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Analysis of intermediate filament structure by chemical cross-linking
Dlabolová, Lada ; Novák, Petr (advisor) ; Šulc, Miroslav (referee)
Intermediate filament proteins create a dynamic cytoskeletal filamentous network, which due to its elastic properties, significantly contributes to the resistance of cells and tissues to mechanical stress. An important protein from the family of intermediate filaments, vimentin, is expressed mainly in cells of mesenchymal origin. Vimentin has been associated with a large number of pathophysiological conditions, and current studies consider vimentin as clinically promising target for the diagnosis, prognosis and treatment of a wide range of diseases from cancer to infectious and inflammatory diseases. Although in terms of structural characterization, vimentin belongs to one of the most studied proteins from the family of intermediate filaments, our knowledge is currently limited to the structure of the vimentin tetramer. Vimentin is capable of self-assembly into filaments formed by homo-oligomeric ULF subunits and the assembly process involves several steps of the organization of subunits. Structural characterization of the oligomeric subunits involved in the assembly of vimentin filaments is a prerequisite for elucidating the architecture of mature filaments, which can significantly contribute to understanding and connecting the mechanisms of many diseases associated with changes in vimentin...
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Intermediate filaments in mammalian cell motility
Čermáková, Kateřina ; Libusová, Lenka (advisor) ; Pelantová, Markéta (referee)
Cell migration is crucial for the formation and maintenance of a multicellular mammalian organism, contributing to important processes such as embryonic development, tissue renewal, and immune surveillance. It is a complex phenomenon involving a plethora of processes, including relevant signalization. An impairment of those processes could be projected into innumerable pathological states, from wound healing disruption to tumour metastasis and invasiveness, thus the interest of many researchers has turned toward migration. The migration of mammalian cells is dependent on a cytoskeleton, which is being considerably rearranged in motile cells. This thesis aims to summarise the role of intermediated filaments in cell motility - the less understood cytoskeletal network in this context. Cytoplasmic, as well as nuclear intermediate filaments, due to their unique mechanical properties, affect cell mechanics. They protect against physical stresses (as cells squeeze through confined pores in a complex intercellular microenvironment) and modulate and direct actomyosin-generated forces, which are the main driving force of migration. In addition, they contribute to important migration- involved steps, such as cell polarisation, cell adhesion to surrounding surfaces, cohesion in collective migration, and...
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Cytoskeletal orchestration of early mammalian development
Novotný, Jan ; Libusová, Lenka (advisor) ; Mašek, Jan (referee)
Structures of the cytoskeleton play a critical role in early mammalian development. Microtubules, intermediate filaments and actin microfilaments provide structural and mechanical basis for major developmental transitions of preimplantation development. Microvilli supported by bundles of actin filaments become restricted to one pole of the blastomere during the process of polarisation. All components of the cytoskeleton network undergo apparent reorganisation in embryo compaction. Cells flatten and adhere to each other thanks to the formation of cytoskeleton-linked intercellular junctions and oscillatory contractions generated by actomyosin complexes. Filopodia, actin based protrusions, are debated for their potential role in blastomere apposition and cell signalling. The first process of cell line specification in mammalian development is that which defines outer cells of morula as presumptive trophectoderm and inner cells as presumptive inner cell mass. Proteins and structures of cytoskeleton are asymmetrically distributed between these two populations helping to determine the fate of each blastomere. The aim of the thesis is to summarise the roles of cytoskeleton in blastomere polarisation, embryo compaction and specification of inner and outer cell populations in mammalian preimplantation...
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The role of desmin in a heart
Šeovićová, Maja ; Horníková, Daniela (advisor) ; Vegrichtová, Markéta (referee)
Desmin forms a muscle specific intermediate filament which participates in a formation of a dynamic intracellular network that links contractile apparatus with a sarcolemma, mitochondria and it provides a communication with a cell nucleus. This network serves to maintain morphological and functional aspects of muscle cells and cell organelles during a muscle contraction. Mutation in a desmin or an absence of desmin is causing a serious disease called desminopathy, which belongs among a group of myofibrillar myopathies. This disease manifests itself in all muscle types, however first observable defects occur in cardiac mitochondria, thus heart is a first organ manifesting symptoms of this disease. The purpose of this thesis was to summarize a current knowledge about physiological and molecular mechanisms that involved in a formation of intracellular desmin network and about patho-physiological states of desminopathies. Key words: desmin, intermediate filaments, heart, desminopathy, cardiomyopathy
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Utilization of chemical cross-linking for studying intermediate filaments organization
Dlabolová, Lada ; Novák, Petr (advisor) ; Sabó, Ján (referee)
Intermediate filaments are cytoskeleton components formed by a large family of fibrous proteins specifically expressed in nearly all differentiated cells. Under physiological conditions, they spontaneously assemble into fibers in a process that involves several stages in the organization of subunits. These fibers provide elastic properties to the cells, allowing them to maintain their structural and mechanical integrity. While the structure of other cytoskeletal components is now well researched, detailed information on the structure of intermediate filaments at various stages of assembly is still not available. Thus, new insights into the structure of these proteins could be of great benefit in understanding of various pathological mechanisms associated with changes in their expression in cells. This thesis studies interactions of dimeric subunits in the tetrameric assembly of vimentin, class III protein of intermediate filaments. By chemical cross-linking of isotopically labeled and unlabeled tetrameric vimentin mixture, followed by proteolytic cleavage and mass spectrometry analysis, interdimeric, intradimeric and intrapeptide cross-linking products were identified. Quantification yielded information on interdimeric and intradimeric distance constraints, which allow the characterization of a...
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Protein composition of the cytoskeleton of protists
Švagr, Eva ; Hampl, Vladimír (advisor) ; Pánek, Tomáš (referee)
While we have a good understanding of the roles of actin and tubulin filaments in the cell cytoskeleton, intermediate filaments (IFs) are often overlooked. However, the importance of IFs becomes quite apparent, as proteins similar to IF proteins have been identified in many protist cells. This holds particularly for the cells of protists, where even the functions of some members of the of actin and tubulin superfamilies remain unclear. Intermediate filaments are still not well established as components of protist cytoskeletons, in contrast to their more thoroughly studied counterparts in Metazoa. Protist and metazoan IF proteins are dissimilar in their sequence; however, they share similarities in structure, and they assemble autonomously into analogous filaments. IF-like proteins have been localized to striated fibers or unique cytoskeletal components in several organisms, even though not much is known about the protein composition of these striated fibers to date. This suggests that IF-like proteins might be a universally present component of these striated fibers often seen in protist cells. Keywords: Protists, cytoskeleton, microtubules, microfilaments, intermediate filaments, fibrils, protein composition
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The role of desmin in a heart
Šeovićová, Maja ; Horníková, Daniela (advisor) ; Vegrichtová, Markéta (referee)
Desmin forms a muscle specific intermediate filament which participates in a formation of a dynamic intracellular network that links contractile apparatus with a sarcolemma, mitochondria and it provides a communication with a cell nucleus. This network serves to maintain morphological and functional aspects of muscle cells and cell organelles during a muscle contraction. Mutation in a desmin or an absence of desmin is causing a serious disease called desminopathy, which belongs among a group of myofibrillar myopathies. This disease manifests itself in all muscle types, however first observable defects occur in cardiac mitochondria, thus heart is a first organ manifesting symptoms of this disease. The purpose of this thesis was to summarize a current knowledge about physiological and molecular mechanisms that involved in a formation of intracellular desmin network and about patho-physiological states of desminopathies. Key words: desmin, intermediate filaments, heart, desminopathy, cardiomyopathy
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