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Experimental system for the mouse polyomavirus life cycle study
Pergner, Jiří ; Španielová, Hana (advisor) ; Mašek, Tomáš (referee)
Experimental system for the mouse polyomavirus life cycle study Abstract: Murine polyomavirus (MPyV) is the prototype of the Polyomaviridae family. This family includes also some important human pathogens (BKV, JCV, Merkel cell polyomavirus). Due to their specific properties viruses within this family may serve as versatile vectors for gene therapy or recombinant vaccine production. New methodological approaches may help to understand some yet unknown facts about MPyV life cycle. Clarification of some processes during murine polyomavirus life cycle may be also important to fully exploit polyomaviruses for therapeutic purposes. The aim of this diploma thesis was to preparare two innovative experimental systems that extend possibilities of studying the life cycle of MPyV. The first part of the diploma thesis focusses on construction of recombinant MPyV which expresses yellow fluorescent protein (EYFP) in the early stages of infection. Such virus can be very useful for studying the infection spreading by live- cell imaging and Fluorescence-Activated Cell Sorting (FACS) and can be employed for co- localization studies of YFP-tagged LT antigen with certain cellular proteins. Second part of the diploma thesis describes preparation of a hybrid cell line prepared by fusion of mouse and monkey cells. This new cell...
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Alterations of the oncogenic properties of tumor cell lines by modulating oncogene expression. Example of v-src transformed chicken cell lines
Kovářová, Denisa ; Hejnar, Jiří (advisor) ; Španielová, Hana (referee)
4 Alterations of the oncogenic properties of tumor cell lines by modulating oncogene expression. Example of v-src transformed chicken cell lines Abstract v-Src protein tyrosine kinase is the product of the transforming gene transduced by avian Rous sarcoma virus. In contrast to proto-oncogene c-src, v-src lacks the negative-regulatory C-terminal domain and consequently shows a higher level of activity and transforming ability. In addition, v-Src contains point mutations throughout its coding region that probably contribute to the high level of intrinsic kinase activity. Long terminal repeats (LTR) comprise strong promoter-enhancer sequences and ensure efficient expression of the v-src gene. v-Src protein has a strong transforming potential in vitro and induces tumor development and growth in vivo. Moreover, it is implicated in metastatic formation. In several cancer types the elevated c-Src kinase activity caused pleiotropic cellular responses inducing transformation and metastasis. The aim of this diploma work was to reveal the role of v-Src in mediating tumor and metastatic progression in chicken cell lines PR9692 and PR9692-E9. Despite the low propensity of the chicken cells to immortalization, comparatively high immoratlization efficiency was observed in cells from ex vivo tumours growing progressively...
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Implication of eukaryotic DNA repair machinery in viral replication
Hron, Tomáš ; Španielová, Hana (advisor) ; Harant, Karel (referee)
Eukaryotic DNA damage response is an important mechanism which ensures genome stability. Its components are also mobilized during viral infection as a reaction against extraneous nucleic acid. Additionally, DNA repair machinery seems to be activated by some viruses purposely to provide their replication. This activation is mediated mainly by viral proteins which are able to interact with cellular factors. In many cases, key components of DNA damage mechanisms are associated with viral replication centre and likely participate in this process. Furthermore, cellular DNA damage signaling is exploited to provide competent environment for viral reproduction. However, particular mechanisms how these cellular factors participate in viral infection are still largely unclear. In this thesis, the principles of relationship between viral infection and eukaryotic DNA damage response are summarized and main viral families which are known to activate and utilize these mechanisms for its genom replication are described.
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New prevention strategies in HIV infection
Buriánková, Lucie ; Španielová, Hana (advisor) ; Drda Morávková, Alena (referee)
This bachelor theses deals with the new preventive methods in the fight against HIV (Human Immunodeficiency Virus). Chapter 1 is aimed at the HIV characterization. The next chapters dwell on the preventive methods, whether these are ABC methods (abstinence from sex, being faithful, use of condoms), which belonging to the old preventive strategy in HIV struggle or innovation methods like male circumcision, vaginal diaphragm, microbicides and HIV vaccines. The final chapter deals with the situation in Czech Republic and various ways of educational training at czech schools.
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Viral and cellular RNA-dependent RNA polymerases
Krupicová, Daniela ; Španielová, Hana (advisor) ; Lišková, Petra (referee)
Viral and cellular RNA dependent RNA polymerases Krupicová Daniela Abstract RNA dependent RNA polymerases synthesize RNA strand from RNA template. They can be divided into two large groups - viral (RdRp) and cellular (RDR) polymerases. Viral RdRp play a key role in the life cycle of RNA viruses, ensuring replication and transcription of their genome. The structure, which resembles the "right hand", is similar to other types single-subunit polymerases. Large amounts of knowledge are available about the RdRp of RNA viruses with positive single strand RNA genome. This information then helps in the development of antiviral drugs targeting the RdRp. The second group consists of cellular RDR. These polymerases are found only in eukaryotes. Their active site is homologous to the active site of multisubunits DNA dependent RNA polymerases. RDR is involved in RNA silencing pathways.
