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Role of helicases UAP56 and URH49 in viral infection
Nováková, Veronika ; Šroller, Vojtěch (advisor) ; Šmahel, Michal (referee)
Helicases are proteins with the catalytic ability to unwind double-stranded nucleic acids. An important group are helicases with a DEAD motif, which includes helicase UA56 and the more recently discovered helicase URH49. These helicases are orthologs and they share some functions. Both helicases are involved in the splicing of pre-mRNA and they take part in the transport of mRNA from the nucleus to the cytoplasm. Slight differences between the two helicases lead to different affinites for different mRNAs. Overproduction of the URH49 helicase has been reported in many cancer tissues and has therefore been suggested to function as a potential biomarker of adverse cancer prognosis. The association of helicases UAP56 and URH49 with nuclear export has led to research of their role in cells infected by viruses which replicate in the nucleus. The helicases UAP56 and URH49 have been shown to promote virus replication in several ways. They either participate in the transport of viral RNAs into the cytoplasm and thus help to translate important proteins for the virus or play a role in their encapsidation. They also help recruit the export complex, which is normally dependent on the formation of a splicing apparatus, to viral transcripts without introns. The URH49 helicase has also been described to suppress...
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Immune reactions induced by SARS-CoV-2 infection
Krausová, Kateřina ; Šmahel, Michal (advisor) ; Šroller, Vojtěch (referee)
Coronavirus disease 2019 (COVID-19) pandemic caused by newly discovered Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes severe health and economic problems all over the world. The disease severity depends mainly on the host's immune response to SARS-CoV-2. This virus uses many mechanisms for escape from the host's immune system. The major evasion mechanisms include suppression of interferon production at the early phase of infection, exhaustion of natural killer cells and induction of a cytokine storm. After the innate immune response, mechanisms of adaptive immunity join the defense against the virus. Patients with severe cases have a significant reduction in the amount of both helper CD4+ T cells and cytotoxic CD8+ T cells. On the contrary, these patients have an increased level of antibodies. Even though there have been many findings about immune reactions to SARS-CoV-2 in the year after its discovery, there are still many unknowns. Vaccines, which are successful at preventing COVID-19, have been developed in a short time. However, an important remaining question for further research is the longevity of immune memory after vaccination or after suffering from COVID-19.
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Major capsid protein of polyomaviruses and its interactions with nuclear lamins
Žáčková, Sandra ; Horníková, Lenka (advisor) ; Šroller, Vojtěch (referee)
In this study, we focused on interactions of structural proteins of mouse polyomavirus (MPyV) and BK virus (BKV) with the nuclear lamina. Our goal was to examine whether and how can the virus, hence viral structural proteins, interact with the nuclear lamina and how would these interactions affect its properties. We supposed, that the expression of viral proteins would induce disintegration of the structure of nuclear lamina, thus enabling nuclear egress of virions in the late phase of infection. Viral structural proteins were expressed transiently in cells transfected with an expression vector pMPyV LATE. In these cells, VP1 was localized in a likewise manner as it shows in infected cells - mostly in a perinuclear area. Concurrently, defects in staining of nuclear lamina were observed in these cells, similarly to infected cells. Also, another expression vector was used in our experiments, the pMPyV mut3 VP1 encoding for a mutated protein VP1. When transiently expressed in cells, the mutated VP1 protein showed mostly diffuse nuclear localization. However, we observed significant morphological deformations and defective staining of the nuclear lamina. These observations imply an important role of VP1 in mechanical and biochemical properties alterations of the nuclear lamina in transfected and...
