Vol 8, No 4 (2016)


Peculiarities of Yeasts and Human Telomerase RNAs Processing

Rubtsova M.P., Vasilkova D.P., Naraykina Y.V., Dontsova O.A.


Telomerase is one of the major components of the telomeres -- linear eukaryotic chromosome ends - maintenance system. Linear chromosomes are shortened during each cell division due to the removal of the primer used for DNA replication. Special repeated telomere sequences at the very ends of linear chromosomes prevent the deletion of genome information caused by primer removal. Telomeres are shortened at each replication round until it becomes critically short and is no longer able to protect the chromosome in somatic cells. At this stage, a cell undergoes a crisis and usually dies. Rare cases result in telomerase activation, and the cell gains unlimited proliferative capacity. Special types of cells, such as stem, germ, embryonic cells and cells from tissues with a high proliferative potential, maintain their telomerase activity indefinitely. The telomerase is inactive in the majority of somatic cells. Telomerase activity in vitro requires two key components: telomerase reverse transcriptase and telomerase RNA. In cancer cells, telomerase reactivates due to the expression of the reverse transcriptase gene. Telomerase RNA expresses constitutively in the majority of human cells. This fact suggests that there are alternative functions to telomerase RNA that are unknown at the moment. In this manuscript, we review the biogenesis of yeasts and human telomerase RNAs thanks to breakthroughs achieved in research on telomerase RNA processing by different yeasts species and humans in the last several years.

Acta Naturae. 2016;8(4):14-22
pages 14-22 views

Therapy of HIV Infection: Current Approaches and Prospects

Prokofjeva M.M., Kochetkov S.N., Prassolov V.S.


The human immunodeficiency virus type 1 (HIV-1) is the causative agent of one of the most dangerous human diseases - the acquired immune deficiency syndrome (AIDS). Over the past 30 years since the discovery of HIV-1, a number of antiviral drugs have been developed to suppress various stages of the HIV-1 life cycle. This approach has enables the suppression of virus replication in the body, which significantly prolongs the life of HIV patients. The main downside of the method is the development of viral resistance to many anti-HIV drugs, which requires the creation of new drugs effective against drug-resistant viral forms. Currently, several fundamentally new approaches to HIV-1 treatment are under development, including the use of neutralizing antibodies, genome editing, and blocking an integrated latent provirus. This review describes a traditional approach involving HIV-1 inhibitors as well as the prospects of other treatment options.

Acta Naturae. 2016;8(4):23-32
pages 23-32 views

Synthetic Fluorophores for Visualizing Biomolecules in Living Systems

Martynov V.I., Pakhomov A.A., Popova N.V., Deyev I.E., Petrenko A.G.


The last decade has witnessed significant advance in the imaging of living systems using fluorescent markers. This progress has been primarily associated with the discovery of different spectral variants of fluorescent proteins. However, the fluorescent protein technology has its own limitations and, in some cases, the use of low-molecular-weight fluorophores is preferable. In this review, we describe the arsenal of synthetic fluorescent tools that are currently in researchers’ hands and span virtually the entire spectrum, from the UV to visible and, further, to the near-infrared region. An overview of recent advances in site-directed introduction of synthetic fluorophores into target cellular objects is provided. Application of these fluorescent probes to the solution of a wide range of biological problems, in particular, to the determination of local ion concentrations and pH in living systems, is discussed.

Acta Naturae. 2016;8(4):33-46
pages 33-46 views

From Slow to Fast: Hypogravity-Induced Remodeling of Muscle Fiber Myosin Phenotype

Shenkman B.S.


Skeletal muscle consists of different fiber types arranged in a mosaic pattern. These fiber types are characterized by specific functional properties. Slow-type fibers demonstrate a high level of fatigue resistance and prolonged contraction duration, but decreased maximum contraction force and velocity. Fast-type fibers demonstrate a high contraction force and velocity, but profound fatigability. During the last decades, it has been discovered that all these properties are determined by the predominance of slow or fast myosin-heavy-chain (MyHC) isoforms. It was observed that gravitational unloading during space missions and simulated microgravity in ground-based experiments leads to the transformation of some slow-twitch muscle fibers into fast-twitch ones due to changes in the patterns of MyHC gene expression in the postural soleus muscle. The present review covers the facts and mechanistic speculations regarding myosin phenotype remodeling under conditions of gravitational unloading. The review considers the neuronal mechanisms of muscle fiber control and molecular mechanisms of regulation of myosin gene expression, such as inhibition of the calcineurin/NFATc1 signaling pathway, epigenomic changes, and the behavior of specific microRNAs. In the final portion of the review, we discuss the adaptive role of myosin phenotype transformations.