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Cell Surface Markers in Colorectal Cancer
Červená, Klára ; Španielová, Hana (advisor) ; Plocek, Vítězslav (referee)
Colorectal cancer is the most frequent cancer in the Czech Republic. Retrospective analysis of patients with colorectal cancer have led and still lead to the discovery of new biomarkers that may be important in treatment, prediction or prognosis of cancer. The main aim of this thesis was to desribe discovered cell surface proteins (biomarkers) of colorectal cancer, that correlate with colorectal cancer progression, metastasis and patient outcome, and discusses the possibilities of their utilization as cancer cell targets for theranostic purposes. Keywords: colorectal cancer, biomarkers, targeting, theranostics, prediction, prognosis
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Innovative method of HIV/AIDS prevention in high schools
Peterková, Julie ; Drda Morávková, Alena (advisor) ; Španielová, Hana (referee)
A B S T R A C T Almost every day each of us can read or hear in the media a lot of distorted information concerning HIV or AIDS. Therefore it is necessary to have knowledge about the possible risks of HIV transmission and all ways the HIV can be transferred. These attitudes and preventive measures of knowledge should be developed in high school students who are experiencing very rapid physical and psychosocial growth. Sufficient amount of information available in the problem group of 15-24 year olds has a positive impact on their sexual behavior and thus on prevention. My research methods of knowledge about HIV were done in two schools, one is an eight-year high school, where 24 students of fifth year were questioned and second is a four- year high school, where 24 students of first year were questioned. I used a cross-sectional pre-test and post-test. Students of the fifth year were taken deeper into the problems through creation of posters on topics connected with the problem of HIV. The aim of my research is to develop proposals for the scientific conference, which will be lead by the students and will be accompanied by the created images (posters). This is a non-violent way of repeating the HIV prevention. Creating a pre-test and post-test to determine the extent of knowledge gained after completing the...
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TOR signalling in yeast
Šimek, Jan ; Palková, Zdena (advisor) ; Španielová, Hana (referee)
TOR ("Target of rapamycin") protein, a highly conserved Ser/Thr protein kinase, is a central component of signalling network that controls cell growth in diverse eukaryotic organism, ranging from yeast to man. TOR proteins were first identified in yeast Saccharomyces cerevisiae in 1991 as the targets of the antifungal and immunosupressive agent rapamycine. In contrast to most eukaryotes, yeast contains two TOR homologues , Tor1p and Tor2p. These proteins are components of multiprotein complexes TORC1 and TORC2. TORC1 is specifically inhibited by rapamycine and controls cell growth in response to quality of the available nutrients. TORC2, which is insensite to rapamycine, regulates actin polymerization, sphingolipid biosynthesis and endocytosis. This work is focused on description of both TOR complexes, especially on downstream and upstream regulation of TORC1. Powered by TCPDF (www.tcpdf.org)
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Expression of sTGFbeta RII-Fc-Jun from recombinant vaccinia virus
Samková, Zuzana ; Němečková, Šárka (advisor) ; Španielová, Hana (referee)
Expression of sTGFbetaRII-Fc-Jun from recombinant vaccinia virus TGFß has a biphasic role in tumorigenesis. In early phases it acts as tumor sup-pressor. However, in late phases when cells have escaped selectively from the antimito-genic response of TGFß, it may act as a promoter of tumor progression and invasion. One way of control tumor formation and progression is blocking of TGFß signalling pathways in late phases of tumorigenesis. We have constructed recombinant vaccinia virus P13 expressing soluble TGFbeta type II receptor fused with the Fc fragment of IgG1 and with Jun fragment (sTbetaRII-Fc-Jun). This sTbetaRII-Fc-Jun is supposed to increase the effect of antitumor vaccinia virus vaccine expressing SigE7LAMP, which is investigated for the treatment of the HPV-16 associated cervical cancer. Binding of sTbetaRII-Fc-Jun to protein G were tested by SDS-PAGE and by im-munoblotting. We found that Jun fragment and sTbetaRII fragment do not block Fc bind-ing site for protein G. sTbetaRII-Fc-Jun was characterised using SDS-PAGE and immunoblot analysis. We observed that the amount of sTbetaRII-Fc-Jun was higher in cell supernatans of in-fected cells in comparison to cell lysates. In cell lysates we observed higher amount of sTbetaRII than sTbetaRII-Fc-Jun. The expression of sTbetaRII-Fc-Jun was stronger under...
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Human endogenous retrovirus ERVWE1: transcriptional activation and modifications of promoter DNA methylation
Dobšová, Martina ; Trejbalová, Kateřina (advisor) ; Španielová, Hana (referee)
Endogenous retrovirus ERVWE1 is an integral part of the human genome. In the course of evolution, a protein encoded by the env gene of this retrovirus - Syncytin-1 - has gained unique function in human development. It mediates cell-to-cell fusion of placental cytotrophoblasts. Receptor that binds to Syncytin-1 is expressed in different cell types. Syncytin-1-mediated fusion is essential in placenta, but it can cause disruption of tissue integrity in other cell types. ERVWE1 expression is regulated by promoter DNA methylation, transcription factor GCM1 and efficient mRNA splicing. This thesis concerns the ERVWE1 expression and its regulation in non-placental tissues. It was found out that the moderate GCM1 overexpression was not sufficient to induce Syncytin-1 expression. Neither treatment with DNA demethylation agent 5-azacytidine nor with Syncytin-1 activator forskolin was able to manage Syncytin-1 expression. This thesis extends previous findings concerning high syncytin-1 expression in seminomas. In same tissues, there was found elevated TET1 expression on mRNA level in comparison with controls. The presence of the TET1 demethylation enzyme can influence ERVWE1 promoter DNA methylation. Previously unreported splicing variant of TET1 has been found during the construction of human TET1 expression...
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