The canonical stop codons of the nuclear genome of the trypanosomatid Blastocrithidia nonstop are recoded. Here, we investigated the effect of this recoding on the mitochondrial genome and gene expression. Trypanosomatids possess a single mitochondrion and protein-coding transcripts of this genome require RNA editing in order to generate open reading frames of many transcripts encoded as 'cryptogenes'. Small RNAs that can number in the hundreds direct editing and produce a mitochondrial transcriptome of unusual complexity. We find B. nonstop to have a typical trypanosomatid mitochondrial genetic code, which presumably requires the mitochondrion to disable utilization of the two nucleus-encoded suppressor tRNAs, which appear to be imported into the organelle. Alterations of the protein factors responsible for mRNA editing were also documented, but they have likely originated from sources other than B. nonstop nuclear genome recoding. The population of guide RNAs directing editing is minimal, yet virtually all genes for the plethora of known editing factors are still present. Most intriguingly, despite lacking complex I cryptogene guide RNAs, these cryptogene transcripts are stochastically edited to high levels.
- MeSH
- buněčné jádro * genetika metabolismus MeSH
- editace RNA * MeSH
- genetický kód MeSH
- genom mitochondriální * MeSH
- guide RNA, Kinetoplastida genetika metabolismus MeSH
- kodon genetika MeSH
- messenger RNA genetika metabolismus MeSH
- mitochondrie genetika metabolismus MeSH
- otevřené čtecí rámce genetika MeSH
- protozoální proteiny genetika metabolismus MeSH
- RNA transferová * genetika metabolismus MeSH
- terminační kodon genetika MeSH
- Trypanosomatina genetika metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Ticks (Family Ixodidae) spend most of their life cycle as immature stages in the soil and litter, and as any other soil invertebrates, are likely to be controlled top-down by soil-dwelling predators. To date, the ability of soil invertebrate predators to control ixodid tick population remains little known, partly due to methodological difficulties. In the current study, we developed and successfully tested a novel method of labeling live Ixodes ricinus (L., 1758) (Ixodida: Ixodidae) nymphs with a 15N isotope label. Labeled ticks were used in a small-scale 8-day-long microcosm experiment to reveal soil predators attacking nymphs. Only a small fraction (4.1% of all samples) of soil generalist predators preyed upon nymphs. A strong 15N label was found in 5 predator species, namely 2 spiders (Pachygnatha listeri Sundevall, 1830, Tetragnathidae and Ozyptila sp., Theridiidae), 2 gamasid mites (Pergamasus beklemischevi Sellnick, 1929 and Pergamasus quisquiliarum [Canestrini, 1882], Parasitidae), and 1 staphylinid beetle (Geostiba circellaris [Gravenhorst, 1806], Staphylinidae). The isotopic labeling can be a useful tool in revealing a range of invertebrate predators that can control tick populations in soil.
We designed 0D, 1D, and 2D supramolecular assemblies made of diaryliodonium salts (functioning as double σ-hole donors) and carboxylates (as σ-hole acceptors). The association was based on two charge-supported halogen bonds (XB), which occurred between IIII sites of the iodonium cations and the carboxylate anions. The sequential introduction of the carboxylic groups in the aryl ring of the benzoic acid added a dimension to the 0D supramolecular organization of the benzoate, which furnished 1D-chained and 2D-layered structures when terephthalate and trimesate anions, correspondingly, were applied as XB acceptors. The structure-directing XB were studied using DFT calculations under periodic boundary conditions and were followed by the one-electron-potential analysis and the Bader atoms-in-molecules topological analysis of electron density. These theoretical methods confirmed the existence of the XB and verified the philicities of the interaction partners in the designed solid-state structures.
