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Tumor cell lines viability testing after exposure to chemicals and chemotherapeutics
Horáčková, Lucie ; Zemanová, Jana (referee) ; Brázda, Václav (advisor)
Individual types of viability tests based on colorimetric changes of the solution are desribed in the theoretical part. Furthermore, HSP proteins are characterized, which are not connected only by heat shock, but also during other cell stresses such as exposure to UV, cold, extreme pH or heavy metals. They are important for the cell, because they help to reformulate proteins that have been damaged by cellular stress and also bind to new unpacked proteins and ensure their correct folding. Proteins that are affected by molecular chaperones are collectively called client proteins. Some HSPs also contribute to membrane transport or degradation. These proteins are co-operative with the cochaperones, which are important for heat shock proteins because they help them to pack protein, in particular by catalyzing the hydrolysis of ATP to ADP. Herein is also described cisplatin and its derivatives, including mechanism of action and adverse effects. This work was focused on detection cytotoxicity of cisplatin and its derivatives. Cells were exposed to stress condition induced by cytostatics and huge changes in heat shock proteins and cochaperon levels were observed. There was also observed colocalization of heat shock proteins and their client protein p53 by confocal microscopy in these stressing conditions.
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HSP90 as a target for antitumor therapy
Drápalová, Kateřina ; Zatloukalová, Pavlína (advisor) ; Dibus, Nikol (referee)
Stability and correct tertiary structure of proteins are necessary for maintaining cellular homeostasis. The cell uses molecular chaperones, including Hsp90, to achieve this balance. Hsp90 is an essential protein for healthy and cancer cells. Overexpression of this chaperone is noticeable in many cancers. This thesis summarizes current understandings of the Hsp90 protein and its role in carcinogenesis. Hsp90 became a target of anticancer therapy in the 90s. Inhibition of this chaperone, though successful in many studies, has not yet reached the wanted results in clinical practice. Current failures are mostly caused by high hepatotoxicity and other side effects accompanying the therapy, or the inhibition having insufficient antitumor effects. Nevertheless, the inhibition of the Hsp90 protein represents an interesting approach in antitumor therapy. New inhibitors are constantly being developed and tested in monotherapy or in combination therapy, which demonstrates significantly higher efficacy and, thanks to the synergistic effect, enables the application of a lower concentration of therapeutics.
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Antioxidant system of tomato plants exposed to triazole fungicides
Žufić, Antoniana ; Hýsková, Veronika (advisor) ; Jaklová Dytrtová, Jana (referee)
Fungal pathogens cause severe crop losses worldwide. Recently, however, chemical fungicides have become dangerous environmental contaminants affecting non-target organisms. The aim of this study was to investigate the effect of triazole fungicides in the form of penconazole (P), tebuconazole (T) or their combination (PT) on tomato plants Solanum lycopersicum L. cv. Cherrola. No significant change in the activity of a key enzyme of the ascorbate-glutathione cycle, ascorbate peroxidase and activity of cytosolic peroxides, was detected in leaves, roots or fruits at the intervals studied, i.e. after two and five weekly treatments of P, T or PT, either as a foliar or a soil application. While slightly increased guaiacol peroxidase activity was found in roots due to T spraying and PT combination, the activity of this enzyme was reduced in leaves similar to catalase. The greatest changes in leaf antioxidant capacity correlated with increased phenolic and flavonoid content were observed after five treatments of penconazole (P) as a soil application both P alone and in combination as PT. The studied triazoles significantly affected the abundance of isoforms of superoxide dismutase and partially total peroxides in leaves, roots and fruits and of acorbate peroxidase in leaves. The highest content of HSP70 was found in...
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The effect of Hsp70 protein on the infection caused by Potato virus Y
Doričová, Vlasta ; Hýsková, Veronika (advisor) ; Hoffmeisterová, Hana (referee)
Whithin their natural environment, plants are subjected to a combination of stress conditions. Since potential interactions between signal pathways, plants respond to multiple stresses differently from how they do to individual stresses, activating a specific programme. Heat shock proteins (HSP70) overexpressed after heat shock influence the viral infection. On one side HSP70 can participate on refolding of aggregated or partially denaturated proteins, on the other side HSP70 can interact with viral proteins and facilitate propagation of viral replication complexes. In this work the effect of heat shock (42řC, 2. hours) applied before or after the inoculation of plants Nicotiana tabacum L. cv. Petit Havana SR1 with Potato virus Y on viral infection was detected. This effect was studied in two biological experiments. The amount of coat protein of PVYNTN and protein HSP70 were detected simultaneously with the activity assays of Hatch-Slack cycle enzymes, glycosidases and peroxidase. Both experimental approaches (heat shock applied before or after the inoculation by PVYNTN ) enhanced amount of the virus and in the 2nd experiment it accelerated infection development. Immediately after application of heat shock the amount of HSP70 was increased. The enhancement of HSP70 by viral infection occurred...
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Role of molecular chaperones Hsp70 and Hsp90 in the replication cycle of DNA viruses
Žáčková, Sandra ; Horníková, Lenka (advisor) ; Poláková, Ingrid (referee)
Molecular chaperones are proteins which enable other proteins to assemble into native conformation and are essential for viability of the cells. Chaperones of the Hsp70 family bind to newly synthetized and denaturated proteins, prevent their aggregation and facilitate their assembly. They participate in assembly and disassembly of oligomers and also in the transport across the membranes. Chaperones of the Hsp90 family do not participate in the assembly of nascent or denaturated proteins. They bind proteins which are nearly in native conformation and enable them to assemble into conformation suitable for ligand binding or interacting with other proteins. These attributes predestinate chaperones to participate in the replication cycle of DNA viruses. A huge amount of proteins is translated during viral infection. These proteins require the chaperones to facilitate their assembly and are also required for assembly into oligomers and macromolecular structures. In addition to capsid assembly the chaperones also participate in transport of genetic information to the sites of replication, disassembly of incoming viral particles or replication of viral DNA. Therefore, the development of specific chaperone inhibitors is a promising approach. They could be used against broad spectrum of viral infections...
