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Regulatory mechanism of the cyclin-dependent kinase 16 through the cyclin Y/14-3-3 complex
Janáčková, Zuzana ; Obšil, Tomáš (advisor) ; Pavlíček, Jiří (referee)
CDK16 is a cell-cycle-related cyclin-dependent kinase that functions primarily in neurite growth processes, axonal transport regulation and sperm development. It also plays a role in the progression of X-linked intellectual disability and, together with its activating partner cyclin Y, in the development and progression of various types of cancer. Consequently, CDK16 and cyclin Y could represent potential new therapeutic targets. Both CDK16 and cyclin Y are structurally atypical among similar proteins, and the mechanism of activation of CDK16 by cyclin Y is unclear. However, it is clear from recent studies that the interaction between CDK16 and cyclin Y requires the prior interaction of cyclin Y with 14-3-3 proteins. In addition, for cyclin Y to interact with 14-3-3 proteins, its phosphorylation on the S100 and S326 binding sites of 14-3- 3 is required. To investigate the mechanism of activation of CDK16 by cyclin Y, we decided to characterize the interaction between CDK16, cyclin Y and 14-3-3 proteins in detail. The interaction between CDK16, cyclin Y and 14-3-3 proteins was characterized using biophysical methods including fluorescence anisotropy measurements, native electrophoresis, size exclusion chromatography and protein crystallography. The for- mation of binary pCCNY-14-3-3 and ternary...
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Trk1 Potassium Importers - key transport systems for yeast cell fitness and stress tolerance
Masaryk, Jakub ; Sychrová, Hana (advisor) ; Heidingsfeld, Olga (referee) ; Malínský, Jan (referee)
One of the key prerequisites for yeast cell growth is the uptake of essential compounds, such as potassium. Potassium is a vital monovalent cation and its sufficient intracellular concentration is crucial for various processes, for instance: regulation of membrane potential and cell turgor, enzymatic activity, and protein synthesis. A sufficient internal concentration of potassium is also one of the pivotal signals for cell division. However, as also excess of potassium might lead to unfavourable physiological consequences in yeast, such as deacidification of vacuoles and depolarization of plasma membrane, it is imperative for the yeast cells that the whole process of potassium acquisition is a tightly regulated affair, in order to maintain proper potassium homeostasis. In yeast Saccharomyces cerevisiae, uniporter Trk1 is considered a key player in potassium uptake. The presented thesis aimed to provide novel knowledge regarding Trk1, more specifically to study its ability to modify its capacity for potassium uptake, putative regulation by phosphorylation, and involvement in the survival of glucose-induced cell death (GICD). Additionally, potassium-uptake systems in selected non-conventional species were characterized as well. The most distinctive feature of Trk1 is its alleged ability to switch...
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