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Functional analysis of plant Arp2/3 complex subunits
Kukla, Jakub ; Schwarzerová, Kateřina (advisor) ; Honys, David (referee)
1. Abstract ARP2/3 complex is well studied in case of animals, it plays key roles in motility of cells and intracellular organels. It's malfunctions result in severe growth disorders and even lethality of affected cells. On the contrary, plant cells do not exhibit such dramatic phenotype of ARP2/3 complex mutations like it is by animals. It is possible that just the different life strategies of plants and animals contribute to differences in a way how animal and plant cells use their cytoskeleton, where ARP2/3 complex is it's part as well. It is highly conserved 7 protein complex from yeast to human. His main functions are creation of new "de novo" actin filaments, actin branched filaments network. Some of the parasite organisms are capable of missusing its nucleator activity to actively move inside of host cell. Because of the plant cells are sourounded by the cell wall, which give them support in creating various shapes and also hinders active movement of the whole cell body, it is likely that ARP 2/3 complex could be possibly involved in novel plant specific functions as well. If we think about the different life strategy of plants and animals we can not ignore all the things these two kingdoms have in common regarding to cytoskeleton processes. That is the need both for vesicular transport and...
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Non-traditional roles of formins besides actin nucleation
Metlička, Jáchym ; Cvrčková, Fatima (advisor) ; Opatrný, Zdeněk (referee)
Formin homology 2 (FH2) domaincontaining proteins (formins) have, since their discovery in 1990, been observed in all analyzed species of eukaryotic kingdoms. Our knowledge of structure and function of the defining FH2 domain has greatly increased over the last couple of years. Its function in nucleation, polymerization and processive capping of actin filaments designates formin protein family an important cytoskeletonremodelling factor. But FH2 domain is just one part of the puzzle additional optional conserved peptide structures surrounding it, as well as concrete variation of the FH2 domain itself, greatly influence the functional properties and cellular localization of the resultant formin protein. Formins have been implicated in variety of cellular processes, which often (but not always) involve the cytoskeleton e.g. Factin network management, crosstalk of Factin filaments and microtubules or plasma membrane. They also partake in processes integral to cell division, function in conserved signalling pathways and much more. This thesis explains the structure and function of FH2 and FH1 domains, outlines the main formin phylogenetic clades in multicellular eukaryotes and reviews various roles that formins fulfill or are thought to fulfill. Such goal, however, is very bold and (considering the...
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Microtubule-associated proteins in plants
Benáková, Martina ; Krtková, Jana (advisor) ; Vinopal, Stanislav (referee)
1. Abstract and key words MTs are one of the basic cellular protein structure. Their features and function are influenced and modified by group of other proteins, i.e. microtubule-associated proteins (MAPs). In the last decades, an extensive research on MAPs and their wide range of functions has been carried out. Therefore we are aware of the involvement of some of the MAPs in MT dynamics, other have been shown to have rather structural function. They bundle MTs with various cell structures, such as the other MTs, proteins, organelles, actin cytoskeleton or plasma membrane. Many described MAPs are homologous in the whole eukaryotic domain, for example MAP65 or EB1 (END BINDING 1) family, therefore it is interesting to follow if and how the functions of plant MAPs differ from their animal counterparts. On the other hand, there are many specific MAPs with unique functions in plants, e.g. ATK5 or SPR1 (SPIRAL 1). This Bachelor thesis is a survey on current knowledge of plant MAPs and it makes an effort to present their characteristic and functions in plant cell and organism. Key words: cytoskeleton, microtubules, microtubule-associated proteins, plant cell, growth and development
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Mechanisms of epidermal cells polarization in plants.
Vojtíková, Zdeňka ; Žárský, Viktor (advisor) ; Soukup, Aleš (referee)
Plant epidermal cells form contact area of the plant, they protect it from impacts of surrounding environment and they mediate the communication with its neighbourhood. In the epidermis there are evenly distributed several cell types with quite specialized morphology (pavement cells, trichomes and guard cells) due to the polarization mechanisms. The cytoskeleton and signal molecules of ROP GTPase family promote the polarized growth. Thanks to polarized growth the cells reach their shapes. The cytoskeleton responds to the signal by expanding the cell, helps with targeting of the secretion to the sites of active growth and mediates polarized formation of the cell wall. On the upper side of the epidermis the cuticle and layer of epicuticular waxex is secreted. The secretion of cuticular components is baso-apicaly polarized. This work summarizes the mechanisms of the polarization in plant epidermal cells discoverd untill now.
