The IκB-protein BCL-3 controls toll-like receptor-induced MAPK activity by promoting TPL-2 degradation in the nucleus

Proinflammatory responses induced by Toll-like receptors (TLRs) are dependent on the activation of the NF-ĸB and mitogen-activated protein kinase (MAPK) pathways, which coordinate the transcription and synthesis of proinflammatory cytokines. We demonstrate that BCL-3, a nuclear IĸB protein that regulates NF-ĸB, also controls TLR-induced MAPK activity by regulating the stability of the TPL-2 kinase. TPL-2 is essential for MAPK activation by TLR ligands, and the rapid proteasomal degradation of active TPL-2 is a critical mechanism limiting TLR-induced MAPK activity. We reveal that TPL-2 is a nucleocytoplasmic shuttling protein and identify the nucleus as the primary site for TPL-2 degradation. BCL-3 interacts with TPL-2 and promotes its degradation by promoting its nuclear localization. As a consequence, Bcl3−/− macrophages have increased TPL-2 stability following TLR stimulation, leading to increased MAPK activity and MAPK-dependent responses. Moreover, BCL-3–mediated regulation of TPL-2 stability sets the MAPK activation threshold and determines the amount of TLR ligand required to initiate the production of inflammatory cytokines. Thus, the nucleus is a key site in the regulation of TLR-induced MAPK activity. BCL-3 links control of the MAPK and NF-ĸB pathways in the nucleus, and BCL-3–mediated TPL-2 regulation impacts on the cellular decision to initiate proinflammatory cytokine production in response to TLR activation

Pharmacological impact of FLT3 mutations on receptor activity and responsiveness to tyrosine kinase inhibitors

Acute myelogenous leukaemia (AML) is an aggressive blood cancer characterized by the rapid proliferation of immature myeloid blast cells, resulting in a high mortality rate. The 5-year overall survival rate for AML patients is approximately 25%. Circa 35% of all patients carry a mutation in the FLT3 gene which have a poor prognosis. Targeting FLT3 receptor tyrosine kinase has become a treatment strategy in AML patients possessing FLT3 mutations. The most common mutations are internal tandem duplications (ITD) within exon 14 and a single nucleotide polymorphism (SNP) that leads to a point mutation in the D835 of the tyrosine kinase domain (TKD). Variations in the ITD sequence and the occurrence of other point mutations that lead to ligand-independent FLT3 receptor activation create difficulties in developing personalized therapeutic strategies to overcome observed mutation-driven drug resistance. Midostaurin and quizartinib are tyrosine kinase inhibitors (TKIs) with inhibitory efficacy against FLT3-ITD, but exhibit limited clinical impact. In this review, we focus on the structural aspects of the FLT3 receptor and correlate those mutations with receptor activation and the consequences for molecular and clinical responsiveness towards therapies targeting FLT3-ITD positive AML

Elucidation and therapeutic targeting of the molecular mechanism of TRIB2-mediated acute myeloid leukaemia

