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The role of Src family kinases in RNA processing
Gemperle, Jakub ; Rösel, Daniel (advisor) ; Mácha, Jaroslav (referee)
Until now, a lot of information have been obtained about the role of Src family kinases in the cytoplasm or at the plasma membrane and their interactions with growth factor receptors or focal adhesion complexes. Their functional importance at the perinuclear membrane, or even inside the nucleus, however, has not been well characterized. This work, using available information, pointed at the fact that Src family kinases can be found in the nucleus. This opens a new field of Src kinases action, such as in RNA metabolism, considering that it has been assumed that their activity is limited to the cytoplasmic compartment. This work summarizes the current knowledge that hints to Src family kinases dependent network of regulation of RNA metabolism; Src family kinases have pleiotropic effects not only on the RNA binding proteins, but also on the remodeling of chromatin structure. These kinases affect by direct interactions with other proteins transport, splicing or RNA stability and gene expression. This summary suggests that Src family kinases could regulate RNA metabolism on many levels.
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The role of NG2 glycoprotein in regulation of Rho/ROCK signaling
Kratochvílová, Magdalena ; Rösel, Daniel (advisor) ; Kuželová, Kateřina (referee)
NG2 is a transmembrane glycoprotein, which takes part in cellular processes such as adhesion, migration or invasivity, i.e., in processes important in tissue development but also in tumor and metastasis formation. Among other things, NG2 leads to an inhibition of neurite growth, and probably plays an important role in amoeboid type of cell invasion. These processes are in many respects similar. Both in inhibition of neurite growth and in mesenchymal-amoeboid transition occur morphological changes which lead to a loss of cell protrusions and a transition to a rounded shape. In both of these processes Rho/ROCK signaling also plays a crucial role. Connection between NG2 and the Rho/ROCK signaling pathway has been indicated in the process of inhibition of neurite growth. The mechanism of Rho/ROCK signaling regulation by NG2 glycoprotein is, however, still unknown. In this thesis is proposed a molecular mechanism of Rho/ROCK pathway activation by glycoprotein NG2 which relies on the NG2/MUPP1/Syx signaling complex where the scaffold protein MUPP1, bound to activated NG2, enables binding and activation of the Syx protein. Syx then as RhoGEF activates Rho/ROCK signaling, and the activated Rho/ROCK pathway leads to inhibition of neurite growth, increased cell contractility and traction forces. These processes are...
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In-vitro analysis of amoeboid-mesenchymal transition of A375m2 melanoma cells
Kasalová, Lenka ; Rösel, Daniel (advisor) ; Vomastek, Tomáš (referee)
The invasion of cancer cells is an important aspect of cancer progression. Single tumor cells exhibit at least two types of invasion in 3D environment, mesenchymal and amoeboid invasion. Tumor cells can switch between these two modes of movement depending on cellular status and surrounding environment. Amoeboid-mesenchymal transition (AMT) is less explored then mesenchymal-amoeboid transition (MAT). We performed a proteomic analysis of amoeboid-mesenchymal transition of human melanoma cell line A375M2. We have induced amoeboid-mesenchymal transition by treatment with a ROCK inhibitor Y27632 in 3D matrigel matrices and in 2D environment. Induction of the amoeboid-mesenchymal transition has changed a level of expression of 92 proteins and a level of phosphorylation of 15 proteins. Expression of only 17 proteins and phosphorylation of 8 proteins was identically changed in both of these environments. We found that PKCα regulates amoeboid migration and that treatment of cells with a PKCα inhibitor Gö6976 induces amoeboid-mesenchymal transition. Analysis of the proteomics data have further shown that induction of AMT by the ROCK inhibitor Y27632 leads to activation of antiapoptotic signals and activation of signaling pathways involved in regulation of actin cytoskeleton especially regulation of focal...
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The role of dipeptidyl peptidase-IV and homologous proteases in migration and invasion.
Fejfarová, Edita ; Rösel, Daniel (referee) ; Bušek, Petr (advisor)
Migration and invasion are processes which naturally occur in organism during embryogenesis, immune reactions or wound healing. These processes are very important factors in some serious diseases like rheumatoid arthritis and carcinogenesis. There is no doubt about contribution of proteases in these processes-many of them degrade extracellular matrix and thereby facilitate the movement of cells. While dipeptidylpeptidase-IV cleaves solely two amino acids from N-terminus so it is not considerably involved in ECM degradation. DPPIV and its homologues recognize peptides with proline on penultimate position, which causes resistance to ordinary types of proteases. Substrates of DPPIV and its activity homologues include chemokines implicated in signalling of migration - their cleavage and thus inactivation present DPPIV and DPPIV-like molecules as modulators of cell migration signalling in choriocacinoma, neuroblastoma, on Sézary cells or epithelial cells migrating in response to injury. Another activity of some DPPIV-like proteases is binding to the extracellular matrix proteins, when are helping in the attachment of cells and thus affect the migratory ability of the cells like ovarian cancer cells, prostate cancer cells, melanoma cells or kidney cells. Effects on migration and invasion have also...
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Crosstalk of integrin and mTOR signaling
Teglová, Lucie ; Libus, Jiří (referee) ; Rösel, Daniel (advisor)
iv Abstract Crosstalk of integrin and mTOR signalling is an essential process that monitors cellular interaction with extracellular matrix and transmits these inputs to cell growth signalling. Although adhesion status of the cell monitored by integrin signalling is clearly important for regulation of cellular growth, a little is known about the crosstalk of integrin and mTOR signalling. In this study, we employed two different approaches to describe and elucidate character of this crosstalk. p130Cas is an adaptor protein phosphorylated by Src kinase and focal adhesion kinase upon integrin ligand binding and implicated in cell adhesion, motility and survival in both Src-transformed and untransformed cells. Recently, p130Cas was also described in cellular pathology, mainly by its ability to stimulate cell invasion and metastasis. In this study, we described that p130Cas affects mTOR signalling in Src-transformed cells. Substrate domain of p130Cas was found to be indispensable for this effect and is also responsible for serum-induced activation of mTOR signalling. In addition, we prepared cell lines overexpressing various Rheb protein versions and characterized them in context of mTOR signalling, integrin signalling and cell cycle progression. Interestingly, a cell line overexpressing constitutively active...
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Molecular mechanisms of signal transduction by the ERK signaling cascade.
Bráborec, Vojtěch ; Rösel, Daniel (referee) ; Vomastek, Tomáš (advisor)
The MAPK (mitogen-activated protein kinase) cascade represents an evolutionary conserved mechanism by which cells sense extracellular signals and convert them into variety of context-dependent responses. The best studied member of the MAPK protein family is protein kinase ERK (extracellular signal-regulated kinase). Together with protein kinases Raf and MEK (MAPK/ERK kinase) comprise a prototypical signaling pathway which regulates broad-spectrum of biological processes such as cellular proliferation, differentiation, cellular migration, adhesion or apoptosis. To modulate such a multitude of distinct responses by a single pathway, cells utilize mechanisms such as signal strength and duration, distinct protein localization, communication with other signaling pathways, differential substrate selection and the selection of interactive partners. All presented means of regulation are influenced by proteins with non-enzymatic functions - scaffold proteins, protein inhibitors and anchoring proteins. These protein modulators channel the signals leading to particular cellular response, and thus represent the key element of signal transduction. Despite increasing importance of protein modulators in cellular signaling, their biological roles remain mostly unknown. The physiological importance of protein modulators is...
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