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Characterization of membrane protein DREPP
Vosolsobě, Stanislav ; Schwarzerová, Kateřina (advisor) ; Cvrčková, Fatima (referee)
Proteins of DREPP family (20-25 kDa, syn. PCaP1 in Arabidopsis thaliana) first appeared in ferns and we have shown that several independent duplications of DREPP protein occurred during evolution of large families (Poaceae, Brassicaceae, Solanaceae and Asteraceae) and in group Coniferophyta. Secondary losses of one paralogue occurred in subfamilies Pooideae and Solanoideae.We have also detected two large-scale modification of DREEP protein in Asparagales and Brassicaceae (this divergent paralogue was previously described as MAP18 protein). We have examined colinearity of chromosome fragments in vicinity both PCaP1 and MAP18 paralogues in Arabidopsis thaliana and we hypothesize that MAP18 gene arose during genome duplication on the origin of Brassicaceae family. DREPP protein was previously identified in detergent-resistant membrane microdomain fraction and a myristyl anchor was shown to be necessary for their membrane localization. Membrane association was shown to be modified by the interaction of unique N-terminal domain with PtdInsPs, which was inhibited by binding of Ca-calmodulin (Nagasaki et al., 2008). The mutation of Gly2 by Ala in the myristilation site, or C-terminal GFP-fusion (GFP-DREPP), affect membrane association in Arabidopsis thaliana (Nagasaki et al., 2008). Several DREPP paralogues in...
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Computational Simulation of Mechanical Tests of Isolated Animal Cells
Bansod, Yogesh Deepak ; Kučera,, Ondřej (referee) ; Florian, Zdeněk (referee) ; Canadas, Patrick (referee) ; Burša, Jiří (advisor)
Buňka tvoří složitý biologický systém vystavený mnoha mimobuněčným mechanickým podnětům. Hlubší pochopení jejího mechanického chování je důležité pro charakterizaci její odezvy v podmínkách zdraví i nemoci. Výpočtové modelování může rozšířit pochopení mechaniky buňky, která může přispívat k vytvoření vztahů mezi strukturou a funkcí různých typů buněk v různých stavech. Za tímto účelem byly pomocí metody konečných prvků (MKP) vytvořeny dva bendotensegritní modely buňky v různých stavech: model vznášející se buňky pro analýzu její globální mechanické odezvy, jako je protažení nebo stlačení, a model buňky přilnuté k podložce, který vysvětluje odezvu buňky na lokální mechanické zatížení, jako třeba vtlačování hrotu při mikroskopii atomárních sil (AFM). Oba zachovávají základní principy tensegritních struktur jako je jejich předpětí a vzájemné ovlivnění mezi komponentami, ale prvky se mohou nezávisle pohybovat. Zahrnutí nedávno navržené bendotensegritní koncepce umožňuje těmto modelům brát v úvahu jak tahové, tak i ohybové namáhání mikrotubulů (MTs) a také zahrnout vlnitost intermediálních filament (IFs). Modely předpokládají, že jednotlivé složky cytoskeletu mohou měnit svůj tvar a uspořádání, aniž by při jejich odstranění došlo ke kolapsu celé buněčné struktury, a tak umožňují hodnotit mechanický příspěvek jednotlivých složek cytoskeletu k mechanice buňky. Model vznášející se buňky napodobuje realisticky odezvu síla-deformace během protahování a stlačování buňky a obě odezvy ilustrují nelineární nárůst tuhosti s růstem mechanického zatížení. Výsledky simulací ukazují, že aktinová filamenta i mikrotubuly hrají klíčovou úlohu při určování tahové odezvy buňky, zatímco k její tlakové odezvě přispívají podstatně jen aktinová filamenta. Model buňky přilnuté k podložce dává odezvu síla-hloubka vtlačení ve dvou různých místech odpovídající nelineární odezvě zjištěné experimentálně při AFM. Výsledky simulací ukazují, že pro chování buňky je rozhodující místo vtlačení a její tuhost určují aktinová povrchová vrstva, mikrotubuly a cytoplazma. Navržené modely umožňují cenný vhled do vzájemných souvislostí mechanických vlastností buněk, do mechanické úlohy komponent cytoskeletu jak individuálně, tak i ve vzájemné synergii a do deformace jádra buňky za různých podmínek mechanického zatížení. Tudíž tato práce přispívá k lepšímu pochopení mechaniky cytoskeletu zodpovědné za chování buňky, což naopak může napomáhat ve zkoumání různých patologických podmínek jako je rakovina a cévní choroby.
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Application of tensegrity structures in modelling of mechanical behaviour of smooth muscle cells
Bauer, David ; Fuis, Vladimír (referee) ; Burša, Jiří (advisor)
The master’s thesis deals with the computational modelling of the mechanical testing of isolated smooth muscle cells. The main aims are to create computational model of a cell, to simulate single-axis tensile test and to modify the model so that the model reflects real mechanical response. The model of the cell includes cytoplasm, nucleus, cell membrane and cytoskeleton which is modelled as a tensegrite structure. On this model the tensile test was simulated in case of the cell with cytoskeleton and the cell with distributed the cytoskeleton. Force-elongation curves, which were obtained from this simulation, were compared with experimental data which were taken from literature. Tensile properties were measured on freshly isolated cells from rat thoracic aorta, cultured cells, and cells treated with cytochalasin D to disrupt their actin filaments. It was found that the cytoskeleton influence on the cell load in computational model was smaller than in the real cell. Therefore the model was modified by changing material propreties and geometry so that the model of the cell corresponded with the different types of experimentally measured cells.
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