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Selected CEITEC Publications - 4Q 2013

Full versions of all publications can be downloaded by clicking on the titles.

 

Physical Review Letters
Interface-Induced Room-Temperature Ferromagnetism in Hydrogenated Epitaxial Graphene

Giesbers A., Uhlířová K., Konečný M., Peters E., Burkhard M., Aarts J., Flipse C.

Research Group:  Fabrication and Characterisation of Nanostructures
Research Programme: Advanced Nanotechnologies and Microtechnologies

Abstract
We show ferromagnetic properties of hydrogen-functionalized epitaxial graphene on SiC. Ferromagnetism in such a material is not directly evident as it is inherently composed of only nonmagnetic constituents. Our results nevertheless show strong ferromagnetism with a saturation of 0.9µB/hexagon projected area, which cannot be explained by simple magnetic impurities. The ferromagnetism is unique to hydrogenated epitaxial graphene on SiC, where interactions with the interfacial buffer layer play a crucial role. We argue that the origin of the observed ferromagnetism is governed by electron correlation effects of the narrow Si dangling bond states in the buffer layer exchange coupled to localized states in the hydrogenated graphene layer. This forms a quasi-three-dimensional ferromagnet with a Curie temperature higher than 300 K.

 

Acta Materialia
Stress dependence of the Peierls barrier of 1/2〈1 1 1〉screw dislocations in bcc metals

Gröger R., Vitek V.

Research Group: Transport and Magnetic Properties
Research Programme: Advanced Nanotechnologies and Microtechnologies

Abstract
The recently formulated constrained nudged elastic band method with atomic relaxations (NEB + r) is used to investigate the dependence of the Peierls barrier of 1/2〈1 1 1〉screw dislocations in body-centered cubic metals on non-glide stresses. These are the shear stresses parallel to the slip direction acting in the planes of the 〈1 1 1〉zone different from the slip plane, and the shear stresses perpendicular to the slip direction. Both these shear stresses modify the structure of the dislocation core and thus alter both the Peierls barrier and the related Peierls stress. Understanding of this effect of loading is crucial for the development of mesoscopic models of thermally activated dislocation motion via formation and propagation of pairs of kinks. The Peierls stresses and related choices of the glide planes determined from the Peierls barriers agree with the results of molecular statics calculations, which demonstrates that the NEB + r method is a reliable tool for determining the variation in the Peierls barrier with the applied stress. However, such calculations are very time consuming, and it is shown here that an approximate approach of determining the stress dependence of the Peierls barrier can be used, combined with test calculations employing the NEB + r method.

 

Physical Review B
Lattice instabilities in bulk EuTiO3

Bessas D., Rushchanskii K., Kachlík M., Disch S., Gourdon O., Bednarčík J., Maca K., Sergueev I., Kamba S., Ležaić M., Hermann R.

Research Group: Advanced Ceramic Materials
Research Programme: Advanced Materials

Abstract
The phase purity and the lattice dynamics in bulk EuTiO3 were investigated both microscopically, using x-ray and neutron diffraction, 151Eu-Mössbauer spectroscopy, and  151Eu nuclear inelastic scattering, and macroscopically using calorimetry, resonant ultrasound spectroscopy, and magnetometry. Furthermore, our investigations were corroborated by ab initio theoretical studies. The perovskite symmetry, Pm3m, is unstable at the M- and R-points of the Brillouin zone. The lattice instabilities are lifted when the structure relaxes in one of the symmetries: I4/mcm, Imma, R3with relative relaxation energy around −25 meV. Intimate phase analysis confirmed phase purity of our ceramics. A prominent peak in the Eu specific density of phonon states at 11.5 meV can be modeled in all candidate symmetries. A stiffening on heating around room temperature is indicative of a phase transition similar to the one observed in SrTiO3, however, although previous studies reported the structural phase transition to the tetragonal I4/mcm phase our detailed sample purity analysis and thorough structural studies using complementary techniques did not confirm a direct phase transition. Instead, in the same temperature range, Eu delocalization is observed which might explain the lattice dynamical instabilities.