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Therapeutic use of bacteriophages and their modifications
Vaško, Michal ; Fraiberk, Martin (advisor) ; Šroller, Vojtěch (referee)
Bacteriophages, which infect the bacteria, form a vast group of viruses, are one of the most wide-spread organisms. Since their discovery they have been used to treat various diseases caused by the bacteria, but the development of antibiotics hindered their usage significantly. Since various bacteria strains acquire multirezistance nowadays more attention is brought to the usage of bacteriophages once again. But since the application of native bacteriophages has its limitations, e.g. too broad or too narrow host range, low efficiency, they can be overcome using molecular modifications. The goal of this thesis is to provide a brief outline of the therapeutic utilization of phages in bacteriophage therapy. The majority of this thesis focuses on potential methods of acquiring modified phages and their subsequent usage. So prepared bacteriophages characterized by their specific modified properties are not only used as bactericidal reagents but also as a tool to enhance the efficiency of antibiotics, drug delivery, diagnostics and vaccine development. Various studies describe the improvement or alteration of properties gained by phage modifications. Their clinical application, however, is still limited due to small number of in vivo trials.
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Indoleamine 2,3-dioxygenase 1 inhibitors in cancer immunotherapy
Muffová, Barbora ; Poláková, Ingrid (advisor) ; Šroller, Vojtěch (referee)
The indoleamine 2,3-dioxygenase (IDO 1) enzyme is expressed in small amounts in most of the mammalian tissues, and its production is detected also in various types of tumours. IDO 1 catalyses the very first step of the kynurenine pathway, the tryptophan conversion to N-formylkynurenine, which is further metabolized to kynurenine (Kyn). The kynurenine/ tryptophan ratio (kyn/trp) may be used as a prognostic marker in research and treatment of IDO 1+ tumours. The kyn/trp demonstrates the activity of IDO 1 in tumours. The goal of cancer immunotherapy based on IDO 1 inhibition is to reverse or reduce the protumour effects of IDO 1, such as avoiding NK and T cells inhibition and activation of regulatory T cells or association with tumour-associated macrophages (TAM). IDO 1 inhibitors have been examined alone in monotherapy or together with cytotoxic T-lymphocytes antigen 4 (CTLA-4) inhibitors and programmed cell death protein 1 (PD-1) inhibitors in combined therapy. Recently, several studies are dedicated to invent inhibitors, which are able to inhibit the activity of other trp-catalysing enzymes, the indoleamine 2,3-dioxygenase 2 (IDO 2) and tryptophan 2,3-dioxygenase (TDO), together with the IDO 1 activity. Cancer immunotherapy based on IDO 1 inhibition may be combined also with chemotherapy.
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Study of exosomes in polyomavirus infection
Hyka, Lukáš ; Šroller, Vojtěch (advisor) ; Saláková, Martina (referee)
Exosomes are extracellular vesicles of endosomal origin. It was thought, that exosomes are used by cells only as carriers for cellular waste, but it was found out, that exosomes serve in the cellular communication and have a role in viral infections. Exosomes are exploited by viruses for example for the transport of viral protein or viral RNA/DNA. One of the viruses, where the mechanism of exploitation is unknown (if any exists) is murine polyomavirus. Murine polyomavirus belongs to the family Polyomaviridae, to which other human viruses belong for example, JC virus or virus of Merkel cell carcinoma. Murine polyomavirus codes for small, large and middle T antigen and three capsid proteins. Middle T antigen is known to bind to cellular membranes. Exosomes are membrane derived structures, so we investigated a possible transfer of middle T antigen. To this goal the successful isolation of exosomes and their characterization was necessary. Exosomes were isolated by ultracentrifugation and further purified by the density gradient OptiPrep. Exosomes were characterized by electron microscopy, NanoSight and by protein exosomal markers. These markers are for example Alix and flotillin-1. The cells were transfected in order to produce middle T antigen. It was shown, that exosomes isolated from these cells...
