MMP-2 and MMP-9, wherein their amount varies with the cultivation time. The peak MMP-9 amount is at early cultivation time, when the reorganization of cardiomyocyte contractile apparatus occurs, and the MMP-2 peak precedes the recovery of the initial organization of their contractile apparatus. Upon cardiomyocyte cultivation in 3D collagen gels, in which case their contractile apparatus does not rearrange, a steady small amount of MMP2 and MMP-9 is observed. These data indicate that the cardiomyocyte contractile apparatus reorganization in culture is associated with synthesis and spatial organization of their own extracellular matrix.
Formation of new polysomes on free mRNAs in a cell-free translation systems is accompanied by partial disassembly of previously formed polysomes by E. A. Sogorin; S. Ch. Agalarov; A. S. Spirin (1327-1330).
A method for detection of the fluorescence-labeled mRNA in translating ribosomal complexes has been developed. It is demonstrated that in the working cell-free translation system with preformed polysomes, formation of new polysomes on free mRNA takes place. For the first time, it is shown that the process is accompanied by partial disassembly of the previously formed polysomes. This result is interpreted as an indication of the direct relationship between processes of translation termination of polysomal ribosomes and translation initiation of free mRNAs.
Truncated variants of Serratia proteamaculans oligopeptidase B having different activities by A. G. Mikhailova; A. N. Nekrasov; A. A. Zinchenko; T. V. Rakitina; D. A. Korzhenevsky; A. V. Lipkin; O. A. Razguljaeva; M. V. Ovchinnikova; V. A. Gorlenko; L. D. Rumsh (1331-1343).
Treatment of native psychrophilic oligopeptidase B from Serratia proteamaculans (PSP, 78 kDa) with chymotrypsin (soluble or immobilized on modified porous glass MPG-PA) in the presence of 50% glycerol leads to production of a truncated enzyme form (PSP-Chtr, ∼66 kDa), which retains activity toward the low molecular weight substrate of PSP, BAPNA, but in contrast to PSP, is active toward the protein substrate azocasein. It has been shown by MALDI-TOF massspectrometry that PSP-Chtr lacks the N-terminal region of the molecule that envelops the catalytic domain of PSP and supposedly prevents hydrolysis of high molecular weight substrates. It has also been established that the lacking fragment corresponds to the N-terminal highest rank element of the informational structure of PSP. This finding confirms the usefulness of the method of informational structure analysis for protein engineering of enzymes. A similar treatment of PSP with immobilized trypsin also led to production of a stable truncated enzyme form (PSP-Tr, ∼75 kDa) which lacked 22 C-terminal amino acid residues and completely lost enzymatic activity, presumably because of changes in the nearest environment of His652 of the catalytic triad.
Overexpression of selenoprotein SelK in BGC-823 cells inhibits cell adhesion and migration by S. B. Ben; B. Peng; G. C. Wang; C. Li; H. F. Gu; H. Jiang; X. L. Meng; B. J. Lee; C. L. Chen (1344-1353).
Effects of human selenoprotein SelK on the adhesion and migration ability of human gastric cancer BGC-823 cells using Matrigel adhesion and transwell migration assays, respectively, were investigated in this study. The Matrigel adhesion ability of BGC-823 cells that overexpressed SelK declined extremely significantly (p < 0.01) compared with that of the cells not expressing the protein. The migration ability of BGC-823 cells that overexpressed SelK also declined extremely significantly (p < 0.01). On the other hand, the Matrigel adhesion ability and migration ability of the cells that overexpressed C-terminally truncated SelK did not decline significantly. The Matrigel adhesion ability and migration ability of human embryonic kidney HEK-293 cells that overexpressed SelK did not show significant change (p > 0.05) with the cells that overexpressed the C-terminally truncated protein. In addition to the effect on Matrigel adhesion and migration, the overexpression of SelK also caused a loss in cell viability (as measured by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H tetrazolium bromide (MTT) colorimetric assay) and induced apoptosis as shown by confocal microscopy and flow cytometry. The cytosolic free Ca2+ level of these cells was significantly increased as detected by flow cytometry. But the overexpression of SelK in HEK-293 cells did not cause either significant loss in cell viability or apoptosis induction. Only the elevation of cytosolic free Ca2+ level in these cells was significant. Taken together, the results suggest that the overexpression of SelK can inhibit human cancer cell Matrigel adhesion and migration and cause both the loss in cell viability and induction of apoptosis. The release of intracellular Ca2+ from the endoplasmic reticulum might be a mechanism whereby the protein exerted its impact. Furthermore, only the full-length protein, but not C-terminally truncated form, was capable of producing such impact. The embryonic cells were not influenced by the elevation of free Ca2+ level in cytosol, probably due to their much greater tolerance to the variation.