 

Acta Materialia
Near-threshold propagation of mode II and mode III fatigue cracks in ferrite and austenite 

Vojtek T., Pippan R., Hohenwarter A., Holáň L., Pokluda J.

Research Group: Advanced Metallic Materials and Metal Based Composites
Research Programme: Advanced Materials

Abstract
The near-threshold behavior of mode II and mode III long fatigue cracks in ferritic (ARMCO iron) and austenitic (X5CrNi18-10) steel were experimentally studied using various samples specially prepared to obtain the effective threshold values ΔKIIeff,th and ΔKIIIeff,th. In both investigated materials, the effective thresholds for mode III were ~ 1.7 times higher than those for mode II. Three-dimensional topological data obtained by the examination of fracture surfaces using stereophotogrammetry were utilized to identify crack growth micromechanisms. In austenite, mode I branching of both the mode II and mode III cracks started at the very onset of crack growth. On the other hand, all cracks in ferrite propagated in crystallographically assisted local mixed mode I + II + III with mode II dominance. These experimental results can be understood in terms of crack growth micromechanisms according to a deformation model in ferrite and a decohesion model in austenite. The dissimilarity of growth mechanisms in ferrite and austenite may be attributed to a different number of available slip systems in body-centered cubic and face-centered cubic metals.

 

Nature Structural & Molecular Biology
Molecular basis of UG-rich RNA recognition by the human splicing factor TDP-43

Lukavsky P.J., Daujotyte D., Tollervey J.R., Ule J., Stuani C., Buratti E., Baralle F.E., Damberger F.F., Allain F. H-T

Research Group: RNA-based Regulation of Gene Expression
Research Programme: Structural Biology

Abstract
TDP-43 encodes an alternative-splicing regulator with tandem RNA-recognition motifs (RRMs). The protein regulates cystic fibrosis transmembrane regulator (CFTR) exon 9 splicing through binding to long UG-rich RNA sequences and is found in cytoplasmic inclusions of several neurodegenerative diseases. We solved the solution structure of the TDP-43 RRMs in complex with UG-rich RNA. Ten nucleotides are bound by both RRMs, and six are recognized sequence specifically. Among these, a central G interacts with both RRMs and stabilizes a new tandem RRM arrangement. Mutations that eliminate recognition of this key nucleotide or crucial inter-RRM interactions disrupt RNA binding and TDP-43–dependent splicing regulation. In contrast, point mutations that affect base-specific recognition in either RRM have weaker effects. Our findings reveal not only how TDP-43 recognizes UG repeats but also how RNA binding–dependent inter-RRM interactions are crucial for TDP-43 function.

 

Nucleic Acids Research
Crystal structures of B-DNA dodecamer containing the epigenetic modifications 5-hydroxymethylcytosine or 5-methylcytosine

Renciuk D., Blacque O., Vorlickova M., Spingler B.

Research Group: CD Spectroscopy of Nucleic Acids and Proteins
Research Programme: Structural Biology

Abstract
5-Hydroxymethylcytosine (5-hmC) was recently identified as a relatively frequent base in eukaryotic genomes. Its physiological function is still unclear, but it is supposed to serve as an intermediate in DNA de novo demethylation. Using X-ray diffraction, we solved five structures of four variants of the d(CGCGAATTCGCG) dodecamer, containing either 5-hmC or 5-methylcytosine (5-mC) at position 3 or at position 9. The observed resolutions were between 1.42 and 1.99 Å. Cytosine modification in all cases influences neither the whole B-DNA double helix structure nor the modified base pair geometry. The additional hydroxyl group of 5-hmC with rotational freedom along the C5-C5A bond is preferentially oriented in the 30 direction. A comparison of thermodynamic properties of the dodecamers shows no effect of 5-mC modification and a sequence-dependent only slight destabilizing effect of 5-hmC modification. Also taking into account the results of a previous functional study, we conclude that the 5 position of cytosine is an ideal place to encode epigenetic information. Like this, neither the helical structure nor the thermodynamics are changed, and polymerases cannot distinguish 5-hmC and 5-mC from unmodified cytosine, all these effects are making the former ones non-mutagenic. 

