THE THEME IS OF THE STUDY AND DOCUMENTATION THE IMPLEMENTATION OF THE CONSTRUCTION OF THE NEW BUILDING OF THE INTEGRATED RESCUE SYSTEM – STATION TYPE P1. THE FIRE STATION IS DESIGNED INTO THREE OPERATING UNITS. THE MAIN ACCESS TO THE BUILDING IS IN THE 1ST FLOOR ADMINISTRATIVE AREA IN THE SOUTH-EAST SIDE OF THE BUILDING. THIS PART OF THE BUILDING HAS 2 FLOORS. THE OBJECT CONTINUES TO THE SOUTHEAST TOWARDS THE PATH OF THE GARAGE WITH 5 SPACES. ON THE NORTHWEST EDGE OF THE OBJECT IS A TECHNICAL BACKGROUND WITH A DISHWASHER AND A DRYER. TO DEVELOP OVER THE OBJECT PERSONAL VISITS FROM TWO FIRE STATIONS. ONE REGIONAL IN CESKE BUDEJOVICE AND SECOND DISTRICT IN TÝN NAD VLTAVOU. FOR INSPECTION AND UNDERSTANDING PROPOSAL FOR A NEW FIRE STATION ALSO HELPED TOUR OF DOCUMENTATION PROJECT ALREADY MADE ON CONSTRUCTION FIRE STATION IN MARIANSKE LAZNE AND FRYMBURK.

NK cells are innate lymphocytes which constitute the first line of organism's defence against infections through their receptor system. These cells represent an important part of antiviral and antitumor immunity, they also play a role in transplant immunity, autoimmunity and reproduction. This diploma thesis inquires into the structure of the transmembrane receptor NKR-P1B of mouse NK cells and the interaction with its ligand Clr-b. The aim was to prepare the expression vector coding the ligand-binding and whole extracellular region of the receptor NKR-P1B and to optimize its production and refolding in vitro. Purified protein samples were analyzed by size-exclusion chromatography, electrophoresis and mass spectrometry. Interaction between NKR-P1B and Clr-b proteins was tested using biophysical (size-exclusion chromatography and surface plasmon resonance) and biological methods (labelling of cellular sample with NKR-P1B proteins marked with fluorescent dye). In vitro binding experiments have not confirmed mutual interaction between NKR-P1B and Clr-b despite the prepared proteins binding to the bone marrow cells.

Sexual conflict is a term that describes the situation where traits can experience opposing selection pressures in the two sexes. Theory suggests this conflict exists in all organisms with separate sexes, and specific chromosome clusters which are possibly sexually antagonistic have been identified in the species Drosophila melanogaster. One of all identified genes is CG3598, which have proved yielding higher fitness for males carrying allele A2 on this gene compared to A1. In this study, factors which contribute to the difference in fitness between these two alleles, with regards to sperm competition and mating success were observed. A double mating design was used, in which males and females were placed in test tubes together in order to examine the number of copulations and the defensive (P1) and offensive (P2) ability of sperm. The relative fitness of males carrying A1 did not differ from males carrying A2, which rejects previous studies, however, A2 had a higher defensive capability compared to A1. On the other hand, A1 instead had better offensive capability and higher amount of rematings. This indicates that the defensive capability of A2 is very strong and opposes the offensive capability of A1, but also the increased rate of rematings in A1. To get a more precise understanding of the fitness relation between A1 and A2 on the gene CG3598, further experiments would need to be performed on the subject. / Sexuell konflikt är ett begrepp som beskriver förhållandet där egenskaper upplever olika selektionstryck beroende på kön. Teorier finns om att den här konflikten existerar i alla organismer med olika kön, och i arten Drosophila melanogaster har det identifierats specifika kromosomkluster som har möjlighet att bidra till sexuell konflikt. En utav alla identifierade gener är CG3598 som har påvisats ge en högre fitness för hanar bärande allel A2 på denna gen jämfört med allel A1. I den här studien undersöks bakomliggande faktorer till skillnaden i fitness mellan dessa två alleler, med avseende på spermiekonkurrens och parningsframgång. Genom en dubbel parningsdesign, där hanar och honor placerades tillsammans i rör undersöktes den defensiva spermieförmågan (P1), den offensiva förmågan (P2) och antal parningar. Den relativa fitnessen hos hanar med A1 skiljde sig inte från A2, vilket motsäger tidigare studier. Däremot var den defensiva förmågan högre för A2 och A1 hade istället högre offensiv förmåga samt en högre andel omparningar. Detta indikerar att A2 har en stark defensiv förmåga som motsätter den offensiva förmågan i A1 men också möjligheten till fler omparningar. För att få en mer precis uppfattning skulle ytterligare experiment behöva utföras då det i denna studie var en väldigt låg andel hanar som parade sig jämfört med vad som förväntas.