Seed storage globulins: Origin and evolution of primary and higher order structures by A. S. Rudakova; A. M. Cherdivară; K. A. Wilson; A. D. Shutov (1354-1361).
Legumin and vicilin are two-domain seed storage globulins similar in primary and higher order structures of their domains to single-domain plant germins as well as to the domains of two-domain and single-domain bacterial oxalate decarboxylases. Independent evolutionary pathways have been shown for the descent of the storage globulins and germins from two-domain and single-domain bacterial oxalate decarboxylases, respectively. As compared to vicilins, the primary and tertiary structures of legumins were found to most closely reflect the ancient features characteristic of a common precursor of storage globulins. During the evolution of the storage globulins, a mechanism specifically controlling their degradation has been formed. We found that limited proteolysis of soybean legumin and kidney bean vicilin in germinating seeds and in vitro leads to their regular changes, which initiate an extensive cleavage of storage globulin molecules by the one-by-one mechanism. As also shown, limited proteolysis of soybean legumin loosens the intersubunit interactions in its oligomeric molecule. Based on these data, we hypothesize that the deep one-by-one degradation of soybean legumin is triggered by its dissociation, which bares peptide bonds potentially susceptible to proteolytic attack but are masked in the oligomer.
Evolutionary divergence of Arabidopsis thaliana classical peroxidases by E. V. Kupriyanova; P. O. Mamoshina; T. A. Ezhova (1362-1372).
Polymorphisms of 62 peroxidase genes derived from Arabidopsis thaliana were investigated to evaluate evolutionary dynamics and divergence of peroxidase proteins. By comparing divergence of duplicated genes AtPrx53-AtPrx54 and AtPrx36-AtPrx72 and their products, nucleotide and amino acid substitutions were identified that were apparently targets of positive selection. These substitutions were detected among paralogs of 461 ecotypes from Arabidopsis thaliana. Some of these substitutions are conservative and matched paralogous peroxidases in other Brassicaceae species. These results suggest that after duplication, peroxidase genes evolved under the pressure of positive selection, and amino acid substitutions identified during our study provided divergence of properties and physiological functions in peroxidases. Our predictions regarding functional significance for amino acid residues identified in variable sites of peroxidases may allow further experimental assessment of evolution of peroxidases after gene duplication.
Role of restriction-modification systems in prokaryotic evolution and ecology by A. S. Ershova; I. S. Rusinov; S. A. Spirin; A. S. Karyagina; A. V. Alexeevski (1373-1386).
Restriction–modification (R-M) systems are able to methylate or cleave DNA depending on methylation status of their recognition site. It allows them to protect bacterial cells from invasion by foreign DNA. Comparative analysis of a large number of available bacterial genomes and methylomes clearly demonstrates that the role of R-M systems in bacteria is wider than only defense. R-M systems maintain heterogeneity of a bacterial population and are involved in adaptation of bacteria to change in their environmental conditions. R-M systems can be essential for host colonization by pathogenic bacteria. Phase variation and intragenomic recombinations are sources of the fast evolution of the specificity of R-M systems. This review focuses on the influence of R-M systems on evolution and ecology of prokaryotes.
Level of blood cell-free circulating mitochondrial DNA as a novel biomarker of acute myocardial ischemia by N. P. Sudakov; T. P. Popkova; A. I. Katyshev; O. A. Goldberg; S. B. Nikiforov; B. G. Pushkarev; I. V. Klimenkov; S. A. Lepekhova; K. A. Apartsin; G. A. Nevinsky; Yu. M. Konstantinov (1387-1392).
Changes in the level of blood cell-free circulating mitochondrial DNA were examined during experimental adrenaline-induced myocardial injury in rats. The amount of mitochondrial DNA in the blood was significantly elevated at 48 and 72 h after subcutaneous injection of adrenaline solution, and it was accompanied by development of multiple smallfocal myocardial ischemia. This suggests that the measured level of blood cell-free circulating mitochondrial DNA might be used as a biomarker of acute myocardial ischemia.

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