Vol 14, No 1 (2022)

Protonophores are compounds capable of electrogenic transport of protons across membranes. Protonophores have been intensively studied over the past 50 years owing to their ability to uncouple oxidation and phosphorylation in mitochondria and chloroplasts. The action mechanism of classical uncouplers, such as DNP and CCCP, in mitochondria is believed to be related to their protonophoric activity; i.e., their ability to transfer protons across the lipid part of the mitochondrial membrane. Given the recently revealed deviations in the correlation between the protonophoric activity of some uncouplers and their ability to stimulate mitochondrial respiration, this review addresses the involvement of some proteins of the inner mitochondrial membrane, such as the ATP/ADP antiporter, dicarboxylate carrier, and ATPase, in the uncoupling process. However, these deviations do not contradict the Mitchell theory but point to a more complex nature of the interaction of DNP, CCCP, and other uncouplers with mitochondrial membranes. Therefore, a detailed investigation of the action mechanism of uncouplers is required for a more successful pharmacological use, including their antibacterial, antiviral, anticancer, as well as cardio-, neuro-, and nephroprotective effects.

The present review examines the use of chemiluminescence detection to evaluate the course of free radical reactions in biological model systems. The application of the method is analyzed by using luminescent additives that enhance the luminescence thanks to a triplet–singlet transfer of the electron excitation energy from radical reaction products and its emission in the form of light with a high quantum yield; these additives are called chemiluminescence enhancers or activators. Examples of these substances are provided; differences between the so-called chemical and physical enhancers are described; coumarin derivatives, as the most promising chemiluminescence enhancers for studying lipid peroxidation, are considered in detail. The main problems related to the use of coumarin derivatives are defined, and possible ways of solving these problems are presented. Intrinsic chemiluminescence and the mechanism of luminescence accompanying biomolecule peroxidation are discussed in the first part of the review.

The use of traditional tools for the targeted delivery of nanostructures, such as antibodies, transferrin, lectins, or aptamers, often leads to an entire range of undesirable effects. The large size of antibodies often does not allow one to reach the required number of molecules on the surface of nanostructures during modification, and the constant domains of heavy chains, due to their effector functions, can induce phagocytosis. In the recent two decades, targeted polypeptide scaffold molecules of a non-immunoglobulin nature, antibody mimetics, have emerged as much more effective targeting tools. They are small in size (3–20 kDa), possess high affinity (from subnano- to femtomolar binding constants), low immunogenicity, and exceptional thermodynamic stability. These molecules can be effectively produced in bacterial cells, and, using genetic engineering manipulations, it is possible to create multispecific fusion proteins for the targeting of nanoparticles to cells with a given molecular portrait, which makes scaffold polypeptides an optimal tool for theranostics.

Tandem exon duplications play an important role in the evolution of eukaryotic genes, providing a generic mechanism for adaptive regulation of protein function. In recent studies, tandem exon duplications have been linked to mutually exclusive exon choice, a pattern of alternative splicing in which one and only one exon from a group of tandemly arranged exons is included in the mature transcript. Here, we revisit the problem of identifying tandem exon duplications in eukaryotic genomes using bioinformatic methods and show that tandemly duplicated exons are abundant not only in the coding parts, but also in the untranslated regions. We present a number of remarkable examples of tandem exon duplications, identify unannotated duplicated exons, and provide statistical support for their expression using large panels of RNA-seq experiments.

The aim of this work is to develop a 3D cell culture model based on cell spheroids for predicting the functional activity of various compounds in vivo. Agarose gel molds were made using 3D printing. The solidified agarose gel is a matrix consisting of nine low-adhesive U-shaped microwells of 2.3 × 3.3 mm for 3D cell spheroid formation and growth. This matrix is placed into a single well of a 12-well plate. The effectiveness of the cell culture method was demonstrated using human ovarian carcinoma SKOVip-kat cells stably expressing the red fluorescent protein Katushka in the cytoplasm and overexpressing the membrane-associated tumor marker HER2. The SKOVip-kat cell spheroids were visualized by fluorescence microscopy. The cell concentration required for the formation of same-shape and same-size spheroids with tight intercellular contacts was optimized. To verify the developed model, the cytotoxicity of the targeted immunotoxin anti-HER2 consisting of the anti-HER2 scaffold DARP 9_29 and a fragment of the Pseudomonas aeroginosa exotoxin, DARP-LoPE, was studied in 2D and 3D SKOVip-kat cell cultures. The existence of a difference in the cytotoxic properties of DARP-LoPE between the 2D and 3D cultures has been demonstrated: the IC50 value in the 3D culture is an order of magnitude higher than that in the monolayer culture. The present work describes a universal tool for 3D cultivation of mammalian cells based on reusable agarose gel molds that allows for reproducible formation of multicellular spheroids with tight contacts for molecular and cell biology studies.

A breakthrough in cattle breeding was achieved with the incorporation of animal genomic data into breeding programs. The introduction of genomic selection has a major impact on traditional genetic assessment systems and animal genetic improvement programs. Since 2010, genomic selection has been officially introduced in the evaluation of the breeding and genetic potential of cattle in Europe, the U.S., Canada, and many other developed countries. The purpose of this study is to develop a system for a genomic evaluation of the breeding value of the domestic livestock of Black-and-White and Russian Holstein cattle based on 3 milk performance traits: daily milk yield (kg), daily milk fat (%), and daily milk protein content (%) and 6 fertility traits: age at first calving (AFC), calving interval (CI), calving to first insemination interval (CFI), interval between first and last insemination (IFL), days open (DO), and number of services (NS). We built a unified database of breeding animals from 523 breeding farms in the Russian Federation. The database included pedigree information on 2,551,529 cows and 69,131 bulls of the Russian Holstein and Black-and-White cattle breeds, as well as information on the milk performance of 1,597,426 cows with 4,771,366 completed lactations. The date of birth of the animals included in the database was between 1975 and 2017. Genotyping was performed in 672 animals using a BovineSNP50 v3 DNA Analysis BeadChip microarray (Illumina, USA). The genomic estimated breeding value (GEBV) was evaluated only for 644 animals (427 bulls and 217 cows) using the single-step genomic best linear unbiased prediction – animal model (ssGBLUP-AM). The mean genetic potential was +0.88 and +1.03 kg for the daily milk yield, -0.002% for the milk fat content, and –0.003 and 0.001% for the milk protein content in the cows and bulls, respectively. There was negative genetic progress in the fertility traits in the studied population between 1975 and 2017. The reliability of the estimated breeding value (EBV) for genotyped bulls ranged from 89 to 93% for the milk performance traits and 85 to 90% for the fertility traits, whereas the reliability of the genomic estimated breeding value (GEBV) varied 54 to 64% for the milk traits and 23 to 60% for the fertility traits. This result shows that it is possible to use the genomic estimated breeding value with rather high reliability to evaluate the domestic livestock of Russian Holstein and Black-and-White cattle breeds for fertility and milk performance traits. This system of genomic evaluation may help bring domestic breeding in line with modern competitive practices and estimate the breeding value of cattle at birth based on information on the animal’s genome.