Les ATPases de type P1 sont des pompes utilisant l'énergie de l'hydrolyse de l'ATP pour transporter les ions lourds (Cu+, Zn2+, Pb2+ Cd2+) à travers la membrane cellulaire. Elles sont difficiles à purifier et cristalliser et leur structure 3D est en général inconnue. Nous nous sommes intéressés à la structure et à la dymanique de la partie membranaire de l'ATPase cadmium CadA et aux domaines de liaison du métal de l'ATPase cuivre humaine de Menkés. Similarité de séquence et analyses d'hydropathie, complétées par des expériences ont montré que les ATPases de type P1 sont constituées de 8 segments transmembranaires (TMs) au lieu de 10 pour l'ATPase calcium de structure connue. En collaboration avec les biochimistes, et en utilisant les programmes MODELLER, CHARMM, XPLOR, AMD ainsi que nos propres programmes, nous avons prédit la structure des TMs de CadA. Nous avons construit plusieurs modéles du paquet de TMs correspondant à plusieurs topologies, calculé les coordonnées atomiques avec une procédure similaire à la détermination de structure à partir d'expériences de RMN et raffiné ces coordonnées en utilisant des simulations de DM en présence de solvant implicite. Le programme AMD de DM Adaptive a été utilisé pour vérifier les modèles de manière interactive. Une autre caractéristique intéressante des ATPases de type P1 est la présence en N-ter de un à six domaine(s) de liaison des métaux (MBDS). Dans le cas de l'ATPase de Menkés, il y a 6 MBDs, chacun pouvant lier un ion Cu$^+$. La structure de chaque MBD est connue. En utilisant des simulations de DM, nous avons étudié la dynamique de chaque MBD en presence ou absence de métal dans le but de comprendre comment le métal est transféré de la métallochaperone qui prend en charge le métal lors de son entrée dans la cellule au MBD. Ces études ont utilisé le travail récent des chercheurs de l'équipe dans le domaine de la paramétrisation des ions métalliques pour des champs de force de mécanique moléculaire. Le bon accord de nos résultats de DM sur les MBDs avec les connaissances expérimentales a montré que des modèles mécaniques sont capables de rendre compte et d'expliquer certaines propriétés de liaison et de transport des métaux dans les métalloprotéines et les ATpases, cibles pharmaceutiques bien connues.

P1 prophage is stably maintained in E. coli as a low-copy-number plasmid. Stable maintenance of P1 is dependent on the function of the plasmid encoded partition system, parABS. ParA is the partition ATPase, ParB is the partition-site binding protein, and parS is the partition site. The concerted action of these proteins results in dynamic movement of the plasmid over the bacterial nucleoid, which results in its stable maintenance. Plasmid movement has been proposed to be caused by interactions between parS bound ParB and nucleoid bound ParA. In this thesis, I have identified a complex of ParA, ParB, and DNA that is capable of promoting plasmid stability. ParA, ParB, DNA interactions required the ATP bound conformation of ParA. The ParA-ParB-DNA complex was dynamically regulated by nucleotide hydrolysis, which promoted complex disassembly. Complex formation resulted from the cooperative binding of ParA and ParB to DNA. ParA-ParB and ParB-DNA interactions were both necessary for complex formation. ParA-ParB-DNA complex size was regulated by ParB stimulation of ParA-ATP hydrolysis. Microscopy demonstrated that complexes resulted in the association of multiple DNA molecules due to protein binding. The properties of complex assembly, dynamics, and DNA grouping lead me to propose a model where associations between ParA bound to the bacterial nucleoid and the partition complex mediated plasmid movement and localization.

