Search Results (35)

Some Cretaceous ants belonging to the stem group of Formicidae exhibit bizarre morphology. This wide range of unusual adaptive features is primarily related to the mouthparts and clypeus. The researchers were perplexed by their specific ecology, as modern ant lineages do not exhibit anything similar. Here, we report and describe a new genus based on an extraordinary and mysterious alate ant from Late Eocene Baltic amber. Undoubtedly, the new ant is classified within the subfamily Formicinae (one of the crown groups), yet it displays a highly specialised morphology and an unusual array of features that are not observed in any extant ant lineages. Neither recent nor extinct ants have such a combination of features. While the exact phylogenetic placement of the new ant remains uncertain, we offer a discussion of its potential affinities based on our constrained phylogenetic analyses. We propose that †Eridanomyrma gen. n. should be considered in the new tribe †Eridanomyrmini trib. n. This new taxon highlights the adaptive diversity of a highly specialised, extinct lineage of Eocene crown-group ants. We also present a 3D model based on X-ray computed microtomography (µCT). Full article

In this paper, we discuss the prediction of the delivery efficiency of magnetic carriers based on their properties and field parameters. We developed a theory describing the behavior of magnetic capsules in the capillaries of living systems. A partial differential equation for the spatial distribution of magnetic capsules has been obtained. We propose to characterize the interaction between the magnetic field and the capsules using a single vector, which we call “specific magnetic force”. To test our theory, we performed experiments on a model of a capillary bed and on a living organism with two types of magnetic capsules that differ in size and amount of magnetic material. The experimental results show that the distribution of the capsules in the field correlated with the theory, but there were fewer actually accumulated capsules than predicted by the theory. In the weaker fields, the difference was more significant than in stronger ones. We proposed an explanation for this phenomenon based on the assumption that a certain level of magnetic force is needed to keep the capsules close to the capillary wall. We also suggested a formula for the relationship between the probability of capsule precipitation and the magnetic force. We found the effective value of a specific magnetic force at which all the capsules attracted by the magnet reach the capillary wall. This value can be considered as the minimum level for the field at which it is, in principle, possible to achieve a significant magnetic control effect. We demonstrated that for each type of capsule, there is a specific radius of magnet for which the effective magnetic force is achieved at the largest possible distance from the magnet’s surface. For the capsules examined in this study, the maximum distance where the effective field can be achieved does not exceed 1.5 cm. The results of the study contribute to our understanding of the behavior of magnetic particles in the capillaries of living organisms when exposed to a magnetic field. Full article

The apurinic/apyrimidinic site (AP site) is a highly mutagenic and cytotoxic DNA lesion. Normally, AP sites are removed from DNA by base excision repair (BER). Methoxyamine (MOX), a BER inhibitor currently under clinical trials as a tumor sensitizer, forms adducts with AP sites (AP-MOX) resistant to the key BER enzyme, AP endonuclease. As AP-MOX remains unrepaired, translesion DNA synthesis is expected to be the main mechanism of cellular response to this lesion. However, the mutagenic potential of AP-MOX is still unclear. Here, we compare the blocking and mutagenic properties of AP-MOX and the natural AP site for major eukaryotic DNA polymerases involved in translesion synthesis: DNA polymerases η, ι, ζ, Rev1, and primase–polymerase PrimPol. The miscoding properties of both abasic lesions remained mostly the same for each studied enzyme. In contrast, the blocking properties of AP-MOX compared to the AP site were DNA polymerase specific. Pol η and PrimPol bypassed both lesions with the same efficiency. The bypass of AP-MOX by Pol ι was 15-fold lower than that of the AP site. On the contrary, Rev1 bypassed AP-MOX 5-fold better than the AP site. Together, our data suggest that Rev1 is best suited to support synthesis across AP-MOX in human cells. Full article