 

RNA a publication of the RNA society
Mammalian DIS3L2 exoribonuclease targets the uridylated precursors of let-7 miRNAs

Ustianenko D., Hrossová D., Potěšil D., Chalupníková K., Hrazdilová K., Pachernik J., Cetkovska K., Uldrijan S.,  Zdráhal Z., Vaňáčová Š.

Research Groups: RNA Quality Control & Proteomics
Research Programmes: Structural Biology & Genomics and Proteomics of Plant Systems

Abstract
The mechanisms of gene expression regulation by miRNAs have been extensively studied. However, the regulation of miRNA function and decay has long remained enigmatic. Only recently, 3′ uridylation via LIN28A-TUT4/7 has been recognized as an essential component controlling the biogenesis of let-7 miRNAs in stem cells. Although uridylation has been generally implicated in miRNA degradation, the nuclease responsible has remained unknown. Here, we identify the Perlman syndromeassociated protein DIS3L2 as an oligo(U)-binding and processing exoribonuclease that specifically targets uridylated pre-let-7 in vivo. This study establishes DIS3L2 as the missing component of the LIN28-TUT4/7-DIS3L2 pathway required for the repression of let-7 in pluripotent cells.

 

Nucleic Acids Research
Compromised telomere maintenance in hypomethylated Arabidopsis thaliana plants

Ogrocka A., Polanska P., Majerová E., Janeba Z., Fajkus J., Fojtová M.

Research Group: Chromatin Molecular Complexes
Research Programme: Genomics and Proteomics of Plant Systems

Abstract
Telomeres, nucleoprotein structures at the ends of linear eukaryotic chromosomes, are important for the maintenance of genomic stability. Telomeres were considered as typical heterochromatic regions, but in light of recent results, this view should be reconsidered. Asymmetrically located cytosines in plant telomeric DNA repeats may be substrates for a DNA methyltransferase enzyme and indeed, it was shown that these repeats are methylated. Here, we analyse the methylation of telomeric cytosines and the length of telomeres in Arabidopsis thaliana methylation mutants ( met 1-3 and ddm 1-8 ), and in their wild-type siblings that were germinated in the presence of hypomethylation drugs.

 

Current Biology

Local Auxin Sources Orient the Apical-Basal Axis in Arabidopsis Embryos

Robert H.S., Grones P., Stepanova A.N., Robles L.M., Lokerse A.S., Alonso J.M., Weijers D., Friml J.

Research Group: Developmental and Cell Biology of Plants
Research Programme: Genomics and proteomics of plant systems

Abstract
Establishment of the embryonic axis foreshadows the main body axis of adults both in plants and in animals, but underlying mechanisms are considered distinct. Plants utilize directional, cell-to-cell transport of the growth hormone auxin to generate an asymmetric auxin response that specifies the embryonic apical-basal axis. The auxin flow directionality depends on the polarized subcellular localization of PIN-FORMED (PIN) auxin transporters. It remains unknown which mechanisms and spatial cues guide cell polarization and axis orientation in early embryos. Herein, we provide conceptually novel insights into the formation of embryonic axis in Arabidopsis by identifying a crucial role of localized tryptophan-dependent auxin biosynthesis.

 

The Plant Cell
The More the Merrier: Recent Hybridization and Polyploidy in Cardamine

Mandáková T., Kovařík A., Zozomová-Lihová J., Shimizu-Inatsugid R., Shimizu K., Mummenhoff K., Marholdc K., Lysak M.

