Die Rolle negativer Phosphorylierungen in der Regulation klassischer NF-kappaB-Signalwege

The classical NF-kappaB-signaling is not only one uniform pathway but consists of several distinct signaling pathways which differ regarding their inducing agents, the intracellular proteins involved in signal transmission, the kind of activated NF-kappaB dimers and the set of induced target genes. However, the precise molecular mechanisms by which these differential NF-kappaB activation patterns are determined still remain largely unknown. The key regulator within the classical NF-kappaB-signaling cascades is the IKK-complex. Since structural alterations in the IKK-complex are thought to participate in the regulation of its kinase activity, protein-protein-interactions within the IKK-complex were characterized. The IBD in NEMO has been shown to mediate the interaction with the IKKs. However, our studies reveal that the IBD additionally constitutes an independent oligomerization interface. Interestingly, the IBD possesses several phosphorylation sites. Yet, the physiological functions are still largely unknown. IKKß-mediated phosphorylation of Ser68 in the IBD caused a reduced NEMO-IKKß interaction and NEMO oligomerization. The negative effect of the Ser68-phosphorylation on the classical NF-kappaB-signaling pathways underline the importance of phosphatases and specific dephosphorylation events for the classical NF-kappaB activation. Therefore the yet poorly defined role of the protein phosphatase calcineurin in the TCR-induced NF-kappaB activation was analyzed. Our results point out that calcineurin acts as a BCL10 phosphatase that removes inhibitory phosphate groups from BCL10 in the early activation state. Therefore a molecular basis of how calcium influences the TCR-induced NF-kiappaB-signaling pathway is the calcineurin-mediated regulation of the CBM-complex. In summary these data underscore the importance of negatively regulating phosphorylations, which are necessary to avoid an unwanted chronic NF-kappaB activation and highlight the relevance of phosphatases in the process of IKK activation

Ulm University Medicine | 2015: Research and Teaching

It is our great pleasure to present this edition of “Ulm University Medicine – Research & Teaching 2015”. You will find details of all the latest developments and newest achievements of the Medical Faculty Ulm and of the Ulm University Medical Center. Ulm University Medicine is dedicated to providing excellent teaching and training for our future scientists and physicians in order to enhance high quality research and health care

The Ca2+-dependent Phosphatase Calcineurin Controls the Formation of the Carma1-Bcl10-Malt1 Complex during T Cell Receptor-induced NF-κB Activation*

T cell receptor (TCR) ligation induces increased diacylglycerol and Ca2+ levels in T cells, and both secondary messengers are crucial for TCR-induced nuclear factor of activated T cells (NF-AT) and NF-κB signaling pathways. One prominent calcium-dependent enzyme involved in the regulation of NF-AT and NF-κB signaling pathways is the protein phosphatase calcineurin. However, in contrast to NF-AT, which is directly dephosphorylated by calcineurin, the molecular basis of the calcium-calcineurin dependence of the TCR-induced NF-κB activity remains largely unknown. Here, we demonstrate that calcineurin regulates TCR-induced NF-κB activity by controlling the formation of a protein complex composed of Carma1, Bcl10, and Malt1 (CBM complex). For instance, increased calcium levels induced by ionomycin or thapsigargin augmented the phorbol 12-myristate 13-acetate-induced formation of the CBM complex and activation of NF-κB, whereas removal of calcium by the calcium chelator EGTA-acetoxymethyl ester (AM) attenuated both processes. Furthermore, inhibition of the calcium-dependent phosphatase calcineurin with the immunosuppressive agent cyclosporin A (CsA) or FK506 as well as siRNA-mediated knockdown of calcineurin A strongly affected the PMA + ionomycin- or anti-CD3 + CD28-induced CBM complex assembly. Mechanistically, the positive effect of calcineurin on the CBM complex formation seems to be linked to a dephosphorylation of Bcl10. For instance, Bcl10 was found to be hyperphosphorylated in Jurkat T cells upon treatment with CsA or EGTA-AM, and calcineurin dephosphorylated Bcl10 in vivo and in vitro. Furthermore, we show here that calcineurin A interacts with the CBM complex. In summary, the evidence provided here argues for a previously unanticipated role of calcineurin in CBM complex formation as a molecular basis of the inhibitory function of CsA or FK506 on TCR-induced NF-κB activity