Apurinic/apyrimidinic (AP) sites are endogenous DNA lesions widespread in human cells. Having no nucleobases, they are noncoding and promutagenic. AP site repair is generally initiated through strand incision by AP endonuclease 1 (APE1). Although AP sites’ repair in regular B-DNA has been studied extensively, their processing in G-quadruplexes (G4s) has received much less attention. Here, we used the hTERT promoter region that is capable of forming three stacked parallel G4s to understand how AP sites can influence higher-order quadruplex folding and stability and how a G4 affects the efficiency of human APE1-mediated AP site processing. We designed a series of synthetic single- and double-stranded DNA constructs of varying lengths containing a stable AP site analog in both G- and C-rich strands at positions corresponding to somatic driver mutations. Using circular dichroism, we studied the effect of the AP site on hTERT G4 structure and stability. Bio-layer interferometry and gel-based approaches were employed to characterize APE1 binding to the designed DNA substrates and AP site processing. It was shown that (i) an AP site leads to G4 destabilization, which depends on the lesion location in the G4 scaffold; (ii) APE1 binds tightly to hTERT G4 structure but exhibits greatly reduced cleavage activity at AP sites embedded in the quadruplex; and (iii) a clear correlation was revealed between AP site-induced hTERT G4 destabilization and APE1 activity. We can hypothesize that reduced repair of AP sites in the hTERT G4 is one of the reasons for the high mutation rate in this promoter region. Full article

The design of controllable and precise RNA-targeted CRISPR/Cas9 (Clustered Regularly Interspaced Short Palindromic Repeats) systems is an important problem of modern molecular biology and genetic technology. Herein, we have designed a series of photocleavable guide CRISPR RNAs (crRNA) and their 2′-modified (2′-fluoro and locked nucleic acid) analogs containing one or two 1-(2-nitrophenyl)-1,2-ethanediol photolabile linkers (PL). We have demonstrated that these crRNAs can be destroyed by relatively mild UVA irradiation with the rate constants 0.24–0.77 min⁻¹ and that the photocleavage markedly slows down the action of Cas9 nuclease in the model in vitro system. Two PLs provide more rapid crRNA destruction than a single linker. PLs in the crRNA structure improve the specificity of DNA cleavage by Cas9 nuclease for the fully complementary target. The application of photocleavable crRNA in CRISPR/Cas9 genome editing permits the system to be switched off in a spatiotemporally controlled manner, thus alleviating its off-target effects. Full article

Clickable nucleosides, most often 5-ethynyl-2′-deoxyuridine (EtU), are widely used in studies of DNA replication in living cells and in DNA functionalization for bionanotechology applications. Although clickable dNTPs are easily incorporated by DNA polymerases into the growing chain, afterwards they might become targets for DNA repair systems or interfere with faithful nucleotide insertion. Little is known about the possibility and mechanisms of these post-synthetic events. Here, we investigated the repair and (mis)coding properties of EtU and two bulkier clickable pyrimidine nucleosides, 5-(octa-1,7-diyn-1-yl)-U (C8-AlkU) and 5-(octa-1,7-diyn-1-yl)-C (C8-AlkC). In vitro, EtU and C8-AlkU, but not C8-AlkC, were excised by SMUG1 and MBD4, two DNA glycosylases from the base excision repair pathway. However, when placed into a plasmid encoding a fluorescent reporter inactivated by repair in human cells, EtU and C8-AlkU persisted for much longer than uracil or its poorly repairable phosphorothioate-flanked derivative. DNA polymerases from four different structural families preferentially bypassed EtU, C8-AlkU and C8-AlkC in an error-free manner, but a certain degree of misincorporation was also observed, especially evident for DNA polymerase β. Overall, clickable pyrimidine nucleotides could undergo repair and be a source of mutations, but the frequency of such events in the cell is unlikely to be considerable. Full article

The DNA building blocks 2′-deoxynucleotides are enantiomeric, with their natural β-D-configuration dictated by the sugar moiety. Their synthetic β-L-enantiomers (βLdNs) can be used to obtain L-DNA, which, when fully substituted, is resistant to nucleases and is finding use in many biosensing and nanotechnology applications. However, much less is known about the enzymatic recognition and processing of individual βLdNs embedded in D-DNA. Here, we address the template properties of βLdNs for several DNA polymerases and the ability of base excision repair enzymes to remove these modifications from DNA. The Klenow fragment was fully blocked by βLdNs, whereas DNA polymerase κ bypassed them in an error-free manner. Phage RB69 DNA polymerase and DNA polymerase β treated βLdNs as non-instructive but the latter enzyme shifted towards error-free incorporation on a gapped DNA substrate. DNA glycosylases and AP endonucleases did not process βLdNs. DNA glycosylases sensitive to the base opposite their cognate lesions also did not recognize βLdNs as a correct pairing partner. Nevertheless, when placed in a reporter plasmid, pyrimidine βLdNs were resistant to repair in human cells, whereas purine βLdNs appear to be partly repaired. Overall, βLdNs are unique modifications that are mostly non-instructive but have dual non-instructive/instructive properties in special cases. Full article

