<> "The repository administrator has not yet configured an RDF license."^^ . <> . . "Spatio-temporal protease activity and cell death induced by\r\nNatural Killer cells and soluble CD95L"^^ . "In the CD95 signaling cascade molecular processes take place, which rely on protein multimerization\r\nfor activity. In this work, I aimed at measuring the initial events in CD95-mediated apoptosis,\r\nin particular the kinetics of the molecular events ligand binding, receptor oligomerization, FADD\r\nrecruitment and caspase-8 activation to understand which of those processes involve critical kinetic\r\nsteps. For this, I used the model cell HeLa.\r\nI have succeeded in showing and comparing the kinetics of ligand-binding, FADD recruitment and\r\ncaspase-8 activation by using confocal microscopy. I have found that a critical step before caspase-\r\n8 activation is the binding of FADD to the activated receptor. Furthermore, I found that the\r\nactivation of the receptor, presumably by its oligomerization, is a limiting step in CD95-mediated\r\napoptosis. Moreover, I showed that the internalization and activation of the receptor CD95 at the\r\nplasma membrane are competing processes.\r\nTo arrive at these findings, two forms of soluble CD95L were compared, one naturally occurring\r\nform with weak activity (sCD95L) and one recombinant form with high activity (IZsCD95L). Next\r\nto biochemical methods, fluorescence-microscopy based techniques were particularly applied and\r\nI have been able to confirm that both forms are trimeric. Notably, I developed a new method\r\nto evaluate single-molecule photobleaching data suitable to infer the subunit stoichiometry of\r\nmolecules at picomolar concentration.\r\nSecondly, I aimed at understanding how natural killer cells (NK cells) mediate cell death in target\r\ncells. In addition to CD95L, NK cells use own proteases to induce the death of certain cells. In\r\nthis context, I have been able to distinguish the activity of caspase-8 and granzymes in individual\r\nHeLa target cells based on the method of Beaudouin et al. 2013 and by using the NK cell line\r\nNK92. While granzyme B activity was measured in cells that died early after NK-cell contact,\r\ncaspase-8 activity was measured in those which died late upon NK-cell contact. Noteworthy was\r\nthe observation that granzyme B was mainly active near the plasma membrane, and in lesser\r\namounts inside the nucleus. This was consistent with my results showing that the granzyme B\r\ninhibitor serpinB9 is localized at the plasma membrane. In contrast, granzyme A was probably not\r\nactive in the cytosol, but in the nucleus of the cell."^^ . "2014" . . . . . . . "Clarissa"^^ . "Liesche"^^ . "Clarissa Liesche"^^ . . . . . . "Spatio-temporal protease activity and cell death induced by\r\nNatural Killer cells and soluble CD95L (PDF)"^^ . . . "ClarissaLiesche_Thesis.pdf"^^ . . . "Spatio-temporal protease activity and cell death induced by\r\nNatural Killer cells and soluble CD95L (Other)"^^ . . . . . . "lightbox.jpg"^^ . . . "Spatio-temporal protease activity and cell death induced by\r\nNatural Killer cells and soluble CD95L (Other)"^^ . . . . . . "preview.jpg"^^ . . . "Spatio-temporal protease activity and cell death induced by\r\nNatural Killer cells and soluble CD95L (Other)"^^ . . . . . . "medium.jpg"^^ . . . "Spatio-temporal protease activity and cell death induced by\r\nNatural Killer cells and soluble CD95L (Other)"^^ . . . . . . "small.jpg"^^ . . . "Spatio-temporal protease activity and cell death induced by\r\nNatural Killer cells and soluble CD95L (Other)"^^ . . . . . . "indexcodes.txt"^^ . . "HTML Summary of #17298 \n\nSpatio-temporal protease activity and cell death induced by \nNatural Killer cells and soluble CD95L\n\n" . "text/html" . . . "570 Biowissenschaften, Biologie"@de . "570 Life sciences"@en . .