Constitutive Stat3, Tyr705, and Ser727 phosphorylation in acute myeloid leukemia cells caused by the autocrine secretion of interleukin-6.
ABSTRACT To explore the activation patterns of signal transducer and activator of transcription 3 (Stat3) in acute myeloid leukemia (AML), we examined whether the phosphorylation of tyrosine705 (Tyr705) and serine727 (Ser727) residues was abnormally regulated in cells from patients with AML. In 5 of 20 (25%) patients with AML, Stat3 was constitutively phosphorylated on Tyr705 and Ser727, which were not further up-regulated by treatment with IL-6. Furthermore, Stat3 was constitutively bound to the IRE response element in these cells as determined by electrophoretic mobility shift assay, and stimulation with IL-6 did not result in increased DNA binding. Interestingly, AML cells with constitutive Stat3 activation also secreted high levels of IL-6 protein. Treating these AML cells with anti-IL-6 resulted in restored IL-6-inducible Stat3 phosphorylation on both Tyr705 and Ser727 with low or undetectable basal phosphorylation levels in unstimulated cells. In contrast, treatment with anti-IL-1 did not result in altered Stat3 phosphorylation patterns. The constitutive IL-6 expression was associated with elevated levels of suppressor of cytokine signaling-1 (SOCS-1) and SOCS-3 mRNA expression, which were not down-regulated by anti-IL-6. These data indicate that the constitutive Stat3 activation in the investigated AML blasts is caused by high IL-6 secretion levels, thus stimulating the Jak/Stat pathway in an autocrine manner, a paracrine manner, or both. (Blood. 2000;95:3765-3770)
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ABSTRACT: We previously showed that CD34+/CD38− acute myelogenous leukemia (AML) cells, which contain leukemia stem cells, expressed a greater amount of the phosphorylated forms of JAK2 and STAT5 (p-JAK2 and p-STAT5) than their CD34+/CD38+ counterparts. To identify candidate cytokines that are involved in the activation of JAK2/STAT5 in CD34+/CD38− AML cells, we compared the cytokine expression profiles of CD34+/CD38− AML cells and their CD34+/CD38+ counterparts. Interestingly, freshly isolated CD34+/CD38− AML cells from patients (n = 17) expressed less interleukin-1β (IL-1β) than their CD34+/CD38+ counterparts and CD34+ normal hematopoietic stem/progenitor cells from healthy volunteers (n = 6), as measured by real-time Reverse Transcription-Polymerase Chain Reaction (RT-PCR). Methylation-specific PCR found that IL-1B gene expression was silenced by methylation of the promoter region. Importantly, exposure of CD34+/CD38− AML cells to IL-1β (100 ng/ml) stimulated cell-cycle progression, induced apoptosis and sensitized these cells to growth inhibition by antileukemia agents. These changes occurred in conjunction with the downregulation of cyclin-dependent kinase inhibitor p21waf1, antiapoptotic proteins and p-STAT5. Forced expression of IL-1β in CD34+/CD38− AML cells by lentiviral transduction significantly impaired the self-renewal capacity of the cells and induced apoptosis. Additionally, when these CD34+/CD38− AML cells with forced expression of IL-1β were transplanted into severely immunocompromised mice, the engraftment of the cells and reconstitution of AML were significantly impaired. Taken together, our results indicate that the inhibition of STAT5 by IL-1β may be a promising treatment strategy to eradicate leukemia stem cells in AML.International Journal of Cancer 10/2013; 133(8). · 6.20 Impact Factor
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ABSTRACT: Mature Large Granular lymphocytes (LGL) disorders include a spectrum of conditions, ranging from polyclonal to clonal indolent and/or overt leukemic LGL proliferations. Most cases are represented by clonal expansions of TCRα/β+ LGL displaying a CD8+ phenotype with expression of cytotoxic T-cell antigens (CD57, CD16, TIA-1, perforin and granzyme B). Proliferations of CD3-CD16+ NK cells with a restricted patter of NK receptors are less common, usually comprising 15% of the cases. Main features are cytopenias, splenomegaly and autoimmune phenomena. Morphology, immunophenotyping and molecular analyses are crucial to establish a correct diagnosis of disease. According to the 2008 WHO classification, two separate entities account for the majority of cases, T-LGL leukemia and Chronic Lymphoproliferative Disease of NK cell (this latter still provisional). Although these disorders are characterized by the expansion of different cells types i.e. T and NK cells, with specific genetic features and abnormalities, compelling evidence supports the hypothesis that a common pathogenic mechanism would be involved in both disorders. As a matter of fact, a foreign antigen driven clonal selection is considered the initial step in the mechanism ultimately leading to generation of both conditions. In this chapter we will discuss recent advances on the pathogenesis of chronic T and NK disorders of granular lymphocytes, challenging the current WHO classification on the opportunity to separate T and NK disorders, which are likely to represent two sides of the same coin.Translational medicine @ UniSa. 01/2014; 8:4-11.
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ABSTRACT: Phosphorylation is a fundamental biochemical reaction that modulates protein activity in cells. While a single phosphorylation event is relatively easy to understand, multisite phosphorylation requires systems approaches for deeper elucidation of the underlying molecular mechanisms. In this paper we develop a mechanistic model for single- and multi-site phosphorylation. The proposed model is compared with previously reported studies. We compare the predictions of our model with experiments published in the literature in the context of inflammatory signaling events in order to provide a mechanistic description of the multisite phosphorylation-mediated regulation of Signal Transducer and Activator of Transcription 3 (STAT3) and Interferon Regulatory Factor 5 (IRF-5) proteins. The presented model makes crucial predictions for transcription factor phosphorylation events in the immune system. The model proposes potential mechanisms for T cell phenotype switching and production of cytokines. This study also provides a generic framework for the better understanding of a large number of multisite phosphorylation-regulated biochemical circuits.PLoS ONE 10/2014; 9(10):e110913. · 3.53 Impact Factor