The pseudokinase TRIB2 is a potent acute myeloid leukaemia (AML) oncogene, capable of inducing transplantable AML with a relatively short latency in murine models. Functionally, the oncogencity of TRIB2 has been linked to its degradation of CCAAT/enhancer binding-protein-alpha (C/EBPα), a transcription factor necessary for regulation of haematopoietic stem cells (HSC) and myeloid differentiation and is mutated in ∼10-15% of cytogenetically normal AMLs. Previously, we have demonstrated that elevated TRIB2 mRNA expression is associated with a small subset of C/EBPα dysregulated AML patients. However in our analysis of primary AML patient samples we reveal detectably high TRIB2 protein expression in a greater number of samples than predicted from mRNA studies compared to normal peripheral blood mononuclear cells. Here, using in vivo ubiquitination assays we determined that TRIB2 exerts its effect through K48 specific ubiquitin-dependent proteasomal degradation of C/EBPα. Peptide array analysis identified the specific amino acids involved in the direct binding of these two proteins. Site-directed mutagenesis of these amino acids demonstrated that the direct binding of TRIB2 and C/EBPα was required for TRIB2-mediated C/EBPα degradation. In order to determine if posttranslational modification of C/EBPα was a trigger for TRIB2-mediated binding and degradation, we assessed the phosphorylation of C/EBPα, often a modification involved in target substrate ubiquitination. We found that TRIB2 decreased the levels of phosphorylated Serine 21 (S21) C/EBPα through preferential binding to the phosphorylated form of S21 C/EBPα and mediating its K48 specific ubiquitin-dependent proteasomal degradation. While TRIB2 retains the canonical amino acid motifs of a kinase and the ability to bind ATP, indicative of kinase activity, the absence of phosphorylated S21 C/EBPα in the presence of TRIB2 suggests that it does not have sufficient kinase activity to enable efficient phosphotransfer. The presence of TRIB2 further blocked the ability of mitogenic stimuli to phosphorylate S21 of C/EBPα. TRIB2 thus acts to perturb the regulation and function of C/EBPα phosphorylation ultimately leading to its degradation. We propose this contributes to the leukaemic phenotype of AML cells which include increased self-renewal and proliferation. Using clinically available inhibitors of the proteasomal degradation pathway we have investigated the targeted inhibition of the TRIB2 degradation function to induce cell death in AML cells. In TRIB2 overexpressing AML cell lines, and in AML patient samples identified to have elevated levels of TRIB2, we have demonstrated that elevated TRIB2 expressing samples are more sensitive than low TRIB2 expressing samples to cell death induced by proteasomal inhibition. Our data shows that in the presence of TRIB2 phosphorylated S21 C/EBPα is a trigger for its ubiquitin dependent degradation. We propose TRIB2 mediates is leukaemogenic effects in AML through direct protein-protein interaction, perturbation of phosphorylation signalling, resulting ultimately in proteasomal mediated degradation of its target C/EBPα. As C/EBPα plays a key role in both stem cell function and myeloid differentiation in AML, the targeted inhibition of TRIB2-mediated C/EBPα degradation may provide therapeutic avenues in AML

Radio sound as material culture in the home

SIGLEAvailable from British Library Document Supply Centre-DSC:DXN024189 / BLDSC - British Library Document Supply CentreGBUnited Kingdo

Nanobodies identity an activated state of the TRIB2 pseudokinase

No abstract available

The presence of C/EBP alpha and its degradation are both required for TRIB2-mediated leukaemia

C/EBP alpha (p42 and p30 isoforms) is commonly dysregulated in cancer via the action of oncogenes, and specifically in acute myeloid leukaemia (AML) by mutation. Elevated TRIB2 leads to the degradation of C/EBP alpha p42, leaving p30 intact in AML. Whether this relationship is a cooperative event in AML transformation is not known and the molecular mechanism involved remains elusive. Using mouse genetics, our data reveal that in the complete absence of C/EBP alpha, TRIB2 was unable to induce AML. Only in the presence of C/EBP alpha p42 and p30, were TRIB2 and p30 able to cooperate to decrease the latency of disease. We demonstrate that the molecular mechanism involved in the degradation of C/EBP alpha p42 requires site-specific direct interaction between TRIB2 and C/EBP alpha p42 for the K48-specific ubiquitin-dependent proteasomal degradation of C/EBP alpha p42. This interaction and ubiquitination is dependent on a critical C terminal lysine residue on C/EBP alpha. We show effective targeting of this pathway pharmacologically using proteasome inhibitors in TRIB2-positive AML cells. Together, our data show that excess p30 cooperated with TRIB2 only in the presence of p42 to accelerate AML, and the direct interaction and degradation of C/EBP alpha p42 is required for TRIB2-mediated AML

Ueber explizite Differenzenverfahren fuer hyperbolische Systeme mit variablen Koeffizienten. T. 2

SIGLECopy held by FIZ Karlsruhe; available from UB/TIB Hannover / FIZ - Fachinformationszzentrum Karlsruhe / TIB - Technische InformationsbibliothekDEGerman