Numerous studies have shown that oxidative modifications of guanine (7,8-dihydro-8-oxoguanine, 8-oxoG) can affect cellular functions. 7,8-Dihydro-8-oxoadenine (8-oxoA) is another abundant paradigmatic ambiguous nucleobase but findings reported on the mutagenicity of 8-oxoA in bacterial and eukaryotic cells are incomplete and contradictory. Although several genotoxic studies have demonstrated the mutagenic potential of 8-oxoA in eukaryotic cells, very little biochemical and bioinformatics data about the mechanism of 8-oxoA-induced mutagenesis are available. In this review, we discuss dual coding properties of 8-oxoA, summarize historical and recent genotoxicity and biochemical studies, and address the main protective cellular mechanisms of response to 8-oxoA. We also discuss the available structural data for 8-oxoA bypass by different DNA polymerases as well as the mechanisms of 8-oxoA recognition by DNA repair enzymes. Full article

Apurinic/apyrimidinic (AP) sites are abundant DNA lesions generated both by spontaneous base loss and as intermediates of base excision DNA repair. In human cells, they are normally repaired by an essential AP endonuclease, APE1, encoded by the APEX1 gene. Other enzymes can cleave AP sites by either hydrolysis or β-elimination in vitro, but it is not clear whether they provide the second line of defense in living cells. Here, we studied AP site repairs in APEX1 knockout derivatives of HEK293FT cells using a reporter system based on transcriptional mutagenesis in the enhanced green fluorescent protein gene. Despite an apparent lack of AP site-processing activity in vitro, the cells efficiently repaired the tetrahydrofuran AP site analog resistant to β-elimination. This ability persisted even when the second AP endonuclease homolog, APE2, was also knocked out. Moreover, APEX1 null cells were able to repair uracil, a DNA lesion that is removed via the formation of an AP site. If AP site hydrolysis was chemically blocked, the uracil repair required the presence of NTHL1, an enzyme that catalyzes β-elimination. Our results suggest that human cells possess at least two back-up AP site repair pathways, one of which is NTHL1-dependent. Full article

Lipophilic oligonucleotide derivatives are a potent approach to the intracellular delivery of nucleic acids. The binding of these derivatives to serum albumin is a determinant of their fate in the body, as its structure contains several sites of high affinity for hydrophobic compounds. This study focuses on the features of self-association and non-covalent interactions with human serum albumin of novel self-penetrating oligonucleotide derivatives. The study revealed that the introduction of a triazinyl phosphoramidate modification bearing two dodecyl groups at the 3′ end region of the oligonucleotide sequence has a negligible effect on its affinity for the complementary sequence. Dynamic light scattering verified that the amphiphilic oligonucleotides under study can self-assemble into micelle-like particles ranging from 8 to 15 nm in size. The oligonucleotides with dodecyl groups form stable complexes with human serum albumin with a dissociation constant of approximately 10⁻⁶ M. The oligonucleotide micelles are simultaneously destroyed upon binding to albumin. Using an electrophoretic mobility shift assay and affinity modification, we examined the ability of DNA duplexes containing triazinyl phosphoramidate oligonucleotides to interact with Ku antigen and PARP1, as well as the mutual influence of PARP1 and albumin or Ku antigen and albumin upon interaction with DNA duplexes. These findings, together with the capability of dodecyl-containing derivatives to effectively penetrate different cells, such as HEK293 and T98G, indicate that the oligonucleotides under study can be considered as a platform for the development of therapeutic preparations with a target effect. Full article

Immune responses to tissue-engineered grafts made of xenogeneic materials remain poorly studied. The scope of current investigations is limited by the lack of information on orthotopically implanted grafts. A deeper understanding of these processes is of great importance since innovative surgical approaches include the implantation of xenogeneic decellularized scaffolds seeded by cells. The purpose of our work is to study the immunological features of tracheal repair during the implantation of tissue-engineered constructs based on human xenogeneic scaffolds modified via laser radiation in rabbits. The samples were stained with hematoxylin and Safranin O, and they were immunostained with antibodies against tryptase, collagen II, vimentin, and CD34. Immunological and inflammatory responses were studied by counting immune cells and evaluating blood vessels and collagen. Leukocyte-based inflammation prevailed during the implantation of decellularized unseeded scaffolds; meanwhile, plasma cells were significantly more abundant in tissue-engineered constructs. Mast cells were insignificantly more abundant in tissue-engineered construct samples. Conclusions: The seeding of decellularized xenogeneic cartilage with chondrocytes resulted in a change in immunological reactions upon implantation, and it was associated with plasma cell infiltration. Tissue-engineered grafts widely differed in design, including the type of used cells. The question of immunological response depending on the tissue-engineered graft composition requires further investigation. Full article