P1 prophage is stably maintained in E. coli as a low-copy-number plasmid. Stable maintenance of P1 is dependent on the function of the plasmid encoded partition system, parABS. ParA is the partition ATPase, ParB is the partition-site binding protein, and parS is the partition site. The concerted action of these proteins results in dynamic movement of the plasmid over the bacterial nucleoid, which results in its stable maintenance. Plasmid movement has been proposed to be caused by interactions between parS bound ParB and nucleoid bound ParA. In this thesis, I have identified a complex of ParA, ParB, and DNA that is capable of promoting plasmid stability. ParA, ParB, DNA interactions required the ATP bound conformation of ParA. The ParA-ParB-DNA complex was dynamically regulated by nucleotide hydrolysis, which promoted complex disassembly. Complex formation resulted from the cooperative binding of ParA and ParB to DNA. ParA-ParB and ParB-DNA interactions were both necessary for complex formation. ParA-ParB-DNA complex size was regulated by ParB stimulation of ParA-ATP hydrolysis. Microscopy demonstrated that complexes resulted in the association of multiple DNA molecules due to protein binding. The properties of complex assembly, dynamics, and DNA grouping lead me to propose a model where associations between ParA bound to the bacterial nucleoid and the partition complex mediated plasmid movement and localization.

While an information retrieval system takes as input a user query and returns a list of relevant documents chosen from a large collection, a question answering system attempts to produce an exact answer. Recent research, motivated by the question answering track of the Text REtrieval Conference (TREC) has focused mainly on answering ‘factoid’ questions concerned with names, places, dates etc. in the news domain. However, questions seeking definitions of terms are common in the logs of search engines. The objective of this project was therefore to investigate methods of retrieving definitions from scientific documents. The subject domain was salmon, and an appropriate test collection of articles was created, pre-processed and indexed. Relevant terms were obtained from salmon researchers and a fish database. A system was built which accepted a term as input, retrieved relevant documents from the collection using a search engine, identified definition phrases within them using a vocabulary of syntactic patterns and associated heuristics, and produced as output phrases explaining the term. Four experiments were carried out which progressively extended and refined the patterns. The performance of the system, measured using an appropriate form of precision, improved over the experiments from 8.6% to 63.6%. The main findings of the research were: (1) Definitions were diverse despite the documents’ homogeneity and found not only in the Introduction and Abstract sections but also in the Methods and References; (2) Nevertheless, syntactic patterns were a useful starting point in extracting them; (3) Three patterns accounted for 90% of candidate phrases; (4) Statistically, the ordinal number of the instance of the term in a document was a better indicator of the presence of a definition than either sentence position and length, or the number of sentences in the document. Next steps include classifying terms, using information extraction-like templates, resolving basic anaphors, ranking answers, exploiting the structure of scientific papers, and refining the evaluation process.

