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Interaction of protein subunits SEC10 and SEC15 of the exocyst tethering complex
Bartáková, Anna ; Ryšlavá, Helena (advisor) ; Čermáková, Michaela (referee)
The exocyst is an evolutionarily conserved tethering complex involved in the regulation of the secretory pathway in eukaryotic cells. As an effector of Small GTPases, the exocyst contributes to efficient targeting of secretory vesicles to the sites of intense exocytosis at the plasmatic membrane through interactions with specific membrane phospholipids. The exocyst complex consists of eight subunits: SEC3, SEC5, SEC6, SEC8, SEC10, SEC15, EXO70, and EXO84. Each subunit binds at least two other subunits, among them the interaction of SEC10 and SEC15 is crucial for the exocyst function. The question remains how this particular interaction is evolutionarily conserved across different plant taxonomic groups. SEC10 and SEC15 genes from the three evolutionary groups of plants (green algae - Klebsormidium nitens, mosses - Marchantia polymorpha, angiosperms - Arabidopsis thaliana) were cloned into yeast two-hybrid vectors for studying protein interactions. Testing the interaction of SEC10 and SEC15 exocyst subunits from different plant species in the yeast-two hybrid system showed that despite partially different protein sequences of the tested subunits, their interaction across different evolutionary groups of plants remains very well conserved.
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Drawings as a tool for investigating lower-secondary students' knowledge of cell structure
VLČKOVÁ, Barbora
This bachelor's thesis focuses on the use of drawing as a tool to assess students' knowledge of cell structure. The aim of the study is to examine how drawing can serve as a means of gathering information about students' knowledge and to evaluate the level of understanding of cell biology among 6th-grade students. The research involved 41 participants from two elementary schools. A questionnaire with three parts was used to collect data from the participants. The first part of the questionnaire inquired about basic personal information and the participants' interest in drawing. The second part consisted of open-ended and closed-ended questions to assess knowledge related to cells. In the final part, the participants were given space to draw and describe organelles of animal and plant cells. The questionnaire was administered to the participants twice. In the first administration, they were asked to demonstrate their knowledge without any preparation. During the second visit to the schools, the participants were provided with an educational text about cells, accompanied by illustrations of individual cells, to quickly familiarize them with basic information about cells. They then completed the same quiz again. The results indicate that drawing can be an effective tool for assessing students' knowledge of cell structure. Through drawing, students express and visualize their understanding of cells. The majority of respondents believe that drawing significantly contributes to their learning. The first quiz yielded average results, with at least half of the respondents answering correctly to most questions. Improvement in results was observed in the second quiz. The evaluation of the third part of the questionnaire was based on predetermined criteria. With one exception, each respondent included at least the nucleus in their cell drawing. Most respondents depicted more than one organelle in the cell.
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Cell biology of iron transport in plants.
Batík, Adam ; Žárský, Viktor (advisor) ; Tylová, Edita (referee)
Plants use iron as a cofactor of proteins used in photosynthetic systems, electron transport chain and many more. Iron bioavailability for plants in soil is low because it tends to oxidise and create insoluable compounds.For this reasonplants haveevolvedtwo distinct iron uptake mechanisms.Because of the iron toxicitycaused by production ofreactiveoxygen species via the Fenton reaction and the unspecific transport of metals other than iron, plants have to regulate cellular iron concentrationtightly.Theyhave evolved a complex system of signalling networks that has recently begun to uncover. In additionto the regulation ofiron uptake, the plant cell combats iron toxicity by sequestering iron into storage organelles and by chelating it. Iron is essential for seed sprouting but this work is focused on transport of iron into the plant from the soil,subcellulartransportandlongdistance transport ofironin the vasculature.
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The role of ARPC2 in plant cells
Šlajcherová, Kateřina ; Schwarzerová, Kateřina (advisor) ; Klíma, Petr (referee)
ARPC2 protein localization in a plant cell Kateřina Šlajcherová 1 Abstract Actin cytoskeleton is an ubiquitous structure which plays numerous irreplacable roles. Actin nucleation is, beside formins, performed by ARP2/3 complex (actin-related protein), comprising of seven subunits (ARP2, 3, C1-C5) and activated by protein SCAR/WAVE complex. ARP2/3 complex is attached to the membrane and branches existing microfilaments, apart from nucleating them de novo. ARP2/3 mutants in most organisms show severe defects. However, plant mutants exhibit only mild phenotype, for example, Arabidopsis thaliana ARPC2 mutant (dis2-1) has deformed trichomes and leaf epidermal cells, but its viability is not impaired. The aim of the thesis is to map ARPC2 localization within the cell and broaden our understanding of ARP2/3 complex role in plant cell morphogenesis. Tobacco ARPC2 (NtArpC2) subunit was visualized in Arabidopsis plants, using the GFP fusion protein as well as imunofluorescence and anti-ARPC2 antibody. Experiments were undertaken to collocalize the subunit with actin and microtubular cytoskeleton, with mitochondrions, endosomes and other membrane organelles. The specimens were observed in confocal and TIRF microscope. The GFP-NtARPC2 protein shows as motile dots; their movement, but not their existence, is dependent...