Base excision DNA repair (BER) is a key pathway safeguarding the genome of all living organisms from damage caused by both intrinsic and environmental factors. Most present knowledge about BER comes from studies of human cells, E. coli, and yeast. Plants may be under an even heavier DNA damage threat from abiotic stress, reactive oxygen species leaking from the photosynthetic system, and reactive secondary metabolites. In general, BER in plant species is similar to that in humans and model organisms, but several important details are specific to plants. Here, we review the current state of knowledge about BER in plants, with special attention paid to its unique features, such as the existence of active epigenetic demethylation based on the BER machinery, the unexplained diversity of alkylation damage repair enzymes, and the differences in the processing of abasic sites that appear either spontaneously or are generated as BER intermediates. Understanding the biochemistry of plant DNA repair, especially in species other than the Arabidopsis model, is important for future efforts to develop new crop varieties. Full article

DNA–protein cross-links remain the least-studied type of DNA damage. Recently, their repair was shown to involve proteolysis; however, the fate of the peptide remnant attached to DNA is unclear. Particularly, peptide cross-links could interfere with DNA polymerases. Apurinuic/apyrimidinic (AP) sites, abundant and spontaneously arising DNA lesions, readily form cross-links with proteins. Their degradation products (AP site–peptide cross-links, APPXLs) are non-instructive and should be even more problematic for polymerases. Here, we address the ability of human DNA polymerases involved in DNA repair and translesion synthesis (POLβ, POLλ, POLη, POLκ and PrimPOL) to carry out synthesis on templates containing AP sites cross-linked to the N-terminus of a 10-mer peptide (APPXL-I) or to an internal lysine of a 23-mer peptide (APPXL-Y). Generally, APPXLs strongly blocked processive DNA synthesis. The blocking properties of APPXL-I were comparable with those of an AP site, while APPXL-Y constituted a much stronger obstruction. POLη and POLκ demonstrated the highest bypass ability. DNA polymerases mostly used dNTP-stabilized template misalignment to incorporate nucleotides when encountering an APPXL. We conclude that APPXLs are likely highly cytotoxic and mutagenic intermediates of AP site–protein cross-link repair and must be quickly eliminated before replication. Full article

As the world’s population continues to increase, ensuring food security becomes a major problem. This often leads to the expansion of agricultural production, even in harsh conditions and becomes a key problem for many countries, including Russia. However, such expansion may entail certain costs, including the potential loss of insect populations, which are vital for ecological balance and agricultural productivity. The development of fallow lands in these regions is necessary to increase food production and increase food security; it is important to balance this with protection from harmful insects and sustainable farming methods. Research into the effects of insecticides on insects is an ongoing challenge, and new, sustainable farming methods are needed to ensure that protection from harmful insects and sustainable development can coexist. This article discusses the use of pesticides to protect the well-being of mankind, the problems of studying the effects of pesticides on insects and the vulnerability of insects to pesticides in regions with harsh conditions. It also discusses successful methods of sustainable agriculture and the importance of the legal framework governing the use of pesticides. The article emphasises the importance of balanced development with insect protection to ensure the sustainability of agricultural expansion in harsh conditions. Full article

The protein encoded by the vaccinia virus D4R gene has base excision repair uracil–DNA N-glycosylase (vvUNG) activity and also acts as a processivity factor in the viral replication complex. The use of a protein unlike PolN/PCNA sliding clamps is a unique feature of orthopoxviral replication, providing an attractive target for drug design. However, the intrinsic processivity of vvUNG has never been estimated, leaving open the question whether it is sufficient to impart processivity to the viral polymerase. Here, we use the correlated cleavage assay to characterize the translocation of vvUNG along DNA between two uracil residues. The salt dependence of the correlated cleavage, together with the similar affinity of vvUNG for damaged and undamaged DNA, support the one-dimensional diffusion mechanism of lesion search. Unlike short gaps, covalent adducts partly block vvUNG translocation. Kinetic experiments show that once a lesion is found it is excised with a probability ~0.76. Varying the distance between two uracils, we use a random walk model to estimate the mean number of steps per association with DNA at ~4200, which is consistent with vvUNG playing a role as a processivity factor. Finally, we show that inhibitors carrying a tetrahydro-2,4,6-trioxopyrimidinylidene moiety can suppress the processivity of vvUNG. Full article