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The role of Spr1851 protein Streptococcus pneumoniae in cell division
Jarošová, Václava ; Ulrych, Aleš (advisor) ; Seydlová, Gabriela (referee)
The role of Spr1851 protein Streptococcus pneumoniae in cell division Human extracellular pathogen S. pneumoniae encodes unique serin/threonine protein kinase of Eucaryotic type StkP and its cognate phosphatase PhpP. This kinase affects number of cellular processes including virulence, competence, cell division and cell wall synthesis by phosphorylating its substrates. Hypothetical protein Spr1851 named Jag was identified as a new StkP substrate in the membrane fraction by comparing the wild-type phosphoproteomes with StkP deleted strain. This protein consists of three domains and an interdomain region that contains T89 phosphorylation site. There is a Jag_N domain with an unknown function at the N-terminus. The C-terminus contains two domains - KH and R3H, which are highly conserved and their expected function is binding of nucleic acid. The main aim of this work is to explain the function of Jag protein, to determine the effect of individual domains on the phenotype and the localization of the protein and to determine the role of phosphorylation on T89. The results confirm that Jag protein could play a role in cell division or cell wall synthesis. Furthermore, the results indicate that the Jag_N domain is essential for the localization of the protein into the membrane, whereas the KH and R3H domains are...
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Metabolic control of bacterial division.
Valtová, Aneta ; Lichá, Irena (advisor) ; Fišer, Radovan (referee)
Metabolic control of cell cycle has been study for a long time, but it is not completely known. Mechanisms of metabolic control described for a several decade has been explained on molecular level with using a modern methods. Regulation of cell cycle in consideration of metabolism and nutritional status is going on at the several level of cell replication. The most known is about assembly of bacterial cell divisiome. Changes in nutrient availability induce stress response that use low-molecular substances in signaling pathways leading to changes in the cell cycle. One of the most studied is (p)ppGpp that participates in stringent response and affect sigma factors, directly inhibits the initiation of replication by binding to the DnaG primase and indirectly inhibits the elongation of replication. Current researches has revealed that some enzymes with already known enzymatic function in the major metabolic pathways (glycolysis or TCA) also has a function as sensors that transmit the nutritional change signal directly into the cell dividing process. These signals most often inhibits FtsZ protein or affect its helper proteins and subsequent ring formation. Analogues of these enzymes were found in gram-positive (Bacillus subtilis) and gram-negative bacteria (Escherichia coli, Caulobacter crescentus)....
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The role of protein kinase StkP in regulation of the cell division in Streptococcus pneumoniae
Malíková, Eliška ; Doubravová, Linda (advisor) ; Kuthan, Martin (referee)
Protein phosphorylation by protein kinases is a key mechanizm that enables both eukaryotic and prokaryotic organizm sense and read environmental signals and convert these signals into changes in gene expression and thus proper biological response. One of the main phosphorylation systems in bacteria consists of eukaryotic-like Ser/ Thr protein kinases. The genome of human pathogen Streptococcus pneumoniae contains single Ser/ Thr protein kinase StkP. StkP regulates virulence, competence, stress resistance, gene expression and plays an important role in the regulation of cell division cycle. Analysis of phosphoproteome maps of both wild type and ΔstkP mutant strain of S. pneumoniae showed that in vivo StkP phosphorylates several putative substrates including the cell division protein DivIVA (NOVÁKOVÁ et al., 2010). DivIVA in S. pneumoniae is localized at midcell and at the cell poles. It was proposed to be primarily involved in the formation and maturation of the cell poles (FADDA et al., 2007). The aim of this thesis was to investigate phosphorylation of the cell division protein DivIVA in S. pneumoniae. Gene divIVA was cloned, expressed in E. coli and protein was purified via affinity chromatography. Phosphorylation of DivIVA by StkP was examined in a kinase assay. We confirmed that DivIVA is a direct...
