Antiapoptotische Wirkungen des Makrophagen-migrationsinhibierenden Faktors (MIF) auf das Tumorsuppressorprotein p53 bei der ROS-induzierten Apoptose

In the present thesis, intrinsic (ROS -> p53-activation / ROS -> mitochondrial apoptosis) and extrinsic (PI3K-Akt-Pathway) effects of the cytokine MIF (Macrophage Migration Inhibitory factor) have been examined and brought in connection with antiapoptotic and pro-proliferative effects. The functions as redox-active cytokine (redox acting cytokine = "Redoxkine"), which have been linked to anti-apoptotic characteristics of MIF could be confirmed and extended to cellular ROS models. In this context it was discovered that MIF is involved in the scavenging of ROS (reactive oxygen species) and thus contributes to maintain redox homeostasis. In the absence of MIF the generation of exogenous ROS favours the breakdown of the redox equilibrium and the emergence of ROS induced damages, which is why a premature and stronger course of mitochondrial apoptosis was observed in MIF-deficient MEF cells. Tissue sections of murine carotid arteries also showed first indices that the ROS production in intimal cells of MIF-/-LDLR-/- animals is favoured during the process of inflammation in a wire injury model. In particular, the mechanistical connection between compensatory effects of MIF during ROS stress and the activation and stabilization of the p53 tumor suppressor protein have been examined. Agreeing with previous postulates, increased protein concentrations of p53 could be observed in MIF-deficient MEF cells. Activation of p53 at the ATM/ATR-phosphorylation site Ser15 was clearly increased in these cells during ROS stress, whereby the accumulation of p53 becomes explainable by an inhibition of the Mdm2-mediated degradation of the protein. Activation of p53 was accompanied by a increased expression of the apoptogenic p53-targets Bax and PUMA (BH-3-Domain-proteins), which promote mitochondrial apoptosis was also detectable in the MIF-/-MEF cells. In addition it was observed, that during ROS stress the COP9/CSN-specific phosphorylation of p53 at Thr155 is supported in the presence of MIF. Thus MIF seems to exert a bivalent influence on the posttranslational stabilization of p53 (inhibition of p53-activation and promotion of p53-degradation). Regarding the COP9/CSN-dependent degradation of p53, the modulation by MIF is probably mediated through protein-protein interactions (MIF-CSN5/JAB1 (CSN5/JAB1-monomer or MIF-mini-CSN5/JAB1-complex)). Although a physical interaction of MIF and p53 was not detected, a competition between MIF and p53 for the common binding partner CSN5/JAB1, which was observed during Coimmunoprecipitation experiments, is likely. As a result of an increased basal transactivation activity of p53 in MIF-deficient MEFs, increased protein concentrations of the cell cycle inhibitor p21 and the tumor suppressor PTEN have been detected in these cells. Since PTEN inhibits Akt-kinase activity, it was examined whether MIF -beside its anti-apoptotic effects on p53- can cause an extrinsic activation of the Akt signaling pathway. In MEF cells, the activation of Akt could be induced by recombinant MIF. In MIF-/-MEFs Akt was activated when the cells were incubated with culture media from MIF positive MEFs. Akt was also induced by removal and re-accumulation of MIF in the culture media of MIF-positive MEFs. The activation of Akt could be blocked with MIF-specific antibodies, why an autocrine effect of MIF on the Akt pathway is assumed. In conjunction with investigations on the degradation of MIF during the process of ROS induced apoptosis a potential monoubiquitination of MIF was discovered, which seems to disappear under hyperoxic conditions. At present, no function regarding the secretion of MIF can be assigned to this modification. First attempts also resulted in no references that MIF is degradated by the proteasome.

Grantor dc:publisher

Publikationsserver der RWTH Aachen University

Year dc:date

2006

Author dc:creator

• Thiele, Michael Robert

Contributors dc:contributor

• Bernhagen, Jürgen

• info:eu-repo/classification/ddc/570
• Biowissenschaften, Biologie

Statement dc:rights

• info:eu-repo/semantics/openAccess

Language dc:language

ger

Identifier

https://publications.rwth-aachen.de/search?p=id:%22RWTH-CONV-123893%22

OAI identifier oai:identifier

oai:publications.rwth-aachen.de:62319