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Phosphorylation of the adaptor protein PSTPIP2 and its role in neutrophilic granulocytes
Dvořáček, Tomáš ; Brdička, Tomáš (advisor) ; Ballek, Ondřej (referee)
PSTPIP2 is an adaptor protein of the F-BAR family, which is an important regulator controlling the effector mechanisms of innate immune cells. The regulatory functions of this protein were discovered thanks to the CMO mouse strain, which lost the expression of this protein. As a result of PSTPIP2 deficiency, mice of the CMO strain develop an autoinflammatory disease affecting bone tissue and skin. The main mechanism that drives its pathology is the loss of regulation of the neutrophil granulocyte activity. These cells then produce excessive amounts of the pro-inflammatory cytokine IL-1β and reactive oxygen species. However, the exact molecular mechanism of action of the PSTPIP2 protein is unknown. When the PSTPIP2 protein is activated, it is phosphorylated and interacts with other proteins, which mediate its regulatory function. Interaction partners described so far in neutrophil granulocytes include phosphatases of the PEST family, the lipid phosphatase SHIP1 and the non-receptor tyrosine kinase CSK. In this thesis, we identified kinases from the SRC family as kinases that phosphorylate PSTPIP2. Furthermore, we found that the main phosphorylation sites of PSTPIP2 are tyrosines at positions 323 and 329. Finally, we proved that SHIP1 can bind to the phosphotyrosine motif around the tyrosine at...
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The role of protein tyrosine phosphatase CD45 in neutrophil granulocytes
Ilievová, Kristýna ; Brdička, Tomáš (advisor) ; Černý, Jan (referee)
Strict regulation of the immune response is critical for appropriate protection against in- fection, preventing tissue damage, and maintaining homeostasis. A significant part of this regulation is mediated at the level of signaling pathways in which tyrosine phosphorylati- on plays a key role. It is regulated by the action of protein tyrosine kinases and protein tyrosine phosphatases (PTP). An important PTP expressed on all nucleated hematopoie- tic cells is the CD45. Its role has been studied primarily in T- and B-lymphocytes. There CD45 plays an important role in antigen-induced signaling and signaling triggered by other stimuli. It becomes apparent that also in neutrophils CD45 plays an importat role in many mechanisms that contribute to appropriate protection against infection. These include, for example, adhesion, extravasation, chemotaxis, phagocytosis, production of cytokines and oxidative burst. In many cases, CD45 affects these processes by regulating Src family kinases. Other means of CD45 participation in specific pathways are often not clear. This thesis summarizes our current understanding of role of CD45 in neutrophil granulocytes and its effects on the function of these cells. 1
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The role of Src-family kinases in the immunological synapse of antigen presenting cells.
Kotlabová, Klára ; Brdička, Tomáš (advisor) ; Brábek, Jan (referee)
Antigen presentation during which antigen fragments in complex with MHC glycoproteins are recognized by T cell antigen-specific receptors is necessary for the initiation of adaptive immune response. During this process, immunological synapse is assembled at the site of contact between the T cell and the antigen-presenting cell (APC). This leads to the activation of receptors on the surface of both cells followed by triggering of multiple signaling pathways. However, our knowledge about the signaling occurring at the APC-side of the IS is limited in comparison to the T cell side. Here, we analyze role of Src family kinases in the APC signaling pathways. For this purpose, constructs targeting Csk kinase to the plasma membrane of APCs were prepared to inhibit SFKs there. We show that expression of these constructs inhibits activation of SFKs, calcium mobilization and cell activation of K46 B cell line. Further, expression of these constructs in hematopoietic progenitors attenuates their differentiation into dendritic cells which then results in their decreased ability to stimulate T cells.
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The role of Src family kinases in RNA processing
Gemperle, Jakub ; Rösel, Daniel (advisor) ; Mácha, Jaroslav (referee)
Until now, a lot of information have been obtained about the role of Src family kinases in the cytoplasm or at the plasma membrane and their interactions with growth factor receptors or focal adhesion complexes. Their functional importance at the perinuclear membrane, or even inside the nucleus, however, has not been well characterized. This work, using available information, pointed at the fact that Src family kinases can be found in the nucleus. This opens a new field of Src kinases action, such as in RNA metabolism, considering that it has been assumed that their activity is limited to the cytoplasmic compartment. This work summarizes the current knowledge that hints to Src family kinases dependent network of regulation of RNA metabolism; Src family kinases have pleiotropic effects not only on the RNA binding proteins, but also on the remodeling of chromatin structure. These kinases affect by direct interactions with other proteins transport, splicing or RNA stability and gene expression. This summary suggests that Src family kinases could regulate RNA metabolism on many levels.
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