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Wingless/Notch crosstalk in Drosophila melanogaster
Hajšmanová, Hana ; Mašek, Jan (advisor) ; Macůrková, Marie (referee)
Fundamental for the existence of multicellular organisms is the presence signalling pathways, complex biochemical cascades, allowing reciprocal communication amongst the cells. Both Notch and Wnt pathways are crucial for embryonic development and their deregulation leads to many severe diseases. Decades of research, largely based on model organism D. melanogaster, showed that components of these pathways do not function strictly separately but on contrary interact with each other (crosstalk). This phenomenon is explored only to a lesser extent in mammals. The aim of this thesis is to recapitulate existing findings of Wnt/ Notch crosstalk in D. melanogaster and outline where crosstalk could play a role in mammals.
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Identification of novel mechanisms controlling emergency granulopoiesis in hematopoietic stem and progenitor cells
Vaníčková, Karolína ; Alberich Jorda, Meritxell (advisor) ; Zadražil, Zdeněk (referee)
Granulocytes represent the first line of defense against bacteria and fungi. Daily production of granulocytes is sustained by steady state granulopoiesis but under stress (e.g., bacterial infection) this program switches to emergency granulopoiesis (EG) which ensures the production of granulocytes at enhanced and accelerated rates. Very little is known about the regulation of EG. In this thesis, we showed that disruption of the β-catenin-TCF/LEF mediated transcription impairs EG in vivo. Further, we demonstrated that lipopolysaccharide (LPS) administration in mice induces accumulation of active β-catenin in hematopoietic stem and progenitor cells (HSPCs) as early as 4 hours (H) after stimulation, with highest increase at 24H. This effect was at least partially mediated in a niche independent manner, since LPS stimulation in vitro induced β-catenin accumulation in c-Kit+ cells after 2H, with a peak activation at 4H. Using single cell RNA sequencing, we determined the cell cluster dynamics of HSPCs following 4H LPS stimulation. Interestingly, we identified a possible upstream activator of β- catenin in one of the clusters - Wnt10b. Indeed, Wnt10b showed a similar expression pattern as EG master regulator Cebpb and β-catenin activation, following in vitro treatment with LPS. Altogether, our data point...
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Function and regulation of ETV4 and MSX1 transcription factors in colon cancer progression
Hrčkulák, Dušan ; Kříž, Vítězslav (advisor) ; Libusová, Lenka (referee)
Colon cancer causes approximately seven percent of all cancer-related deaths in the world and presumably due to modern lifestyle, it is also one of the most frequently diagnosed cancers. The inefficiency of standard treatment indicates the need for intensive research of molecular mechanisms of cancer development. The canonical Wnt signaling pathway is essential for maintenance of the progenitor phenotype of stem cells in crypts of the intestine and controls repopulation of the epithelia, in physiological conditions. However, aberrant activation leads to tumor formation. Although Wnt signaling in cancer has been subjected to thorough investigation, there is still a lot of questions concerning further branching of the pathway. As a model of Wnt/β-catenin triggered colorectal cancer, we use mice with mutated APC, which is the tumor suppressor involved in this pathway. Previous expression profiling of the intestinal tumors from relevant mice revealed two transcription factors: ETV4 and MSX1 which are significantly overexpressed in cancer cells. In this project we elucidate whether the overexpression is really tumor restricted and Wnt dependent or there is a crosstalk with another signal transduction pathway. We investigate the function and regulation of these transcription factors by synthetic reporter assays,...
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Adjusting Wnt signaling, new regulatory mechanisms of the Wnt pathway
Fafílek, Bohumil ; Kořínek, Vladimír (advisor) ; Stopka, Tomáš (referee) ; Machoň, Ondřej (referee)
4 Abstract The Wnt pathway is one of the major signaling cascades contributing to multiple cellular processes during embryogenesis, and adult tissue homeostasis and regeneration. Moreover, aberrant activation of the Wnt signaling pathway is connected with development of neoplasia, notably colorectal cancer. The aim of the thesis was to identify new ways of the Wnt pathway regulation to understand better physiological as well as non-physiological mechanisms of Wnt signaling. The results are summarized in four publications. The first article deals with TROY, a member of tumor necrosis factor receptor family. We identified TROY as a Wnt target gene during our search for Wnt responsive genes in colorectal cancer cell lines. Additionally, we detected expression of Troy in tumors of two mouse models of intestinal cancer. In the healthy gut, Troy is produced in fast cycling intestinal stem cells where negatively regulates the Wnt pathway. The second study focuses on processing and posttranslational modification of murine Wnt1 and Wnt3a. Wnts are glycosylated and double acetylated by lipid adducts and our results revealed that O-linked acylation of serine is required for the subsequent S-palmitoylation of cysteine. Moreover, acylation of Wnts is connected with their signaling activity which is related to Wnt1 and...
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Changes in beta-catenin expression during ontogenesis in the transgenic minipigs for human mutant huntingtin
Žižková, Martina ; Motlík, Jan (advisor) ; Jarkovská, Karla (referee)
Huntington's disease (HD) is an inherited autosomal dominant neurodegenerative disorder caused by an unstable expansion of the CAG repeat sequence within the huntingtin gene. Huntingtin associates with ubiquitin-proteasome system that ensures degradation of particular proteins including β-catenin which is an important molecule whose equilibrated degradation is necessary for the proper functioning of the Wnt signaling pathway. The binding of β-catenin to the destruction complex is altered in HD, leading to the toxic stabilization of β-catenin. The main goal of my thesis was to determine whether the accumulation of β-catenin due to the presence of mutant huntingtin is also characteristic of Liběchov minipigs, a large animal model of Huntington's disease stably expressing N-truncated human mutant huntingtin. Using immunoblot and specific antibodies, we have revealed age-dependent accumulation of mutant huntingtin in transgenic minipigs. Unlike endogenous huntingtin, no decrease of the level of mutant huntingtin was observed in the striatum of transgenic animals. Surprisingly, this was followed by a decrease of phosphorylated β-catenin. Nevertheless, our results demostrate the accumulation of β-catenin in mesenchymal stem cells isolated from the oldest boars during ontogenesis. Furthermore, we have revealed a...
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