Research Group: Plant Cytogenomics
Research Programme: Genomics and proteomics of plant systems

Abstract
This article describes the use of cytogenomic and molecular approaches to explore the origin and evolution of Cardamine schulzii, a textbook example of a recent allopolyploid, in its ∼110-year history of human-induced hybridization and allopolyploidy in the Swiss Alps. Triploids are typically viewed as bridges between diploids and tetraploids but rarely as parental genomes of high-level hybrids and polyploids. The genome of the triploid semifertile hybrid Cardamine × insueta (2n = 24, RRA) was shown to combine the parental genomes of two diploid (2n = 2x = 16) species, Cardamine amara (AA) and Cardamine rivularis (RR).

 

Carcinogenesis
Identification of novel sequence variations in microRNAs in chronic lymphocytic leukemia

Kmínková J., Mráz M., Zápražná K., Navrkalová V., Tichý B., Plevová K., Malčíková J., Černá K., Rausch T., Beneš V., Brychtová Y., Doubek M., Mayer J., Pospíšilová S.

Research Group: Medical genomics
Research Programme: Molecular medicine

Abstract
MicroRNA (miRNA) expression is deregulated in many tumors including chronic lymphocytic leukemia (CLL). Although the particular mechanism responsible for their aberrant expression is not well characterized, the presence of mutations and single nucleotide polymorphisms (SNP) in miRNA genes, possibly affecting their secondary structure and expression, has been described. In CLL, however, the impact and frequency of such variations have yet to be elucidated. Using a custom resequencing microarray, we screened sequence variations in 109 cancer related pre-miRNAs in 98 CLL patients. Additionally, the primary regions of miR-29b-2/29c were analyzed by Sanger sequencing in another cohort of 213 CLL patients. Altogether, we describe 6 novel miR-sequence variations and the presence of SNPs (n=27), most of which changed the miR-secondary structure. Moreover, some of the identified SNPs have a significantly different frequency in CLL when compared to a control population. Additionally, we identified a novel variation in miR-16-1 that had not been previously described in CLL patients. We show that this variation affects the expression of mature miR-16-1. We also show that the expression of another miRNA with pathogenetic relevance for CLL, namely miR-29b-2, is influenced by the presence of a polymorphic insertion which is more frequent in CLL than in a control population. Altogether, these data suggest that sequence variations may occur during CLL development and/or progression.

 

Nucleic Acids Research
Quadruplex-forming sequences occupy discrete regions inside plant LTR retrotransposons

Lexa M., Kejnovský E., Šteflová P., Konvalinová H., Vorlíčková M., Výskot B.

Research Group: Genome Dynamics
Research Programme: Molecular Medicine

Abstract
Retrotransposons with long terminal repeats (LTR) form a significant proportion of eukaryotic genomes, especially in plants. They have
gag and pol genes and several regulatory regions necessary for transcription and reverse transcription. We searched for potential quadruplex-forming sequences (PQSs) and potential triplex-forming sequences (PTSs) in 18 377 full-length LTR retrotransposons collected from 21 plant species. We found that PQSs were often located in LTRs, both upstream and downstream of promoters from which the whole retrotransposon is transcribed. Upstream-located guanine PQSs were dominant in the minus DNA strand, whereas downstreamlocated guanine PQSs prevailed in the plus strand,indicating their role both at transcriptional and posttranscriptional levels. Our circular dichroism spectroscopy measurements confirmed that these PQSs readily adopted guanine quadruplex structures – some of them were paralell-stranded, while others were anti-parallel-stranded. The PQS often formed doublets at a mutual distance of up to 400 bp. PTSs were most abundant in 30UTR (but were also present in 50UTR). We discuss the potential role of quadruplexes and triplexes as the regulators of various processes participating in LTR retrotransposon life cycle and as potential recombination sites during post-insertional retrotransposonbased genome rearrangements.

 

Neural Plasticity
The Mechanisms of Movement Control and Time Estimation in Cervical Dystonia Patients 

Filip P., Lungu O.V., Shaw D.J., Kasparek T., Bareš M.

