National Repository of Grey Literature 9 records found  Search took 0.01 seconds. 
Ap4A-RNA in IgE activated mast cells
Potužník, Jiří František ; Macíčková Cahová, Hana (advisor) ; Černý, Jan (referee)
Mast cells are tissue resident members of the immune system. They have a wide range of functions and receptors including the FcεRI receptor, which gets activated by binding to IgE bound to an antigen. When the cells are activated in this manner, a process termed the LysRS- Ap4A-MITF signalling pathway occurs, resulting in the translocation of the Lys tRNA synthetase into the nucleus and an activation of its moonlighting activity - the production of diadenosine tetraphosphate (Ap4A). Ap4A is a dinucleoside polyphosphate, a type of ubiquitous molecule present in all domains of life. They are made up of two nucleosides joined together by a 5' to 5' phosphodiester bridge of variable lengths. Recently, these molecules have been shown to serve as non-canonical initiating nucleotides during bacterial transcription, where they function as 5' RNA caps, similar to the well-known 7- methylguanosine eukaryotic mRNA cap. In this thesis, I present proof of existence of Ap 4A capped RNA in mast cells, a previously unknown 5' RNA structure in eukaryotic cells, and I attempt to pinpoint its role in the activation of these cells and in the wider context of mast cell mediated immune response. Keywords: mast cells, RNA caps, Dinucleoside polyphosphates, Ap 4A, RNA modification, IgE, FcεRI receptor, Lysine tRNA synthetase
Characterization of non-canonical RNA polymerase encoded by the yeast linear plasmids
Sýkora, Michal ; Vopálenský, Václav (advisor) ; Macíčková Cahová, Hana (referee) ; Valášek, Leoš (referee)
Transcription is the control point of gene expression. This process relies on protein complex of multisubunit RNA polymerases, which are extremely conserved among all cellular organisms. Transciption of extrachromosomal hereditary elements such as organelles, viruses and plasmids is dependent on host cellular RNA polymerases or intrinsic RNA polymerase is contained within these elements. Putative non-canonical two-subunit RNA polymerase is also encoded by linear cytoplasmic plasmids of the yeast Kluyveromyces lactis and most likely transcribes genes of these plasmids. Besides the two subunits of RNA polymerase encoded by linear plasmids of Kluyveromyces lactis there are another two estimated components of the transcription apparatus, namely capping enzyme that adds the cap to 5' mRNA ends and putative DExD/H box helicase. Characterization of the unique and underexplored transcription machinery of Kluyveromyces lactis plasmids was the principal objective of this work. The main goal was to: 1) clarify evolutionary origin of the linear plasmid transcription apparatus; 2) describe architecture of the linear plasmid transcription complex in vivo focused on putative RNA polymerase binding partners; 3) reveal mechanisms of transcription initiation and termination of the yeast linear plasmids. The main...
The function of 2'-O-methylated RNA in the context of viral infection
Potužník, Jiří ; Macíčková Cahová, Hana (advisor) ; Forstová, Jitka (referee)
RNA is subject to a wide array of post-transcriptional modifications. 2'-O-methylation is an essential intrinsic modification of RNA. It affects the structure and reactivity of the molecule as well as its function. 2'-O-methylation is highly conserved, present in all three domains of life. Viral RNA uses this modification to mimic the host and evade detection by the immune system. There are two main mechanisms, through which viral 2'-O-methylated RNA does this. The first is evading detection by a pattern recognition receptor form the RIG-I-like receptor family Mda5. Mda5 is capable of detecting unmethylated RNA and recognising it as non-self, thus initiating an immune response. The second mechanism the evasion and restriction of an effector molecule IFIT. IFIT proteins are capable of detecting the absence of 2'-O- methylation on viral RNAs and inhibiting their translation. They do this by interfering with the formation of the ternary complex, an essential member of ribosomal formation. Using viral 2'- O-methylation as a target for therapy, it is possible to develop attenuated vaccines. Keywords: viral RNA, RNA modifications, 2'-O-methylation, Mda5, IFIT, RIG-I-like receptors, epitranscriptomics, WNV, JEV
Redox DNA Labeling - from Simple DNA Detection by Osmium Tetroxide Complexes Modification to Ratiometric Sequence Analysis
Havran, Luděk ; Balintová, Jana ; Vidláková, Pavlína ; Macíčková-Cahová, Hana ; Pivoňková, Hana ; Hocek, Michal ; Fojta, Miroslav
DNA is electroactive molecule producing analytically useful intrinsic voltammetric signals. For some applications is a useful use redox active tag to improve specificity of the analysis. One from approaches how to prepare DNA bearing redox label is its modification by complexes of osmium tetroxide with nitrogen ligands. This method find wide used in highly sensitive DNA detection and in development of electrochemical DNA hybridization sensors. Another possibility for preparation of redox labeled DNA is incorporation of redox tags modified deoxynucleotide triphosphates by DNA polymerases. In this contribution will be demonstrated application of various redox labels in electrochemical analysis of DNA.