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moderadas, ela proporciona efeitos benéficos sobre as funções cognitivas na vida adulta e no decorrer do envelhecimento. No entanto, a ingestão crescente de bebidas contendo cafeína por adolescentes tem causado preocupação, pois os efeitos desta substância sobre as funções cognitivas e a maturação do encéfalo durante a adolescência são pouco conhecidos. A cafeína atravessa a placenta e a barreira hemato-encefálica e o seu consumo tem sido associado ao maior risco de aborto espontâneo e baixo peso ao nascer. Portanto, nos estágios iniciais do desenvolvimento encefálico o consumo de cafeína também carece de maiores eslcarecimentos. Nesta tese, o impacto do consumo de cafeína durante diferentes fases de desenvolvimento do encéfalo foi investigado sobre o comportamento e proteínas sinápticas em ratos. No primeiro capítulo, ratos adolescentes machos consumiram cafeína na água de beber nas doses de 0,1; 0,3 e 1,0 g/L (correspondendo ao consumo baixo, moderado e elevado, respectivamente) somente durante o seu período ativo (das 19 às 7 horas). Nenhuma das doses testadas teve efeito sobre a atividade locomotora, porém todas desencadearam efeitos ansiogênicos. A cafeína (0,3 e 1,0 g/L) melhorou o desempenho na tarefa de reconhecimento ao objeto, enquanto na dose mais elevada (1,0 g/L) os animais não habituaram ao campo aberto, uma forma de avaliar o aprendizado não-associativo. Todas as doses testadas reduziram a densidade de proteína glial fibrilar ácida (GFAP) e proteína associada ao sinaptossoma (SNAP-25) sem causar alterações na imunorreatividade da proteína nuclear específica para neurônios (NeuN) no hipocampo e no córtex cerebral No hipocampo, a cafeína (em todas as doses testadas) aumentou a densidade de receptor de adenosina A1 e reduziu a do factor neurotrófico derivado do encéfalo (BDNF) e sua forma precursora (proBDNF) (1,0 g/L). No córtex cerebral, a cafeína (1,0 g/L) reduziu a densidade do receptor A1 e aumentou a do BDNF e do proBDNF (0,3 e 1,0 g/L). Estes resultados revelam que o consumo de cafeína por ratos adolescentes exacerba a ansiedade, mas provoca diferentes efeitos sobre a memória, melhorando a de reconhecimento e prejudicando o aprendizado não associativo. Parte destes efeitos foi associada às mudanças nos níveis de BDNF, GFAP e SNAP-25, porém sem perda da viabilidade neuronal aparente no hipocampo e no córtex cerebral. No segundo capítulo, o impacto do consumo de cafeína (0,1; 0,3 e 1,0 g/L na água de beber, das 19 às 7 horas) foi investigado sobre o comportamento e proteínas sinápticas na vida adulta dos animais que consumiram cafeína no decorrer do desenvolvimento encefálico. O consumo de três diferentes doses de cafeína iniciou 15 dias antes do acasalamento e permaneceu durante a prenhez e lactação. A partir do desmame os animais foram divididos em dois grupos: os que consumiram cafeína até a vida adulta (ao longo da vida) e os que interromperam o consumo (desenvolvimento). Esses dois grupos também foram subdivididos e analisados de acordo com o sexo. Foram comparados os efeitos destes protocolos sobre o comportamento e a densidade de proteínas sinápticas do hipocampo e córtex de fêmeas e machos adultos. A memória de reconhecimento foi prejudicada nas fêmeas que receberam cafeína (0,3 e 1,0 g/L) durante o desenvolvimento, o que coincidiu com o aumento do proBDNF e níveis inalterados de BDNF no hipocampo Ambos os protocolos de exposição causaram hiperlocomoção nos machos, enquanto que nas fêmeas somente a exposição ao longo da vida aumentou a atividade locomotora de forma significativa. Já no comportamento relacionado à ansiedade, ambos os sexos apresentaram um perfil ansiolítico ao consumir cafeína (1,0 g/L) ao longo da vida. Ambos os regimes de administração diminuíram os níveis de GFAP e SNAP-25 no hipocampo dos ratos machos. A densidade do receptor de TrkB foi reduzida no hipocampo em ambos os sexos e protocolos de exposição. No córtex cerebral BDNF e proBDNF aumentaram com o consumo de cafeína ao longo da vida nos machos. Nas fêmeas houve aumento no BDNF, mas não no proBDNF, em ambos regimes de administração. O receptor TrkB diminuiu no córtex dos ratos machos que receberam cafeína somente durante o desenvolvimento. Ambas proteínas – GFAP e SNAP-25 – aumentaram suas densidades nos machos que receberam ambos regimes de administração. Estes resultados revelaram que o consumo de cafeína ao longo da vida pode recuperar o prejuízo na memória de reconhecimento das fêmeas que consumiram a substância durante o desenvolvimento e indicam que a exposição durante um período específico do desenvolvimento do encéfalo promove alterações comportamentais dependentes do sexo, as quais nós relacionamos com modificações na sinalização BDNF. Os resultados desta tese destacam a importância de controlar o consumo de cafeína em períodos críticos para o desenvolvimento encefálico de