Acta Naturae. 2016;8(4):47-59
pages 47-59 views

The Role of BAR Domain Proteins in the Regulation of Membrane Dynamics

Stanishneva-Konovalova T.B., Derkacheva N.I., Polevova S.V., Sokolova O.S.


Many cellular processes are associated with membrane remodeling. The BAR domain protein family plays a key role in the formation and detection of local membrane curvatures and in attracting other proteins, including the regulators of actin dynamics. Based on their structural and phylogenetic properties, BAR domains are divided into several groups which affect membrane in various ways and perform different functions in cells. However, recent studies have uncovered evidence of functional differences even within the same group. This review discusses the principles underlying the interactions of different groups of BAR domains, and their individual representatives, with membranes.

Acta Naturae. 2016;8(4):60-69
pages 60-69 views

Design of Stable α-Helical Peptides and Thermostable Proteins in Biotechnology and Biomedicine

Yakimov A.P., Afanaseva A.S., Khodorkovskiy M.A., Petukhov M.G.


α-Heliсes are the most frequently occurring elements of the secondary structure in water-soluble globular proteins. Their increased conformational stability is among the main reasons for the high thermal stability of proteins in thermophilic bacteria. In addition, α-helices are often involved in protein interactions with other proteins, nucleic acids, and the lipids of cell membranes. That is why the highly stable α-helical peptides used as highly active and specific inhibitors of protein-protein and other interactions have recently found more applications in medicine. Several different approaches have been developed in recent years to improve the conformational stability of α-helical peptides and thermostable proteins, which will be discussed in this review. We also discuss the methods for improving the permeability of peptides and proteins across cellular membranes and their resistance to intracellular protease activity. Special attention is given to the SEQOPT method (http://mml.spbstu.ru/services/seqopt/), which is used to design conformationally stable short α-helices.

Acta Naturae. 2016;8(4):70-81
pages 70-81 views

Research Articles

A Cascade of Thermophilic Enzymes As an Approach to the Synthesis of Modified Nucleotides

Esipov R.S., Abramchik Y.A., Fateev I.V., Konstantinova I.D., Kostromina M.A., Muravyova T.I., Artemova K.G., Miroshnikov A.I.


We propose a new approach for the synthesis of biologically important nucleotides which includes a multi-enzymatic cascade conversion of D-pentoses into purine nucleotides. The approach exploits nucleic acid exchange enzymes from thermophilic microorganisms: ribokinase, phosphoribosylpyrophosphate synthetase, and adenine phosphoribosyltransferase. We cloned the ribokinase gene from Thermus sp. 2.9, as well as two different genes of phosphoribosylpyrophosphate synthetase (PRPP-synthetase) and the adenine phosphoribosyltransferase (APR-transferase) gene from Thermus thermophilus HB27 into expression vectors, generated high-yield E. coli producer strains, developed methods for the purification of the enzymes, and investigated enzyme substrate specificity. The enzymes were used for the conversion of D-pentoses into 5-phosphates that were further converted into 5-phospho-α-D-pentofuranose 1-pyrophosphates by means of ribokinase and PRPP-synthetases. Target nucleotides were obtained through the condensation of the pyrophosphates with adenine and its derivatives in a reaction catalyzed by APR-transferase. 2-Chloro- and 2-fluoroadenosine monophosphates were synthesized from D-ribose and appropriate heterobases in one pot using a system of thermophilic enzymes in the presence of ATP, ribokinase, PRPP-synthetase, and APR-transferase.

Acta Naturae. 2016;8(4):82-90
pages 82-90 views

Downregulation of Purkinje Cell Activity by Modulators of Small Conductance Calcium-Activated Potassium Channels In Rat Cerebellum

Karelina T.V., Stepanenko Y.D., Abushik P.A., Sibarov D.A., Antonov S.M.


Small-conductance calcium-activated potassium channels (SK channels) are widely expressed in CNS tissues. Their functions, however, have not been well studied. Participation of SK channels in Purkinje cell (PC) pacemaker activity has been studied predominantly in vitro. Here we studied for the first time the effects of SK channel activation by NS309 or CyPPA on the PC simple spike frequency in vivo in adult (3 - 6 months) and aged (22 - 28 months) rats using extracellular microelectrode recordings. Both pharmacological agents caused a statistically significant decrease in the PC simple spike frequency. The maximum value of the decrease in the simple spike frequency did not depend on age, whereas a statistically significant inhibition of the spike frequency was achieved faster in aged animals than in adult ones. In experiments on cultured neurons PCs were identified by the expression of calbindin as the PC-specific marker. Registration of transmembrane currents in cerebellar neurons revealed the direct action of NS309 and CyPPA on the SK channels of PC consisted in the enhancement of outward potassium currents and action potential after-hyperpolarization. Thus, SK channel activators can compensate for age-related changes of the autorhythmic functions of the cerebellum.