Research Group: Behavioural and Social Neuroscience
Research Programme: Brain and Mind Research

Abstract
Traditionally, the pathophysiology of cervical dystonia has been regarded mainly in relation to neurochemical abnormities in the basal ganglia. Recently, however, substantial evidence has emerged for cerebellar involvement. While the absence of neurological "cerebellar signs" in most dystonia patients may be considered at least provoking, there are more subtle indications of cerebellar dysfunction in complex, demanding tasks. Specifically, given the role of the cerebellum in the neural representation of time, in the millisecond range, dysfunction to this structure is considered to be of greater importance than dysfunction of the basal ganglia. In the current study, we investigated the performance of cervical dystonia patients on a computer task known to engage the cerebellum, namely, the interception of a moving target with changing parameters (speed, acceleration, and angle) with a simple response (pushing a button). The cervical dystonia patients achieved significantly worse results than a sample of healthy controls. Our results suggest that the cervical dystonia patients are impaired at integrating incoming visual information with motor responses during the prediction of upcoming actions, an impairment we interpret as evidence of cerebellar dysfunction.

 

Schizophrenia Research
A detailed analysis of the effect of repetitive transcranial magnetic stimulation on negative symptoms of schizophrenia: a double-blind trial

Prikryl R., Ustohal L., Prikrylova Kucerova H., Kasparek T., Venclikova S., Vrzalova M., Ceskova E.

Research Group: Behavioural and Social Neuroscience
Research Programme: Brain and Mind Research

Abstract

Objective: The aim of the study was to assess the effect of rTMS not only on the general severity of negative schizophrenia symptoms, but also particularly on their individual domains, such as affective flattening or blunting, alogia, avolition or apathy, anhedonia, and impaired attention.
Methods: Forty schizophrenic male patients on stable antipsychotic medication with prominent negative symptoms were included in the study. They were divided into two groups: 23 were treated with active and 17 with placebo rTMS. Both treatments were similar, but placebo rTMS was administered using a purpose-built sham coil. Stimulation was applied to the left dorsolateral prefrontal cortex (DLPFC). The stimulation frequency was 10 Hz; stimulation intensity was 110% of the individual motor threshold intensity. Each patient received 15 rTMS sessions on 15 consecutive working days (five working days "on" and two weekend days "off" design). Each daily session consisted of 20 applications of 10-second duration with 30-second intervals between sequences. The patients and raters were blind to condition of stimulation treatment.
Results: The active rTMS led to a statistically significantly higher reduction of the Scale for the Assessment of Negative Symptoms (SANS) total score and of all domains of negative symptoms of schizophrenia. After Bonferroni adjustments for multiple testing, the statistical significance disappeared in alogia only.
Conclusion: High-frequency rTMS stimulation over the left DLPFC at a high stimulation intensity with a sufficient number of applied stimulating pulses may represent an efficient augmentation of antipsychotics in alleviating the negative symptoms of schizophrenia.

 

Chromosome Research
Subchromosomal karyotype evolution in Equidae

Musilova P., Kubickova S., Vahala J., Rubes J.

Research Group: Animal Genomics
Research Programme: Molecular Veterinary Medicine 

Abstract
Equidae is a small family which comprises horses, African and Asiatic asses, and zebras. Despite equids having diverged quite recently, their karyotypes underwent rapid evolution which resulted in extensive differences among chromosome complements in respektive species. Comparative mapping using wholechromosome painting probes delineated genome-wide chromosome homologies among extant equids, enabling us to trace chromosome rearrangements that occurred during evolution. In the present study, we performed subchromosomal comparative mapping among seven Equidae species, representing the whole family. Region-specific painting and bacterial artificial chromosome probes were used to determine the orientation of evolutionarily conserved segments with respect to centromere positions. This allowed assessment of the configuration of all fusions occurring during the evolution of Equidae, as well as revealing discrepancies in centromere location caused by centromere repositioning or inversions. Our results indicate that the prevailing type of fusion in Equidae is centric fusion. Tandem fusions of the type telomere–telomere occur almost exclusively in the karyotype of Hartmann’s zebra and are characteristic of this species’ evolution.We revealed inversions in segments homologous to horse chromosomes 3p/10p and 13 in zebras and confirmed inversions in segments 4/31 in African ass, 7 in horse and 8p/20 in zebras. Furthermore, our mapping results suggested that centromere repositioning events occurred in segments homologous to horse chromosomes 7, 8q, 10p and 19 in the African ass and an element homologous to horse chromosome 16 in Asiatic asses. Centromere repositioning in chromosome 1 resulted in three different chromosome types occurring in extant species. Heterozygosity of the centromere position of this chromosome was observed in the kiang. Other subtle changes in centromere position were described in several evolutionary conserved chromosomal segments, suggesting that tiny centromere repositioning or pericentric inversions are quite frequent in zebras and asses.