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Non-conventional bacterial signaling pathways
Krupička, Jiří ; Branny, Pavel (advisor) ; Beranová, Jana (referee)
Two component systems were traditionally considered as main phosphorylation systems of bacteria involved in cell signalling. Recently, attention focuses increasingly on bacterial eukaryote-like Ser/Thr protein kinases (eSTKs). These protein kinases are structurally similar to their eukaryotic counterparts. Some eSTKs possess additional domains such as extracellular PASTA domains that were discovered in a variety of gram-positive bacteria. It has been proved that these domains can act as sensors for unlinked peptidoglycan fragments. However, majority of environmental signal molecules still remains unknown. eSTKs phosphorylate a broad spectrum of substrates including proteins involved in various cell processes such as virulence, cell wall biosynthesis, cell division, and central and secondary metabolism. Cross talk between eSTKs and two component systems also occurs. In this thesis, the current knowledge about eSTKs and their significant substrates in different bacterial species is discussed.
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Spr0334, new protein of cell division in Streptococcus pneumoniae.
Štekerová, Nela ; Doubravová, Linda (advisor) ; Konopásek, Ivo (referee)
Spr0334, new protein of cell division in Streptococcus pneumoniae Streptococcus pneumoniae is an important human pathogen. The geonome of this bacteria encodes a single gene for eukaryotic-like serine / threonine protein kinase called StkP. StkP regulates many physiological processes such as pathogenesis, competence for genetic transformation, resistance to various stresses and resistance to antibiotics. It also affects the transcription of many genes involved in cell wall biosynthesis, pyrimidine metabolism, DNA repair and iron uptake. Recent studies have shown that StkP is located in the cell division septum and significantly regulates cell division and morphology. Its substrates include, among others, cell division protein DivIVA, FtsZ and FtsA. Analysis of phosphoproteome maps of wild type and ΔstkP mutant strain of S. pneumoniae showed that in vivo StkP phosphorylates several putative substrates including the protein Spr0334. Mass spectrometry analysis identified phosphorylation sites of the protein Spr0334: threonine 67 and threonine 78. Furthermore, it was found that the protein Spr0334 is located in the cell division septum, which led to the hypothesis that it could be newly identified cell division protein in S. pneumoniae. The main aim of this thesis was to describe the function of the...
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Live-cell tracking in time-lapse sequences
Zámečník, Tomáš ; Šorel, Michal (advisor) ; Křivánek, Jaroslav (referee)
Title: Live-cell tracking in time-lapse sequences Author: Tomáš Zámečník Department: Department of Software and Computer Science Education Supervisor: RNDr. Michal Šorel Ph.D., Oddělení zpracování obrazu ÚTIA AV ČR Abstract: This diploma thesis deals with methods of tracking particles in image sequences. It's goal is to design and implement a complete system for tracking of live cells, their motion and division. The thesis uses conclusions of published scientific papers, studies their application and analyzes possibilities for their mo- difications or improvement. As a result, there are two applications. First of them is a demonstrational pro- gram, provided as an attachment of this thesis. Second implementation is a mo- dule of commercial software NIS-Elements, by Laboratory Imaging, Ltd., which is used by both scientific and commercial institutions in the whole world. Keywords: cell tracking, particle tracking, cell division 1
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24-Epibrassinolide at Subnanomolar Concentrations Modulates Growth of Mouse Hybridoma Cells
Franěk, František ; Kohout, Ladislav ; Eckschlager, T.
Brassinolides are known to stimulate plant growth and to posses antistresiTactivities in plants. This work was aimed at exploring possible beneficial effects of 24-epibrassinolide on cultured mammalian cells. A mouse hybridoma, i.e. a hybrid ymphocyte cell line, was cultured either in standard serum-free medium, or in diluted medium in which the cell grew inder nutritional stress. Steady-state parameters of semicon-finuous cultures conducted at 24-epibrassinolide concentra-ions from 10~16 M to 1CT9 M were evaluated. Typical effects of he agent that were observed both in standard and in diluted i [media were suppression of intracellular antibody level, increa- / ,ed value of mitochondrial membrane potential, higher frac- / ion of cells in the G(/Gl phase and lower fraction of cells in / he S phase. Alleviation of nutritional stress was observed in j (cultures conducted in diluted media. Viable cell density was j ignificantly higher at 24-epibrassinolide concentrations 10~13[ knd 10~12 M. Results of this exploratory study show that thej plant hormone 24-epibrassinolide may induce alterations of he cell division mechanism of the energetic metabolism, and of secreted protein synthesis in a mammalian cell line.
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