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The role of the interaction of LCK with CD4/CD8 coreceptors
Cesneková, Michaela ; Štěpánek, Ondřej (advisor) ; Černý, Jan (referee)
LCK kinase is an essential regulator of T-cell signalling that interacts with CD4 and CD8 coreceptors, which are crucial for T-cell development and T-cell lineage commitment. Their role, as well as the role of their interaction with LCK in the peripheral T cells, remains disputable, despite being studied for decades. This thesis aims to investigate the importance of LCK-coreceptor interaction in CD8+ T cell signalling and development and to determine the significance of the serine residues in LCK-mediated CD4 endocytosis. We used LCK variants bearing mutations of the coreceptor binding site or its catalytic domain in both mice and cell lines to solve this perplexity. First, the enzymatic activity of LCK variants was evaluated in this thesis. Second, we demonstrate that the function of CD8 is both, LCK-CD8 interaction dependent and independent. Then we examined the late stage of CD8+ T cell development, showing that the absence of the interaction has very mild consequences. It affects only the response of post- selection CD8 single-positive, but not double-positive, thymocytes to sub-optimal antigenic stimulation. Finally, we observed that CD4 with the mutation of all three intracellular serines to alanines shows similar LCK-dependency as wild type CD4. Overall, this study sheds light on the...
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The Role of Lck Kinase in T-cell Antigen Receptor Signaling
Němec, Dušan ; Štěpánek, Ondřej (advisor) ; Rösel, Daniel (referee)
LCK activity is crucial for the triggering of the entire T cell activation process. The primary function of LCK is to convert the signal of TCR:pMHC ligation into the intracellular environment. The outcome of the LCK-triggered pathway is T cell activation, cytokine production, differentiation, and clonal expansion. This thesis provides a summary of recent knowledge about the unique position of LCK in the T cell signaling machinery as well as an overview of molecules and interacting partners that regulate LCK activity. It describes the importance of the LCK-coreceptor association for optimal TCR signaling and physiological thymocyte development and mentions discussed adaptor role of LCK in the T cells. Keywords: LCK, T-cell, antigen, kinase, enzyme
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Identification of a new mechanism of Lck regulation via its C-terminal sequence
Valečka, Jan ; Filipp, Dominik (advisor) ; Vomastek, Tomáš (referee)
T-cell activation is a complex process crucial for a proper function of immune system. It has been extensively studied and its main features are well understood. However, some of the events involved in T-cell signalling are still unclear. After T-cell receptor stimulation, Src-family kinase Lck drives the initiation of signalling by tyrosine phosphorylation. Phosphorylation of several downstream targets is dependent on the redistribution of Lck to the different compartment of the plasma membrane, called lipid rafts. In lipid rafts, active Lck is juxtaposed and activates raft-resident substrates which then trigger downstream signalling. The critical in this process is the mechanism of Lck translocation to lipid rafts which has not been studied so far and represents the topic of great academic and clinical interests. Previously, we identified the adaptor protein RACK1 as a candidate protein mediating the redistribution of Lck to lipid rafts by linking it to the microtubular network. In this thesis, we analysed the structural features and functional role of RACK1 in its interaction with Lck. We show here, using the SYF cell lines expressing the wild type and various mutated forms of Lck, that intact SH3 or SH2 domains of Lck are required for an effective RACK1-Lck complex formation. We also documented...
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