Acta Naturae. 2016;8(4):91-99
pages 91-99 views

Evolution of Tumor Clones in Adult Acute Lymphoblastic Leukemia

Smirnova S.Y., Sidorova Y.V., Ryzhikova N.V., Sychevskaya K.A., Parovichnikova E.N., Sudarikov A.B.


Clonal instability of a tumor cell population in acute lymphoblastic leukemia (ALL) may complicate the monitoring of a minimal residual disease (MRD) by means of patient-specific targets identified at the disease onset. Most of the data concerning the possible instability of rearranged clonal TCR and IG genes during disease recurrence were obtained for ALL in children. The appropriate features of adult ALL, which are known to differ from those of childhood ALL in certain biological characteristics and prognosis, remain insufficiently studied. The aim of this study was to assess the stability of IG and TCR gene rearrangements in adult ALL. Rearrangements were identified according to the BIOMED-2 protocol (PCR followed by fragment analysis). Mismatch in clonal rearrangements at onset and relapse was identified in 83% of patients, indicating clonal instability during treatment. Clonal evolution and diversity of IG and TCR gene rearrangements may be one of the tumor progression mechanisms. New rearrangements may emerge due to residual VDJ-recombinase activity in tumor cells. Also, many clonal IG and TCR gene rearrangements may be present at different levels at a diagnosis, but less abundant clones may be “invisible” due to limited detection sensitivity. Later, major clones may disappear in the course of chemotherapy, while others may proliferate. Investigation of clonal evolution and heterogeneity in ALL and their impact on the treatment efficacy will contribute to the identification of new prognostic factors and the development of therapeutic approaches.

Acta Naturae. 2016;8(4):100-109
pages 100-109 views

Adjuvant-Induced Arthritis in Guinea Pigs

Taranov O.S., Yakubitskiy S.N., Nepomnyashchikh T.S., Nesterov A.E., Shchelkunov S.N.


We propose a model of rheumatoid arthritis (RA) induced in outbred guinea pigs using a single subcutaneous injection of complete Freund’s adjuvant to the hind paw. Histological examination of this model shows fibrin deposition on the surface of the synovial membrane, leukocyte infiltration of the synovial membrane and adjacent tissues, proliferation of the granulation tissue, and emergence of angioid areas, characteristic of RA. The cell response appears as an increase in the plasma cell count and development of follicle-like lymphoid infiltrates; erosion of the articular surface of the cartilage, frequently with deep cartilage destruction over large areas; and epiphysiopathy. The high reproducibility of arthritis induction in this RA model has been demonstrated. The proposed model is promising for the assessment of anti-arthritis preparations and dosage regimens.

Acta Naturae. 2016;8(4):110-117
pages 110-117 views

Flavoprotein miniSOG Cytotoxisity Can Be Induced By Bioluminescence Resonance Energy Transfer

Shramova E.I., Proshkina G.M., Chumakov S.P., Khodarovich Y.M., Deyev S.M.


In this study, we investigated the possibility of phototoxic flavoprotein miniSOG (photosensitizer) excitation in cancer cells by bioluminescence occurring when luciferase NanoLuc oxidizes its substrate, furimazine. We have shown that the phototoxic flavoprotein miniSOG expressed in eukaryotic cells in fusion with NanoLuc luciferase is activated in the presence of its substrate, furimazine. Upon such condition, miniSOG possesses photoinduced cytotoxicity and causes a 48% cell death level in a stably transfected cell line.

Acta Naturae. 2016;8(4):118-123
pages 118-123 views


GMOs in Russia: Research, Society and Legislation

Korobko I.V., Georgiev P.G., Skryabin K.G., Kirpichnikov M.P.


Russian legislation lags behind the rapid developments witnessed in genetic engineering. Only a scientifically based and well-substantiated policy on the place of organisms that are created with the use of genetic engineering technologies and an assessment of the risks associated with them could guarantee that the breakthroughs achieved in modern genetic engineering technologies are effectively put to use in the real economy. A lack of demand for such breakthroughs in the practical field will lead to stagnation in scientific research and to a loss of expertise.

Acta Naturae. 2016;8(4):6-13
pages 6-13 views

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