Biochemistry (Moscow)最新文献

Long non-coding RNAs (lncRNAs) are a novel class of regulators of key cellular processes and biomarkers of various pathologies. The lncRNA JPX is a multifunctional RNA involved in the regulation of transcription, translation, and chromatin architecture. JPX influences transcription and enhancer-promoter communication by regulating binding of proteins to DNA, particularly by interacting with the chromatin architectural protein CTCF. Additionally, JPX can interact with microRNAs, repressor proteins, or mRNA stabilizers, regulating translation in pathogenesis of oncological and other diseases. This review summarizes the accumulated knowledge about the structure, evolutionary origin, and functions of the long non-coding RNA JPX in normal and pathological conditions.

Recent studies have highlighted the pivotal role of biomolecular condensates (liquid-like complexes) in gene control. Biomolecular condensates create a specific microenvironment around enhancers and gene promoters, which can activate transcription, repress it, or maintain at an appropriate level. They can also influence the chromatin structure and are important participants in the enhancer–promoter communication. Finally, biomolecular condensates represent promising therapeutic targets, as their dysregulation results in a broad spectrum of pathologies. The review present most recent, as well as fundamental studies establishing the role of condensates in the regulation of gene expression and enhancer–promoter communication.

Increasing resistance of human immunodeficiency virus type 1 (HIV-1) to the drugs targeting viral proteins stimulates the search for new therapeutic targets, among which are blockers of virus–host protein interactions. For two cellular proteins (LEDGF/p75 and Ku70) that interact with viral integrase, binding inhibitors have already been identified that reduce replication efficiency. Previously, using the methods of cross-linking and co-immunoprecipitation followed by mass spectrometry, several novel potential cellular partners of HIV-1 integrase were identified, including the Hsp60 chaperonin and S-adenosylhomocysteine hydrolase (SAHH). In the present study, we demonstrate that these purified recombinant proteins co-precipitate in vitro with integrase, indicating their ability to directly interact with the enzyme. Knockdown of Hsp60 and SAHH in the human cells was found to stimulate transduction efficiency by the HIV-1-based pseudovirus. This effect occurs specifically at the early stages of HIV-1 replication, not at the stage of proviral transcription. Furthermore, we were able to determine the stage of HIV-1 replication influenced by these proteins. It was revealed that the Hsp60 knockdown stimulates integration, while the SAHH knockdown enhances efficiency of the viral reverse transcription, in which integrase is also involved.

The rise of antimicrobial resistance among pathogenic bacteria poses a critical challenge to modern medicine, highlighting an urgent need for novel therapeutic agents. Bottromycin A₂ (BotA2) is a promising candidate for future drug development, demonstrating potent activity against clinically relevant pathogens, including methicillin-resistant Staphylococcus aureus, vancomycin-resistant Enterococcus, and Mycoplasma species, although its molecular mechanism of action has remained unclear until now. Here, we demonstrate that BotA2 inhibits bacterial translation with unique context specificity determined by the mRNA coding sequence. Using high-throughput toe-printing coupled with deep sequencing (Toe-seq analysis), we show that BotA2 induces ribosome pausing predominantly when a glycine codon enters the A-site of the ribosome, regardless of the codon identities in the P- and E-sites. Our biochemical and biophysical data indicate that BotA2 specifically arrests glycine-delivering ternary complexes on the ribosome, thereby preventing full accommodation of incoming Gly-tRNA^Gly within the peptidyl transferase center. Altogether, our findings uncover a previously undescribed mechanism of translation inhibition, driven by the context-specific immobilization of ternary complexes on elongating ribosomes.

Analysis of the genomes and transcriptomes of the beetle Tenebrio molitor revealed a group of six serine peptidase homologs (SPH) of the S1A chymotrypsin subfamily containing a conservative substitution of the catalytic residue Ser195 with Thr (Ser195Thr) in the active center. All six SPH are secreted proteins with prepropeptides and lack regulatory domains in the propeptide. The most highly expressed homolog, SerPH122, shares 57% sequence identity with the most highly expressed elastase-like peptidase of T. molitor, SerP41. Both proteins exhibit similar domain organization, localization in the posterior midgut, and expression patterns in the feeding stages of the fourth instar larva and imago. Testing hydrolytic activity of the recombinant rSerPH122 preparation demonstrated that the conservative substitution of Ser for Thr in the active center did not abolish its catalytic activity. rSerPH122 exhibits low specific activity but broad substrate specificity, most effectively hydrolyzing substrates of chymotrypsin-like and trypsin-like peptidases. The homolog has a pH optimum at 8.5 and is stable in the pH range 4.0-8.0. This study addresses the question of activity of the homologs with the Ser195Thr substitution and contributes to understanding of the poorly studied area of SPH functions, providing a basis for elucidating relationship between the structure and function of serine peptidases and their homologs.

