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Vesicular roles of Arp2/3 nucleation-promoting factors
Dostál, Vojtěch ; Libusová, Lenka (advisor) ; Malínský, Jan (referee) ; Befekadu, Asfaw (referee)
F-actin is involved in key aspects of vesicular traffic, such as membrane deformation, tubulation and vesicle motion. Branching of F-actin is mediated by Arp2/3 but this complex must first be activated by so-called nucleation-promoting factors (NPFs). These factors play an essential role in the decision where and when branched actin should form on the membrane surface. The thesis focuses on the mechanisms which underlie localization and activation of NPFs, especially in terms of the phosphoinositide composition of the vesicle membranes. I show that one of the NPFs, the WASH complex, does not exclusively depend on the retromer complex for its membrane anchoring, as previously theorized. Rather, its understudied subunit SWIP enables the complex to independently bind to the membrane. I also present data showing that the WASH complex has essential roles in maintaining lysosomal function. Additionally, I elucidate the function of another NPF known as WHAMM in the ERGIC compartment, showing that it depends on the presence of myotubularin 9 for its ability to form membrane tubules. The thesis improves our understanding of the interface between the actin cytoskeleton and intracellular membrane system.
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The role of CUP-4 protein in Wnt signalling
Žídek, Radim ; Macůrková, Marie (advisor) ; Pospíchalová, Vendula (referee)
Wnt signalling is indispensible for proper development of organisms and maintaining of adult tissue homeostasis. Its disruption often leads to disease. In nematode Caenorhabditis elegans, Wnt signalling governs vast array of developmental processes, among others also migration of the Q neuroblasts and their descendants. The sole Wnt acting in this process, EGL-20, triggers the canonical β-catenin Wnt signal transduction pathway in QL but not in QR which leads to QL remaining in the posterior while the QR migrates anteriorly. This represents a useful tool for studying Wnt signalling. Recently, mutation of gene cup-4 was found to disrupt migration of the QL neuroblast in a small proportion of the mutant population. cup-4 encodes a ligand-gated ion channel family homologue and it was shown to participate in endocytosis by coelomocytes, specialized phagocytic cells in the C. elegans body cavity. Here, I present the results of my effort to determine the place of CUP-4 action in Wnt signalling and to elucidate the mechanism of its function. I found that CUP-4 acts upstream of PRY- 1/Axin, which is involved in signal transduction in signal receiving cells, and most probably downstream of adaptin AP2, which is important for recycling of Wnt cargo receptor Wntless (Wls) in Wnt producing cell. cup-4 also...
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Mechanisms of retromer - dependent protein recycling from endosomes
Horázná, Monika ; Macůrková, Marie (advisor) ; Černý, Jan (referee)
Most processes in nature are very effective concerning saving energy and minimizing waste. A good example of saving on cellular level is receptor recycling. Whether it concerns receptors for lysosomal enzymes or for proteins destined for secretion, after releasing their cargo protein the fate of the receptor would be sealed in lysosomes. Nevertheless, some transmembrane receptors contain a signal motif through which they are recognized by specific proteins or protein complexes and they escape the degradation in lysosomes. One such complex is the retromer. Its first discovered function was the recycling of receptors for lysosomal hydrolases in yeast. Later it was proved that it has a similar role in transport of many other proteins in other eukaryotes. The task for retromer is to sort the cargo proteins on the endosomal membrane and together with others auxiliary proteins create a transport vesicle which is then transported to the Golgi. This makes the cell able to recycle proteins that would otherwise be transported from endosomes to lysosomes for degradation.
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Mechanismy regulace aktivity proteinu MTM-6 na endosomech.
Horázná, Monika ; Macůrková, Marie (advisor) ; Fafílek, Bohumil (referee)
Wnt signalling belongs to conserved pathways and mediates cell fate decision, development, regeneration and adult tissue homeostasis. Disruption or misregulation of Wnt signalling pathway often leads to disease. Wnt proteins are hydrophobic glycoproteins which need a special receptor for transport from Golgi Apparatus to cell surface, which is called MIG-14 in Caenorhabditis elegans and Wntless (Wls) in mammals. In this study, I focus on understanding mechanisms that regulate MTM-6 protein activity. MTM-6, a lipid phosphatase associated with endosomal membrane, has been recently identified as a regulator of MIG-14/Wls trafficking in Caenorhabditis elegans. Silencing of mtm-6 leads to misregulation of some Wnt-directed processes, such as migration of Q neuroblasts progeny. This study reports identification of novel mtm-6 genetic interactors that have been found to influence migration of Q neuroblasts progeny through Wnt signalling. New knowledge about mtm-6 genetic interactions bring us near to understanding of Wnt signalling regulation. Keywords: Caenorhabditis elegans, MTM-6, SEL-5, Wntless, Wnt, endosomes, phosphoinositides, retromer
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The role of CUP-4 protein in Wnt signalling
Žídek, Radim ; Macůrková, Marie (advisor) ; Pospíchalová, Vendula (referee)
Wnt signalling is indispensible for proper development of organisms and maintaining of adult tissue homeostasis. Its disruption often leads to disease. In nematode Caenorhabditis elegans, Wnt signalling governs vast array of developmental processes, among others also migration of the Q neuroblasts and their descendants. The sole Wnt acting in this process, EGL-20, triggers the canonical β-catenin Wnt signal transduction pathway in QL but not in QR which leads to QL remaining in the posterior while the QR migrates anteriorly. This represents a useful tool for studying Wnt signalling. Recently, mutation of gene cup-4 was found to disrupt migration of the QL neuroblast in a small proportion of the mutant population. cup-4 encodes a ligand-gated ion channel family homologue and it was shown to participate in endocytosis by coelomocytes, specialized phagocytic cells in the C. elegans body cavity. Here, I present the results of my effort to determine the place of CUP-4 action in Wnt signalling and to elucidate the mechanism of its function. I found that CUP-4 acts upstream of PRY- 1/Axin, which is involved in signal transduction in signal receiving cells, and most probably downstream of adaptin AP2, which is important for recycling of Wnt cargo receptor Wntless (Wls) in Wnt producing cell. cup-4 also...
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Mechanisms of retromer - dependent protein recycling from endosomes
Horázná, Monika ; Macůrková, Marie (advisor) ; Černý, Jan (referee)
Most processes in nature are very effective concerning saving energy and minimizing waste. A good example of saving on cellular level is receptor recycling. Whether it concerns receptors for lysosomal enzymes or for proteins destined for secretion, after releasing their cargo protein the fate of the receptor would be sealed in lysosomes. Nevertheless, some transmembrane receptors contain a signal motif through which they are recognized by specific proteins or protein complexes and they escape the degradation in lysosomes. One such complex is the retromer. Its first discovered function was the recycling of receptors for lysosomal hydrolases in yeast. Later it was proved that it has a similar role in transport of many other proteins in other eukaryotes. The task for retromer is to sort the cargo proteins on the endosomal membrane and together with others auxiliary proteins create a transport vesicle which is then transported to the Golgi. This makes the cell able to recycle proteins that would otherwise be transported from endosomes to lysosomes for degradation.
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