Instability is an intriguing characteristic of many protist genomes, and trypanosomatids are not an exception in this respect. Some regions of trypanosomatid genomes evolve fast. For instance, the trypanosomatid mitochondrial (kinetoplast) genome consists of fairly conserved maxicircle and minicircle molecules that can, nevertheless, possess high nucleotide substitution rates between closely related strains. Recent experiments have demonstrated that rapid laboratory evolution can result in the non-functionality of multiple genes of kinetoplast genomes due to the accumulation of mutations or loss of critical genomic components. An example of a loss of critical components is the reported loss of entire minicircle classes in Leishmania tarentolae during laboratory cultivation, which results in an inability to generate some correctly encoded genes. In the current work, we estimated the evolutionary rates of mitochondrial and nuclear genome regions of multiple natural Leishmania spp. We analyzed synonymous and non-synonymous substitutions and, rather unexpectedly, found that the coding regions of kinetoplast maxicircles are among the most variable regions of both genomes. In addition, we demonstrate that synonymous substitutions greatly predominate among maxicircle coding regions and that most maxicircle genes show signs of purifying selection. These results imply that maxicircles in natural Leishmania populations remain functional despite their high mutation rate.
- Publikační typ
- časopisecké články MeSH
Melatonin has been reported to cause myocardial electrophysiological changes and prevent ventricular tachycardia or fibrillation (VT/VF) in ischemia and reperfusion. We sought to identify electrophysiological targets responsible for the melatonin antiarrhythmic action and to explore whether melatonin receptor-dependent pathways or its antioxidative properties are essential for these effects. Ischemia was induced in anesthetized rats given a placebo, melatonin, and/or luzindole (MT1/MT2 melatonin receptor blocker), and epicardial mapping with reperfusion VT/VFs assessment was performed. The oxidative stress assessment and Western blotting analysis were performed in the explanted hearts. Transmembrane potentials and ionic currents were recorded in cardiomyocytes with melatonin and/or luzindole application. Melatonin reduced reperfusion VT/VF incidence associated with local activation time in logistic regression analysis. Melatonin prevented ischemia-related conduction slowing and did not change the total connexin43 (Cx43) level or oxidative stress markers, but it increased the content of a phosphorylated Cx43 variant (P-Cx43368). Luzindole abolished the melatonin antiarrhythmic effect, slowed conduction, decreased total Cx43, protein kinase Cε and P-Cx43368 levels, and the IK1 current, and caused resting membrane potential (RMP) depolarization. Neither melatonin nor luzindole modified INa current. Thus, the antiarrhythmic effect of melatonin was mediated by the receptor-dependent enhancement of impulse conduction, which was associated with Cx43 phosphorylation and maintaining the RMP level.
- MeSH
- antiarytmika farmakologie terapeutické užití MeSH
- kardiomyocyty metabolismus MeSH
- konexin 43 * metabolismus MeSH
- krysa rodu rattus MeSH
- melatonin * farmakologie terapeutické užití MeSH
- melatoninové receptory metabolismus MeSH
- srdeční arytmie farmakoterapie prevence a kontrola MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
BACKGROUND: FLNC is one of the few genes associated with all types of cardiomyopathies, but it also underlies neuromuscular phenotype. The combination of concomitant neuromuscular and cardiac involvement is not often observed in filaminopathies and the impact of this on the disease prognosis has hitherto not been analyzed. RESULTS: Here we provide a detailed clinical, genetic, and structural prediction analysis of distinct FLNC-associated phenotypes based on twelve pediatric cases. They include early-onset restrictive cardiomyopathy (RCM) in association with congenital myopathy. In all patients the initial diagnosis was established during the first year of life and in five out of twelve (41.7%) patients the first symptoms were observed at birth. RCM was present in all patients, often in combination with septal defects. No ventricular arrhythmias were noted in any of the patients presented here. Myopathy was confirmed by neurological examination, electromyography, and morphological studies. Arthrogryposes was diagnosed in six patients and remained clinically meaningful with increasing age in three of them. One patient underwent successful heart transplantation at the age of 18 years and two patients are currently included in the waiting list for heart transplantation. Two died due to congestive heart failure. One patient had ICD instally as primary prevention of SCD. In ten out of twelve patients the disease was associated with missense variants and only in two cases loss of function variants were detected. In half of the described cases, an amino acid substitution A1186V, altering the structure of IgFLNc10, was found. CONCLUSIONS: The present description of twelve cases of early-onset restrictive cardiomyopathy with congenital myopathy and FLNC mutation, underlines a distinct unique phenotype that can be suggested as a separate clinical form of filaminopathies. Amino acid substitution A1186V, which was observed in half of the cases, defines a mutational hotspot for the reported combination of myopathy and cardiomyopathy. Several independent molecular mechanisms of FLNC mutations linked to filamin structure and function can explain the broad spectrum of FLNC-associated phenotypes. Early disease presentation and unfavorable prognosis of heart failure demanding heart transplantation make awareness of this clinical form of filaminopathy of great clinical importance.
