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Escherichia coli and its bacteriophages are among the most studied model microorganisms. Bacteriophages for various E. coli strains can typically be easily isolated from environmental sources, and many of these viruses can be harnessed to combat E. coli infections in humans and animals. However, some relatively rare E. coli strains pose significant challenges in finding suitable phages. The uropathogenic strain E. coli UPEC124, isolated from a patient suffering from neurogenic bladder dysfunction, was found to be resistant to all coliphages in our collections, and initial attempts to isolate new phages failed. Using an improved procedure for phage enrichment, we isolated the N4-related phage Mimir124, belonging to the Gamaleyavirus genus, which was able to lyse this "difficult" E. coli strain. Although Mimir124 is a narrow-spectrum phage, it was effective in the individualized treatment of the patient, leading to pathogen eradication. The primary receptor of Mimir124 was the O antigen of the O101 type; consequently, Mimir124-resistant clones were rough (having lost the O antigen). These clones, however, gained sensitivity to some phages that recognize outer membrane proteins as receptors. Despite the presence of nine potential antiviral systems in the genome of the UPEC124 strain, the difficulty in finding effective phages was largely due to the efficient, non-specific cell surface protection provided by the O antigen. These results highlight the importance of an individualized approach to phage therapy, where narrow host-range phages—typically avoided in pre-fabricated phage cocktails—may be instrumental. Furthermore, this study illustrates how integrating genomic, structural, and functional insights can guide the development of innovative therapeutic strategies, paving the way for broader applications of phage therapy in combating multidrug-resistant bacterial pathogens. [ABSTRACT FROM AUTHOR]

Biotechnology continues to drive innovation in the production of pharmaceuticals, biofuels, and other valuable compounds, leveraging the power of microbial systems for enhanced yield and sustainability. Genome-scale metabolic (GSM) modeling has become an essential approach in this field, which enables a guide for targeting genetic modifications and the optimization of metabolic pathways for various industrial applications. While single-species GSM models have traditionally been employed to optimize strains like Escherichia coli and Lactococcus lactis, the integration of these models into community-based approaches is gaining momentum. Herein, we present a pipeline for community metabolic modeling with a user-friendly GUI, applying it to analyze interactions between Methylococcus capsulatus, a biotechnologically important methanotroph, and Escherichia coli W3110 under oxygen- and nitrogen-limited conditions. We constructed models with unmodified and homoserine-producing E. coli strains using the pipeline implemented in the original BioUML platform. The E. coli strain primarily utilized acetate from M. capsulatus under oxygen limitation. However, homoserine produced by E. coli significantly reduced acetate secretion and the community growth rate. This homoserine was taken up by M. capsulatus, converted to threonine, and further exchanged as amino acids. In nitrogen-limited modeling conditions, nitrate and ammonium exchanges supported the nitrogen needs, while carbon metabolism shifted to fumarate and malate, enhancing E. coli TCA cycle activity in both cases, with and without modifications. The presence of homoserine altered cross-feeding dynamics, boosting amino acid exchanges and increasing pyruvate availability for M. capsulatus. These findings suggest that homoserine production by E. coli optimizes resource use and has potential for enhancing microbial consortia productivity. [ABSTRACT FROM AUTHOR]

Russian Snow White (RSW) chickens are characterized by high egg production, extreme resistance to low temperatures, disease resistance, and by the snow-white color of the day-old chicks. Studying the genome of this unique chicken breed will reveal its evolutionary history and help to understand the molecular genetic mechanisms underlying the unique characteristics of this breed, which will open new breeding opportunities and support future studies. We have sequenced and made a de novo assembly of the whole RSW genome using deep sequencing (250×) by the short reads. The genome consists of 40 chromosomes with a total length of 1.1 billion nucleotide pairs. Phylogenetic analysis placed the RSW near the White Leghorn, Fayoumi, and Houdan breeds. Comparison with other chicken breeds revealed a wide pool of mutations unique to the RSW. The functional annotation of these mutations showed the adaptation of genes associated with the development of the nervous system, thermoreceptors, purine receptors, and the TGF-beta pathway, probably caused by selection for low temperatures. We also found adaptation of the immune system genes, likely driven by selection for resistance to viral diseases. Integration with previous genome-wide association studies (GWAS) suggested several causal single nucleotide polymorphisms (SNPs). Specifically, we identified an RSW-specific missense mutation in the RALYL gene, presumably causing the snow-white color of the day-old chicks, and an RSW-specific missense mutation in the TLL1 gene, presumably affecting the egg weight. [ABSTRACT FROM AUTHOR]