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The role of BK polyomavirus in human cancer
Cirbusová, Adéla ; Saláková, Martina (advisor) ; Šroller, Vojtěch (referee)
BK polyomavirus is small uncoated DNA virus which is ubiquitous in human population. In imunosupresed individuals can cause severe diseases, specifically BK polyomavirus associated nephropathy (BKVaN) and hemoragic cystitidis. Except that, it is presumed that BKV may be responsible for some cases of human cancer. Some features of BKV could support this idea. There are three encoded oncoproteins in BKV genome. BKV is oncogenic in rodents, where induces multiple types of tumors and it can transform cell lines as well. Moreover, BKV DNA was found in many types of human cancer. All these facts suggest a possible role of BKV in human cancer. Bladder carcinoma in pacients after transplatation and prostate adenocarcinoma are the most likely candidates to link with BKV participation. There is no complete evidence though. Therefore, future studies are necessary to proof or even exclude BKV as a possible cause of human cancer. key words: BKV, cancer, LTag, prostate, bladder
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Dengue vaccines
Holšteinová, Aneta ; Šroller, Vojtěch (advisor) ; Poláková, Ingrid (referee)
Since the middle of the 20th century the number of people infected by the dengue virus (DENV) has been constantly increasing. Specific antiviral drugs are not available, so the objective for eliminating transmission has become the construction of an effective and safe vaccine with an emphasis on inducing a balanced immune reaction against all of the DENV serotypes. Momenteraly, the only licenced vaccine is CYD-TDV (Chimeric Yellow Fever- Dengue, Live-Attenuated, Tetravalent Dengue Vaccine). This vaccine uses the structure of the effective and safe vaccine against the yellow fever virus (YFV) from which surface protein sequences are substituted for corresponding DENV sequences. While preclinical studies displayed promising results, complications have arised during clinical studies, thus creating limiting criteria for their use. For this reason the search for new vaccines that could ensure better safety from DENV for a wider range of people and replace CYD-TDV is ongoing. In the III. phase of the clinical studies there are two live-attenuated vaccines (TetraVax-DV and DENVax). The subunite vaccine (V180), DNA vaccine (TVDV) or inactivated vaccine (TDENV PIV) was forwarded into the I. phase. The combination of several vaccine schedules is also being used (tetravalent live-attenuated...
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Viruses in the pathogenesis of coeliac disease
Chudá, Kateřina ; Cinek, Ondřej (advisor) ; Šroller, Vojtěch (referee)
Celiac disease is a chronic inflammatory disorder affecting the small bowel. It develops in genetically susceptible individuals upon yet unknown environmental stimuli. Environmental triggers such as infections, dietary change or other "hits" are clearly required for disease development, as only a tiny fraction of genetically susceptible subjects develops celiac disease upon gluten exposure. This thesis aims to summarize the current evidence on viruses in the pathogenesis of celiac disease regarding their relevance in population or their involvement in immune processes leading to celiac disease. Rotavirus, orthoreovirus, adenovirus, astrovirus, respiratory syncytial virus, hepatitis viruses and herpesviruses are discussed. In addition, prospective cohort studies are presented that investigate environmental triggers of type 1 diabetes and celiac disease, two diseases sharing genetic predispositions. Keywords: celiac disease, orthoreovirus, rotavirus, adenovirus, astrovirus, respiratory syncytial virus, hepatitis C virus, hepatitis B virus, prospective cohort study
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Experimental and clinically used vaccines based on vaccinia virus
Pilná, Hana ; Mělková, Zora (advisor) ; Šroller, Vojtěch (referee)
Vaccinia virus (VACV) is an enveloped DNA virus belonging in the Orthopoxviridae genus. It is a laboratory virus in which the natural host and exact origin remain unclear. However it is of great significance for human kind. First of all, different VACV strains were used for preparation of vaccines used in the smallpox eradication campaign. Even today a significant effort is made to prepare more efficient and safer vaccines against smallpox, namely because of still remaining concerns that variola virus - causative agent of smallpox - could be misused as a biological weapon. Advances in genetic engineering allowed use of VACV for additional purposes, namely as a vaccination and expression vector. VACV enables insertion of large pieces of foreign DNA into its genome and expression of this DNA in a host. Furthermore VACV replicates exclusively in a cytoplasm, decreasing a risk of incorporation of the viral DNA into the host genome. These and other features make VACV an ideal candidate as a vector for preparation of recombinant vaccines against various infectious and oncological diseases. This thesis provides a summary of both clinically used and experimental vaccines derived from VACV. Powered by TCPDF (www.tcpdf.org)
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