ratos, e aponta para um efeito dependente do sexo. No entanto, mais estudos são necessários para ampliar nosso conhecimento sobre as possíveis vias de sinalização envolvidas nestes processos. / Caffeine is the most consumed psychostimulant substance worldwide, with benefits for cognitive functioning. Caffeine intake at moderate doses also prevents age-related cognitive decline. However, health experts have raised concerns about the growing intake of caffeine-containing drinks by adolescent population. In fact, the effects of caffeine on cognitive functions and neurochemical aspects of late brain maturation during adolescence are poorly understood. In addition, caffeine consumption in the early stages of fetal development has been associated with miscarriage and low birth weight, since it penetrates placenta and blood-brain barrier during pregnancy. Therefore, the impact of caffeine intake was investigated during different stages of brain development. In the first chapter of this thesis, adolescent male rats consumed caffeine in the drinking water (0.1; 0.3 and 1.0 g/L corresponding to low, moderate and high doses, respectively) only during their active period (from 7:00 p.m. to 7:00 a.m.). None of the doses tested had effect on locomotor activity, whereas all triggered anxiogenic effects. Caffeine (0.3 and 1.0 g/L) improved the performance in the object recognition task, but the higher dose of caffeine (1.0 g/L) decreased habituation in open field arena, suggesting a non-associative learning impariment. All tested doses reduced glial fibrillary acidic protein density (GFAP) and synaptosome-associated protein (SNAP-25) without causing any changes in immunoreactivity for neuronspecific nuclear protein (NeuN) in the hippocampus and cerebral cortex. In the hippocampus, caffeine (all doses tested) increased adenosine A1 receptor density and reduced brain-derived neurotrophic factor (BDNF) and proBDNF (1.0 g/L). In the cerebral cortex, caffeine (1.0 g/L) reduced adenosine A1 receptor and increased BDNF and proBDNF density (0.3 and 1.0 g/L) These findings document the effects of caffeine consumption in adolescent rats with a dual impact on anxiety and recognition memory, associated with changes in BDNF, GFAP and SNAP-25 levels without apparent neuronal loss in hippocampus and cerebral cortex. In the second chapter, it was tested whether caffeine consumption (0.1; 0.3 and 1.0 g/L in drinking water, from 7:00 p.m. to 7:00 a.m) throughout life may reverse the negative effects caused by the consumption of caffeine in the early stages of development. For this, we used exposure protocols with the end in postnatal days (PND) 21 (development) or 90 (throughout life); both protocols starting 15 days before mating. The effects of these protocols on the behavior and hippocampal synaptic proteins density of adult female and male rats were compared. Recognition memory was impaired in females receiving caffeine (0.3 and 1.0 g/L) during development, which coincided with increased proBDNF levels and unchanged BDNF in the hippocampus. Both exposure protocols caused hyperlocomotion in males, whereas in females only the exposure throughout life significantly increased locomotor activity. Considering the anxiety related behavior, both sexes presented an anxiolytic profile when consuming caffeine (1.0 g/L) throughout life. Both exposure regimens decreased hippocampal GFAP and SNAP-25 of male rats. The hipocampal TrkB receptor was reduced in both sexes and protocols of exposure In the cortex, both proBDNF and BDNF increased in males receiving caffeine throughout life as well as GFAP and SNAP-25 increased in both treatments regimen. The results revealed that caffeine consumption throughout life can recover the impairment in recognition memory of females that consumed caffeine during development and indicate that exposure for a specific period of brain development promotes sex-dependent behavioral changes, which we relate to alterations in BDNF signaling. The results of this thesis emphasize the importance of controlling caffeine intake during critical periods of brain development of rats and points to a sex dependent effect. However, more studies are needed to expand our knowledge about the possible signaling pathways involved in these processes.

Behavioral perceptions and cortical auditory evoked potentials (CAEPs) from native English (N=10) and Spanish speakers (N=10) were recorded for speech stimuli that constitute phonetically relevant categories of voicing. The stimuli were synthesized bilabial stop consonant-vowel syllables in a continuum ranging from/ba/to/pa/that varied in voice onset time (VOT) from 0 to 60ms. Different behavioral perceptions were evidenced by significantly different categorical phonetic boundaries between the two groups (p.05). Peak P1, N1, and P2 response latencies and P1–N1 and N1–P2 amplitudes increased significantly with increasing VOTs (p