Magnetic Beads-Based Electrochemical Techniques for DNA-Protein Interaction Monitoring
Fojta, Miroslav ; Pivoňková, Hana ; Němcová, Kateřina ; Horáková Brázdilová, Petra ; Havran, Luděk ; Orság, Petr ; Vidláková, Pavlína ; Macíčková-Cahová, Hana ; Balintová, Jana ; Hocek, Michal
Electrochemical techniques, in connection with separation of nucleoprotein complexes at magnetic beads, are suitable for the monitoring of DNA-protein interactions. For the detection of complexes captured at the beads it is possible to utilize intrinsic electrochemical activity of the protein, intrinsic structure-selective signals of the DNA, or indicator DNA substrates tail-labeled with electroactive moieties.
Electrochemical analysis of DNA using switchable redox moieties
Fojta, Miroslav ; Daňhel, Aleš ; Horáková Brázdilová, Petra ; Plucnara, Medard ; Pivoňková, Hana ; Havran, Luděk ; Vidláková, Pavlína ; Raindlová, Veronika ; Balintová, Jana ; Macíčková-Cahová, Hana ; Hocek, Michal
Labelling of DNA with electrochemically active moieties proved to be a convenient way to the development of electrochemical techniques for the sequence-specific DNA sensing. Through combinations of various labels differing in redox potentials, independent redox coding of different DNA sequences or individual nucleobases can be attained. Applications possibilities of electrochemistry in analysis of modified DNAs are further extended by facile monitoring of chemical conversion of reactive groups on DNA during post-labelling with ultimate redox labels. In addition, controlled in situ electrochemical conversions of specific intrinsic and extrinsic DNA components can be utilized to switch their electrochemical signals and improve signal resolution.
Tail labelled oligonucleotide probes for the detection of DNA-protein interactions
Pivoňková, Hana ; Němcová, Kateřina ; Horáková Brázdilová, Petra ; Havran, Luděk ; Macíčková-Cahová, Hana ; Hocek, Michal ; Fojta, Miroslav
DNA-protein interactions can be monitored via different ways. We introduce novel, fast and simple approaches in DNA-protein interaction detection based on electrochemical measurements of DNA alone (structure-sensitive DNA sensing), or DNA modified with osmium tetroxide bearing nitrogenous ligands, or measurements of redox-active moieties enzymatically attached to the end of a DNA substrate thus forming a labeled tail (by terminal transferase).
Redox labelling of nucleic acids for analyzing nucleotide sequences and monitoring DNA-protein interactions
Fojta, Miroslav ; Havran, Luděk ; Horáková Brázdilová, Petra ; Pivoňková, Hana ; Kostečka, Pavel ; Macíčková-Cahová, Hana ; Raindlová, Veronika ; Vrábel, Milan ; Hocek, Michal
Nucleobase labelling of DNA for electrochemical sensing was attained through chemical modification of thymine bases with osmium tetroxide in the presence of nitrogenous ligands, or via enzymatic incorporation of nucleotide conjugates with redox-active moieties using labelled deoxynucleoside triphosphates. DNA hybridization, primer extension and PCR techniques were used for sequence-specific DNA assays. Tail-labelled DNA substrates were applied to monitor DNA binding by tumour suppressor p53 protein.
Polymerase construction of base-modified DNA for chemical biology
Hocek, Michal ; Macíčková-Cahová, Hana ; Kielkowski, Pavel ; Raindlová, Veronika ; Kalachová, Lubica ; Riedl, Jan ; Balintová, Jana ; Ménová, Petra
An efficient two-step methodology for construction of base-modified DNA was developed based on aqueous-phase cross-coupling reactions of halogenated nucleoside triphosphates (dNTPs) followed by polymerase incorporation. A number of diverse chemical modifications have been successfully incorporated in this way for the use in chemical biology (modulation of cleavage by restriction enzymes, interactions with diverse DNA-binding proteins and bioconjugations).

Interested in being notified about new results for this query?
Subscribe to the RSS feed.