A series of novel organogermanium(IV) catecholates 1–9 of the general formula R'2Ge(Cat), where R' = Ph, Et, have been synthesized. Compounds were characterized by 1H, 13C NMR, IR spectroscopy, and elemental analysis. The molecular structures of 1–3, 6, and 8 in crystal state were established using single-crystal X-ray analysis. The complexes are tetracoordinate germanium(IV) compounds containing a dioxolene ligand in a dianion (catecholato) form. Electrochemical transformations of target germanium(IV) complexes have been studied by cyclic voltammetry. The electro-oxidation mechanism of complexes 1–5, 7, and 10 (the related complex Ph2Ge(3,5-Cat) where 3,5-Cat is 3,5-di-tert-butylcatecholate) involves the consecutive formation of mono- and dicationic derivatives containing the oxidized forms of redox-active ligands. The stability of the generated monocations depends both on the hydrocarbon groups at the germanium atom and on the substituents in the catecholate ring. Compounds 6, 8, and 9 are oxidized irreversibly under the electrochemical conditions with the formation of unstable complexes. The radical scavenging activity and antioxidant properties of new complexes were estimated in the reaction with DPPH radical, ABTS radical cation, and CUPRACTEAC assay. It has been found that compounds 8 and 9 with benzothiazole or phenol fragments are more active in DPPH test. The presence of electron-rich moieties in the catecholate ligand makes complexes 5 and 7–9 more reactive to ABTS radical cation. The value of CUPRACTEAC for organogermanium(IV) catecholates varies from 0.23 to 1.45. The effect of compounds 1–9 in the process of lipid peroxidation of rat liver (Wistar) homogenate was determined in vitro. It was found that most compounds are characterized by pronounced antioxidant activity. A feature of complexes 1, 3, and 5–9 is the intensification of the antioxidant action with the incubation time. In the presence of additives of complexes 3, 5, 6, and 8, an induction period was observed during the process of lipid peroxidation. [ABSTRACT FROM AUTHOR]

Early evidence suggests a strong impact of tumour-infiltrating lymphocytes (TILs) on both the prognosis and clinical behaviour of ovarian cancer. Proven associations, however, have not yet translated to successful immunotherapies and further work in the field is urgently needed. We aimed to analyse the tumour microenvironment of a well-characterised cohort of ovarian cancer samples. Tumour markers were selected owing to their comparative underrepresentation in the current literature. Paraffin-embedded, formalin-fixed tumour tissue blocks of 138 patients representative of the population and including early stage disease were identified, stained for CD3, CD20, CD68 and CD163 and analysed for both the stromal and intertumoral components. Data were statistically analysed in relation to clinical details, histological subtype, borderline vs. malignant status, survival and management received. Mean stromal CD3, total CD3 count, mean stromal CD20 and total CD20 count all correlated negatively with survival. Malignant ovarian tumours consistently demonstrated significantly higher infiltration of all analysed immune cells than borderline tumours. Assessment of the stromal compartment produced a considerably higher proportion of significant results when compared to the intra-tumoural infiltrates. Customary assessment of solely intra-tumoural cells in advanced stage disease patients undergoing primary debulking surgery should be challenged, with recommendations for future scoring systems provided. [ABSTRACT FROM AUTHOR]

During gastrulation and neurulation, the chordamesoderm and overlying neuroectoderm of vertebrate embryos converge under the control of a specific genetic programme to the dorsal midline, simultaneously extending along it. However, whether mechanical tensions resulting from these morphogenetic movements play a role in long-range feedback signaling that in turn regulates gene expression in the chordamesoderm and neuroectoderm is unclear. In the present work, by using a model of artificially stretched explants of Xenopus midgastrula embryos and full-transcriptome sequencing, we identified genes with altered expression in response to external mechanical stretching. Importantly, mechanically activated genes appeared to be expressed during normal development in the trunk, i.e., in the stretched region only. By contrast, genes inhibited by mechanical stretching were normally expressed in the anterior neuroectoderm, where mechanical stress is low. These results indicate that mechanical tensions may play the role of a long-range signaling factor that regulates patterning of the embryo, serving as a link coupling morphogenesis and cell differentiation. [ABSTRACT FROM AUTHOR]

mRNA vaccines have been shown to be effective in combating the COVID-19 pandemic. The amount of research on the use of mRNAs as preventive and therapeutic modalities has undergone explosive growth in the last few years. Nonetheless, the issue of the stability of mRNA molecules and their translation efficiency remains incompletely resolved. These characteristics of mRNA directly affect the expression level of a desired protein. Regulatory elements of RNA—5′ and 3′ untranslated regions (UTRs)—are responsible for translation efficiency. An optimal combination of the regulatory sequences allows mRNA to significantly increase the target protein's expression. We assessed the translation efficiency of mRNA encoding of firefly luciferase with various 5′ and 3′UTRs in vitro on cell lines DC2.4 and THP1. We found that mRNAs containing 5′UTR sequences from eukaryotic genes HBB, HSPA1A, Rabb, or H4C2, or from the adenoviral leader sequence TPL, resulted in higher levels of luciferase bioluminescence 4 h after transfection of DC2.4 cells as compared with 5′UTR sequences used in vaccines mRNA-1273 and BNT162b2 from Moderna and BioNTech. mRNA containing TPL as the 5′UTR also showed higher efficiency (as compared with the 5′UTR from Moderna) at generating a T-cell response in mice immunized with mRNA vaccines encoding a multiepitope antigen. By contrast, no effects of various 5′UTRs and 3′UTRs were detectable in THP1 cells, suggesting that the observed effects are cell type specific. Further analyses enabled us to identify potential cell type-specific RNA-binding proteins that differ in landing sites within mRNAs with various 5′UTRs and 3′UTRs. Taken together, our data indicate high translation efficiency of TPL as a 5′UTR, according to experiments on DC2.4 cells and C57BL/6 mice. [ABSTRACT FROM AUTHOR]