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The role of ARP2/3 complex in plant cells
Schiebertová, Petra ; Schwarzerová, Kateřina (advisor) ; Martinec, Jan (referee)
2 Abstract ARP2/3 protein complex is formed from seven proteins (ARP2, ARP3 and ARC1- ARPC5) with a relatively conserved structure. ARP2/3 complex branches and nucleates new actin filaments. This thesis focuses on the study of the role and importance of the individual subunits of the complex ARP2/3 in plants. One of the principal aims of this work is to determine whether complex ARP2/3 may at least partially maintain its role when one or more of the subunits are not available. Furthermore if the individual subunits play another, plant-specific role and if the subunits are functionally equivalent in the complex. The main way how to achieve this objective is the analysis of multiple mutants of Arabidopsis thaliana in subunits of ARP2/3 complex. After comparing several phenotypes of mutant lines it is obvious that all the subunits are functionally equivalent. A loss of ARPC5 subunit usually manifests the strongest phenotypic expression. On the contrary, loss ARPC3 and ARPC2b subunits have weak phenotypic manifestations. Because some phenotypes, such as phenotype distorted trichomes was detected only in some mutant lines, whereas the phenotype of faster roots gravitropic response or vacuolar system fragmentation that was detected in all analyzed mutants suggests, that different subunits play varying roles...
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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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Analysis of endomembrane markers in the cortical cytoplasm and their co-localization with Arp2/3 complex
Jelínková, Barbora ; Schwarzerová, Kateřina (advisor) ; Fendrych, Matyáš (referee)
ARP2/3 is an evolutionarily conserved heteroheptameric protein complex. Its main activity lies in the nucleation of dendritic actin filaments that are involved in membrane remodeling. ARP2/3 takes part in plasma membrane remodeling and the formation of cytoplasmic protrusions that serve in the amoeboid motion of mammalian cells and some protists and plays role in exocytosis and endocytosis of animal and yeast cells. The main objective of this work was to find a connection between the ARP2/3 complex and the regulation of the plant endomembrane system. Using TIRF microscopy we visualized the localization of the ARP2/3 complex in the cortical layer of plant cells and compared it to the localization of several endomembrane markers from the Rab family and an exocytotic marker Exo84b. In the vicinity of the plasma membrane, the ARP2/3 complex subunits localized to dynamic dots very similar to the localization of Exo84b protein. Colocalization analysis showed that a small portion of Exo84b marker and ARP2/3 complex signals colocalize and this result was seconded by the biochemical approach of coimmunoprecipitation. Key words: ARP2/3, endomembrane system, cortical layer, RabA1g, RabC1, RaD2a, Exo84b
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Mechanisms of Microtubules Dynamics and Nucleation in a Plant Cell
Mauerová, Zdeňka ; Schwarzerová, Kateřina (advisor) ; Žárský, Viktor (referee)
Nucleation of microtubules co-determines organization of this cytoskeleton component in cells and makes a significant contribution to shaping its dynamics. In plant cells, micro- tubules are mainly nucleated on preexisting ones and nucleation takes place in the cortex and also within the mitotic spindle and the fragmoplast. Recruiting the γTuRC, a pre- served universal nucleator, to the wall of microtubules is provided by augmin in cooperation with NEDD1/GCP-WD. The function of the γTuRC is at least in the case of nucleation in the spindle, but apparently in other situations as well further enhanced by XMAP215/MOR1, which raises efficiency of the γTuRC through its own polymerization activity, and TPX2, or its homologs, which for one thing, directly activates the complex and for another, locally increases concentration of tubulins by forming condensate with them, which also augments the probability of success of nucleation. Not much is known about regulatory pathways controlling this process, with the exception of the TTP complex, which is functional in the cortex. Overall, knowledge covering nucleation in plants is rather meager and information concerning the molecular mechanisms of functioning of mentioned factors comes mainly from research in animals. Keywords augmin, cytoskeleton, γTuRC,...
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Plant tubulin code
Ničová, Klára ; Schwarzerová, Kateřina (advisor) ; Cvrčková, Fatima (referee)
Microtubules, which are made of polymers of α- and β-tubulin, are an integral part of the cytoskeleton. Both types of tubulin share a considerable sequential homology across eukaryotic organisms. Tubulins are encoded by relatively large gene families. The expression of these genes produces different tubulin isotypes, some of which may exhibit different properties. Tubulin isotypes can be further posttranslationally modified. The best known posttranslational modifications of tubulin include acetylation, phosphorylation, tyrosination, polyglutamylation and polyglycylation. The tubulin code arises from the combination of expression of different tubulin isotypes and their posttranslational modifications. As a consequence, microtubules in cells can be composed of a "mixture" of different tubulins with distinct functions and properties. Even though the existence of the tubulin code has been proven in every model organism, plants included, the precise understanding of the meaning of microtubules being composed of different tubulins is still subjected to research. Much of the research on the tubulin code has been carried out on animal models. In contrast, relatively little is known about the existence of the tubulin code in plants. This theses summarizes current knowledge on the localization and regulation...
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