Hepatitis B virus (Orthohepadnavirus hominoidei, HBV) is a hepatotropic virus from the Hepadnaviridae family and the causative agent of both acute and chronic hepatitis B (CHB). The possible outcomes of CHB include liver cirrhosis and hepatocellular carcinoma (HCC) that pose a significant burden on the healthcare systems worldwide. In the nuclei of infected hepatocytes of patients with CHB, the HBV genome persists as a pool of covalently closed circular DNA (cccDNA) molecules. Current therapeutic strategies cannot directly target cccDNA. Instead, the available treatments focus on long-term suppression of viral replication and require lifelong administration. Development and evaluation of novel antiviral agents capable of achieving complete HBV eradication require relevant in vivo and in vitro models of HBV infection. Among the available animal models, the following categories are distinguished: (i) animals naturally susceptible to HBV; (ii) surrogate models using animal species susceptible to the related hepadnaviruses; (iii) non-susceptible animals receiving HBV genome via recombinant viral vectors; (iv) models utilizing human hepatocyte xenografts. Among the available in vitro models, primary human and northern treeshrew (Tupaia belangeri) hepatocytes fully support the HBV replication cycle, but they rapidly lose susceptibility to the virus in cell culture. In turn, unmodified human hepatoma cell lines are not susceptible to HBV but can support viral replication after transfection with the viral genome. This review discusses key characteristics, advantages, limitations, and areas of application of the currently available in vivo and in vitro models of HBV infection.

The review discusses the functional role of the ribosomal E-site in the context of recent structural data. Traditionally, the E-site has been considered to serve only as a binding site for deacylated tRNA (E-tRNA) prior to its dissociation from the protein synthesis complex. Here, we examine specific contacts formed between E-tRNA and rRNA of the large ribosomal subunit in different organisms, as well as the sequence of their formation and disruption. The mechanism of translation suppression by inhibitors that bind to the ribosomal E-site is discussed. Based on current evidence regarding the location of aminoacyl-tRNA synthetases (ARSs) in the immediate vicinity of the ribosome, we propose a hypothesis that one of the primary functions of the ribosomal E-site is to prepare tRNA (through its modulation) for the formation of a specific complex with ARS, in the content of which it is released from the ribosome.

The review discusses main characteristics and analytical features of the circulating tumor DNA (ctDNA), which accounts for a minor fraction of the cell-free DNA (cfDNA) in cancer patients. Currently, ctDNA is considered to be a promising biomarker for assessing treatment efficacy, prognosis, and disease monitoring in oncology, including breast cancer (BC). A significant proportion of BC patients receive neoadjuvant drug therapy, effectiveness of which largely determines necessity and extent of subsequent treatment. Determination of ctDNA could be the most sensitive method for evaluating response to neoadjuvant therapy, as it enables real-time monitoring of molecular changes during the treatment, prediction of therapeutic response, and assessment of recurrence risk. This approach could become an additional tool for personalization of BC therapy.

Species of the genus Salicornia (Amaranthaceae s.l.) are globally distributed and highly salt-tolerant. They are used as food and for biofuel production. Formation of pure lines through self-pollination, combined with sporadic cross-pollination, polyploidy, high phenotypic plasticity, and a limited number of diagnostic characters, significantly complicates taxonomy of the genus. Salicornia (glassworts) is an evolutionarily young group, where the number of informative substitutions in the traditionally analyzed regions of nuclear and plastid DNA is insufficient to establish relationships between the species. The very concept of a species in this genus remains a subject of debate. To clarify relationships among the Eastern European species, we used high-throughput sequencing to determine sequences and perform phylogenetic analysis of the plastomes of 11 samples representing all major morphotypes of Eastern European glassworts. We also analyzed variability of the nuclear rDNA external transcribed spacer (nrETS). The sizes of the assembled plastomes ranged from 153,290 bp to 153,504 bp and exhibited a typical architecture with a large single-copy region (84,625-84,797 bp), a small single-copy region (18,818-18,870 bp), and two inverted repeats (24,898-24,908 bp). Comparison of phylogenetic trees reconstructed from all currently available plastome data and nrETS alignments of the same glasswort accessions revealed a discrepancy in the placement of the tetraploid S. procumbens subsp. pojarkovae and S. brachiata accessions, which show affinities to different lineages depending on the use of either plastid or nuclear (nrETS) data. Our results highlight the role of reticulate evolution in the genus Salicornia.

Two types of experiments are used to study RNA-chromatin interactions: the interactome search for individual RNAs (“one-to-all” or OTA) and genome-wide contact mapping for all RNAs (“all-to-all” or ATA). Comparative analysis of ATA and OTA data revealed fundamental differences in resolution, completeness, and specificity. OTA data exhibit high resolution (~1000 bp) and reproducibility (>90%), serving as a “gold standard”. ATA data, however, have lower resolution (~5000 bp), and their reproducibility (<10%) is critically dependent on the protocol, with two-step fixation using disuccinimidyl glutarate and formaldehyde (GRID-seq) showing a clear advantage over formaldehyde alone. The introduced “chromatin potential” metric and BaRDIC peak filtering effectively isolate the specific signal. This study proposes a strategy for reliable interactome analysis: combining RNA selection based on chromatin potential with the use of concordant contacts from peaks.