Cellular respiration is associated with at least six distinct but intertwined biological functions. (1) biosynthesis of ATP from ADP and inorganic phosphate, (2) consumption of respiratory substrates, (3) support of membrane transport, (4) conversion of respiratory energy to heat, (5) removal of oxygen to prevent oxidative damage, and (6) generation of reactive oxygen species (ROS) as signaling molecules. Here we focus on function #6, which helps the organism control its mitochondria. The ROS bursts typically occur when the mitochondrial membrane potential (MMP) becomes too high, e.g., due to mitochondrial malfunction, leading to cardiolipin (CL) oxidation. Depending on the intensity of CL damage, specific programs for the elimination of damaged mitochondria (mitophagy), whole cells (apoptosis), or organisms (phenoptosis) can be activated. In particular, we consider those mechanisms that suppress ROS generation by enabling ATP synthesis at low MMP levels. We discuss evidence that the mild depolarization mechanism of direct ATP/ADP exchange across mammalian inner and outer mitochondrial membranes weakens with age. We review recent data showing that by protecting CL from oxidation, mitochondria-targeted antioxidants decrease lethality in response to many potentially deadly shock insults. Thus, targeting ROS- and CL-dependent pathways may prevent acute mortality and, hopefully, slow aging. [ABSTRACT FROM AUTHOR]

Optimizing physical training regimens to increase muscle aerobic capacity requires an understanding of the internal processes that occur during exercise that initiate subsequent adaptation. During exercise, muscle cells undergo a series of metabolic events that trigger downstream signaling pathways and induce the expression of many genes in working muscle fibers. There are a number of studies that show the dependence of changes in the activity of AMP-activated protein kinase (AMPK), one of the mediators of cellular signaling pathways, on the duration and intensity of single exercises. The activity of various AMPK isoforms can change in different directions, increasing for some isoforms and decreasing for others, depending on the intensity and duration of the load. This review summarizes research data on changes in the activity of AMPK, Ca2+/calmodulin-dependent protein kinase II (CaMKII), and other components of the signaling pathways in skeletal muscles during exercise. Based on these data, we hypothesize that the observed changes in AMPK activity may be largely related to metabolic and signaling transients rather than exercise intensity per se. Probably, the main events associated with these transients occur at the beginning of the exercise in a time window of about 1–10 min. We hypothesize that these transients may be partly due to putative trigger-like kinase/protein phosphatase interactions regulated by feedback loops. In addition, numerous dynamically changing factors, such as [Ca2+], metabolite concentration, and reactive oxygen and nitrogen species (RONS), can shift the switching thresholds and change the states of these triggers, thereby affecting the activity of kinases (in particular, AMPK and CaMKII) and phosphatases. The review considers the putative molecular mechanisms underlying trigger-like interactions. The proposed hypothesis allows for a reinterpretation of the experimental data available in the literature as well as the generation of ideas to optimize future training regimens. [ABSTRACT FROM AUTHOR]

The mRubyFT is a monomeric genetically encoded fluorescent timer based on the mRuby2 fluorescent protein, which is characterized by the complete maturation of the blue form with the subsequent conversion to the red one. It has higher brightness in mammalian cells and higher photostability compared with other fluorescent timers. A high-resolution structure is a known characteristic of the mRubyFT with the red form chromophore, but structural details of its blue form remain obscure. In order to obtain insight into this, we obtained an S148I variant of the mRubyFT (mRubyFTS148I) with the blocked over time blue form of the chromophore. X-ray data at a 1.8 Å resolution allowed us to propose a chromophore conformation and its interactions with the neighboring residues. The imidazolidinone moiety of the chromophore is completely matured, being a conjugated π-system. The methine bridge is not oxidized in the blue form bringing flexibility to the phenolic moiety that manifests itself in poor electron density. Integration of these data with the results of molecular dynamic simulation disclosed that the OH group of the phenolic moiety forms a hydrogen bond with the side chain of the T163 residue. A detailed comparison of mRubyFTS148I with other available structures of the blue form of fluorescent proteins, Blue102 and mTagBFP, revealed a number of characteristic differences. Molecular dynamic simulations with the combined quantum mechanic/molecular mechanic potentials demonstrated that the blue form exists in two protonation states, anion and zwitterion, both sharing enolate tautomeric forms of the C=C–O− fragment. These two forms have similar excitation energies, as evaluated by calculations. Finally, excited state molecular dynamic simulations showed that excitation of the chromophore in both protonation states leads to the same anionic fluorescent state. The data obtained shed light on the structural features and spectral properties of the blue form of the mRubyFT timer. [ABSTRACT FROM AUTHOR]