 

PLOS One
Preliminary Insights into the Impact of Dietary Starch on the Ciliate, Neobalantidium coli, in Captive Chimpanzees

Schovancova K.,  Pomajbikova K., Prochzka D.,  Modry D., Bolechova P.,  Petrzelkova K.J.

Research Group: Parasitology
Research ProgrammeMolecular Veterinary Medicine

Abstract
Infections caused by the intestinal ciliate Neobalantidium coli are asymptomatic in most hosts. In humans and captive African great apes clinical infections occasionally occur, manifested mainly by dysentery; however, factors responsible for development of clinical balantidiasis have not been fully clarified. We studied the effect of dietary starch on the intensities of infection by N. coli in two groups of captive chimpanzees. Adult chimpanzees infected by N. coli from the Hodonín Zoo and from the Brno Zoo, Czech Republic, were fed with a high starch diet (HSD) (average 14.7% of starch) for 14 days, followed by a five-day transition period and subsequently with a period of low starch diet (LoSD) (average 0.1% of starch) for another 14 days. We collected fecal samples during the last seven days of HSD and LoSD and fixed them in 10% formalin. We quantified trophozoites of N. coli using the FLOTAC method. The numbers of N. coli trophozoites were higher during the HSD (mean 6 SD: 49.06134.7) than during the LoSD (3.566.8). A generalized linear mixed-effects model revealed significantly lower numbers of the N. coli trophozoites in the feces during the LoSD period in comparison to the HSD period (treatment contrast LoSD vs. HSD: 2.760.06 (SE), z = 47.7; p,,0.001). We conclude that our data provide a first indication that starch-rich diet might be responsible for high intensities of infection of N. coli in captive individuals and might predispose them for clinically manifested balantidiasis. We discuss the potential nutritional modifications to host diets that can be implemented in part to control N. coli infections. 

 

Bioorganic & Medicinal Chemistry
New derivatives of salicylamides: Preparation and antimicrobial activity against various bacterial species

Pauk K., Zadražilova I., Imramovský A., Vinšova J., Pokorná M., Masaříkova M., Cizek A., Jampilek, J.

Research Group: Molecular Bacteriology
Research Programme: Molecular Veterinary Medicine

Abstract
Three series of salicylanilides, esters of N-phenylsalicylamides and 2-hydroxy-N-[1-(2-hydroxyphenylamino)-1-oxoalkan-2-yl]benzamides, in total thirty target compounds were synthesized and characterized. The compounds were evaluated against seven bacterial and three mycobacterial strains. The antimicrobial activities of some compounds were comparable or higher than the standards ampicillin, ciprofloxacin or isoniazid. Derivatives 3f demonstrated high biological activity against Staphylococcus aureus (60.03 μmol/L), Mycobacterium marinum (60.40 μmol/L) and Mycobacterium kansasii (1.58 μmou/L), 3g shows activity against Clostridium perfringens (60.03 μmou/L) and Bacillus cereus (0.09 μmol/L), 3h against Pasteurella multocida (60.03 μmol/L) and M. kansasii (60.43 μmol/L), 3i against methicillin-resistant S. aureus and B. cereus (60.03 μmol/L). The structure–activity relationships are discussed for all the compounds.

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