- MeSH
- fenotyp MeSH
- filaminy chemie genetika metabolismus MeSH
- kardiomyopatie * genetika metabolismus MeSH
- lidé MeSH
- nemoci svalů * MeSH
- restriktivní kardiomyopatie * genetika MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Leishmania tarentolae is a non-pathogenic trypanosomatid isolated from lizards widely used for heterologous protein expression and extensively studied to understand the pathogenic mechanisms of leishmaniasis. The repertoire of leishmanolysin genes was reported to be expanded in L. tarentolae genome, but no proteolytic activity was detected. Here, we analyzed L. tarentolae leishmanolysin proteins from the genome to the structural levels and evaluated the enzymatic activity of the wild-type and overexpressing mutants of leishmanolysin. A total of 61 leishmanolysin sequences were retrieved from the L. tarentolae genome. Five of them were selected for phylogenetic analysis, and for three of them, we built 3D models based on the crystallographic structure of L. major ortholog. Molecular dynamics simulations of these models disclosed a less negative electrostatic potential compared to the template. Subsequently, L. major LmjF.10.0460 and L. tarentolae LtaP10.0650 leishmanolysins were cloned in a pLEXSY expression system into L. tarentolae. Proteins from the wild-type and the overexpressing parasites were submitted to enzymatic analysis. Our results revealed that L. tarentolae leishmanolysins harbor a weak enzymatic activity about three times less abundant than L. major leishmanolysin. Our findings strongly suggest that the less negative electrostatic potential of L. tarentolae leishmanolysin can be the reason for the reduced proteolytic activity detected in this parasite.
- MeSH
- fylogeneze MeSH
- Leishmania * genetika metabolismus MeSH
- leishmanióza * parazitologie MeSH
- metaloendopeptidasy metabolismus MeSH
- paraziti * MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Melatonin treatment was reported to reduce the risk of cardiac arrhythmias, and crucial for this antiarrhythmic action was the effect of melatonin on activation spread. The aim of the present study was evaluation of the mechanisms of this activation enhancement. Experiments were performed in a total of 123 control and melatonin-treated (10 mg/kg, daily, for 7 days) male Wistar rats. In epicardial mapping studies (64 leads, interlead distance 0.5 mm) in the anesthetized animals, activation times (ATs) were determined in each lead as dV/dt minimum during QRS complex under sinus rhythm. Epicardial pacing was performed to measure conduction velocity (CV) across the mapped area. Average left ventricular ATs were shorter in the treated animals as compared to the controls, whereas the minimal epicardial ATs indicating the duration of activation propagation via the ventricular conduction system did not differ between the groups. CV was higher in the treated groups indicating that melatonin affected conduction via contractile myocardium The area of Cx43-derived fluorescence, as well as the expression of Cx43 protein, was similar in ventricles in the control and melatonin-treated groups. Expression of Gja1 gene transcripts encoding Cx43, was increased in the last group. An uncoupling agent octanol modified myocardial conduction properties (time of activation, action potential upstroke velocity, passive electrotonic phase duration) similarly in both groups. On the other hand, the expression of both Scn5a gene transcripts encoding Nav1.5 proteins, as well as peak density of transmembrane sodium current were increased in the ventricular myocytes from the melatonin-treated animals. Thus, a week-long melatonin treatment caused the increase of conduction velocity via enhancement of sodium channel proteins expression and increase of sodium current in the ventricular myocytes.