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Tumor cell lines viability testing after exposure to chemicals and chemotherapeutics
Horáčková, Lucie ; Zemanová, Jana (referee) ; Brázda, Václav (advisor)
Individual types of viability tests based on colorimetric changes of the solution are desribed in the theoretical part. Furthermore, HSP proteins are characterized, which are not connected only by heat shock, but also during other cell stresses such as exposure to UV, cold, extreme pH or heavy metals. They are important for the cell, because they help to reformulate proteins that have been damaged by cellular stress and also bind to new unpacked proteins and ensure their correct folding. Proteins that are affected by molecular chaperones are collectively called client proteins. Some HSPs also contribute to membrane transport or degradation. These proteins are co-operative with the cochaperones, which are important for heat shock proteins because they help them to pack protein, in particular by catalyzing the hydrolysis of ATP to ADP. Herein is also described cisplatin and its derivatives, including mechanism of action and adverse effects. This work was focused on detection cytotoxicity of cisplatin and its derivatives. Cells were exposed to stress condition induced by cytostatics and huge changes in heat shock proteins and cochaperon levels were observed. There was also observed colocalization of heat shock proteins and their client protein p53 by confocal microscopy in these stressing conditions.
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Detection of Hsp70 protein in plants exposed to various stress factors.
Lengálová, Alžběta ; Hýsková, Veronika (advisor) ; Liberda, Jiří (referee)
! Plants are continuously exposed to various stress conditions. Being sessile, they are not able to escape from adverse conditions. Therefore, they have developed specific defence mechanisms. Most studies focus on plant responses to a single type of stress. However, plants in nature must cope with a variety of stresses at the same time. In this work, the effects of heat shock on the interaction of tabaco (Nicotiana tabacum L.) with the Potato virus Y (PVY) were investigated. Obviously, heat stress is associated with the synthesis of Hsp70 protein, which has many important functions alleviating adverse effects of stress conditions (e.g. Hsp70 participates in refolding or degradation of damaged proteins and protein syntetis de novo). The effect of Hsp70 during viral infection is still not fully understood, some studies revealed Hsp70 as a part of viral multiplication and transport processes in plant. In the first experiment performed in this work, higher levels of PVYNTN virus was found in tobacco plants that have been exposed to heat shock after inoculation than in plants only infected. The amount of the virus corresponded with the amount of Hsp70 protein detected immunochemically using a specific antibody. It seems that the plant response to combination of heat stress with viral infection is a...
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The effect of Hsp70 protein on the infection caused by Potato virus Y
Doričová, Vlasta ; Hýsková, Veronika (advisor) ; Hoffmeisterová, Hana (referee)
Whithin their natural environment, plants are subjected to a combination of stress conditions. Since potential interactions between signal pathways, plants respond to multiple stresses differently from how they do to individual stresses, activating a specific programme. Heat shock proteins (HSP70) overexpressed after heat shock influence the viral infection. On one side HSP70 can participate on refolding of aggregated or partially denaturated proteins, on the other side HSP70 can interact with viral proteins and facilitate propagation of viral replication complexes. In this work the effect of heat shock (42řC, 2. hours) applied before or after the inoculation of plants Nicotiana tabacum L. cv. Petit Havana SR1 with Potato virus Y on viral infection was detected. This effect was studied in two biological experiments. The amount of coat protein of PVYNTN and protein HSP70 were detected simultaneously with the activity assays of Hatch-Slack cycle enzymes, glycosidases and peroxidase. Both experimental approaches (heat shock applied before or after the inoculation by PVYNTN ) enhanced amount of the virus and in the 2nd experiment it accelerated infection development. Immediately after application of heat shock the amount of HSP70 was increased. The enhancement of HSP70 by viral infection occurred...
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The effect of abiotic stress on cucumber plants (Cucumis sativa L.)
Plisková, Veronika ; Hýsková, Veronika (advisor) ; Müller, Karel (referee)
The exposure of plants to high salt concentrations causes accumulation of sodium ions. This leads to the inability of the plants to uptake water, a disturbance of ion homeostasis, a decrease in photosynthesis and oxidative stress. As a result of the salt stress, the availability of NADPH decreases. The adaptation to the concentrations of salt depends on plant's ability to compensate for the decreased availability of NADPH, which can be further used in antioxidative cycles and the synthesis of antioxidative compounds and osmoprotectants. In this work, the reduction of relative water content, a decrease in the Rubisco enzyme activity, an increase of Hsp70 in the leaves and an increase in the accumulation of sodium ions was shown in cucumber plants (Cucumis sativa L. convar. Jogger F1) exposed to salt stress (100 mM NaCl). As a consequence of salt stress, an increase in the activity of NADPH providing enzymes was found. Particularly on the second and third day of salt stress, an increase in the activity (up to 270 %) of: NADP-isocitrate dehydrogenase, glucose-6-phosphate dehydrogenase, NADP-malic enzyme, non-phosphorylating glyceraldehyde-3-phosphate dehydrogenase in leaves was detected. The activity of less abundant NADP-dehydrogenases (glucose 1-dehydrogenase, gluconate 2-dehydrogenase, galactose...
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