- MeSH
- konexin 43 * genetika MeSH
- krysa rodu rattus MeSH
- melatonin * farmakologie MeSH
- napěťově řízený sodíkový kanál, typ 5 * genetika MeSH
- potkani Wistar MeSH
- převodní systém srdeční * účinky léků MeSH
- sodík MeSH
- srdce fyziologie MeSH
- upregulace MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Actin is a fundamental member of an ancient superfamily of structural intracellular proteins and plays a crucial role in cytoskeleton dynamics, ciliogenesis, phagocytosis, and force generation in both prokaryotes and eukaryotes. It is shown that actin has another function in metazoans: patterning biosilica deposition, a role that has spanned over 500 million years. Species of glass sponges (Hexactinellida) and demosponges (Demospongiae), representatives of the first metazoans, with a broad diversity of skeletal structures with hierarchical architecture unchanged since the late Precambrian, are studied. By etching their skeletons, organic templates dominated by individual F-actin filaments, including branched fibers and the longest, thickest actin fiber bundles ever reported, are isolated. It is proposed that these actin-rich filaments are not the primary site of biosilicification, but this highly sophisticated and multi-scale form of biomineralization in metazoans is ptterned.
- MeSH
- aktiny * MeSH
- kostra MeSH
- oxid křemičitý * chemie MeSH
- sklo MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
AIM: To test a novel method of assessment of platelet adhesion to a fibrinogen-coated surface in whole blood under flow conditions. METHODS: We developed a fluidic device that mimics blood flow in vessels. The method of detection of platelet adhesion is based on recording of a scattered laser light signal from a fibrinogen-covered surface. Testing was performed in platelet-rich plasma (PRP) and whole blood of healthy volunteers. Control measurements were performed, followed by tests with inhibition of platelet GPIIa/IIIb and GPIb receptors. Then, the same testing sequence was performed in whole blood of persons with autoimmune thrombocytopenia and type 3 von Willebrand disease. RESULTS: The change in intensity of scattered light was 2.7 (2.4; 4.1) times higher in whole blood (0.2 ± 0.08V, n = 7) than in PRP (0.05 ± 0.02 V, n = 7), p < 0.01. The blocking of GP IIb/IIIa receptors decreased the intensity of scattered light to 8.5 (6.5;12)%; the blocking of GPIb receptors decreased it to 34 (23;58)%, p < 0.01. In the whole blood of a person with autoimmune thrombocytopenia, the inhibition of GPIb receptors decreased platelet adhesion, but no effect was observed in type 3 von Willebrand disease. Inhibition of platelet GPIIb/IIIa receptors alone or combined inhibition of GPIb and GPIIb/IIIa receptors resulted in almost total suppression of adhesion in both cases. CONCLUSION: Our system effectively registers platelet adhesion to a fibrinogen-coated surface under controlled-flow conditions and may successfully be applied to the investigation of platelet adhesion kinetics.
- MeSH
- adhezivita trombocytů * MeSH
- agregace trombocytů MeSH
- fibrinogen metabolismus MeSH
- inhibitory agregace trombocytů farmakologie MeSH
- kinetika MeSH
- lidé MeSH
- trombocytový glykoproteinový komplex Ib-IX antagonisté a inhibitory metabolismus MeSH
- trombocytový glykoproteinový komplex IIb-IIIa antagonisté a inhibitory metabolismus MeSH
- trombocyty metabolismus MeSH
- zdraví dobrovolníci pro lékařské studie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
