Signaling by the Mpl receptor involves IKK and NF‐κB*

Abstract

Binding of tumor necrosis factor-alpha (TNF-alpha) to its receptor activates IKK complex, which leads to inducement of NF-kappaB activity. Here we report that activation of Mpl ligand is also linked to IKK and NF-kappaB activity. Mpl ligand, also known as thrombopoietin (TPO) or megakaryocyte growth and development factor (MGDF), induces megakaryocyte differentiation and inhibition of mitotic proliferation, followed by induction of polyploidization and fragmentation into platelets. The latter process is often observed in megakaryocytes undergoing apoptosis. Treatment of a Mpl ligand-responding megakaryocytic cell line with this cytokine led to an immediate, transient increase in IKK activity followed by a profound decrease in this kinase activity over time. This decrease was not due to an effect on the levels of the IKK regulatory components IKKalpha and IKKbeta. Proliferating megakaryocytes displayed a constitutive DNA-binding activity of NF-kappaB p50 homodimers and of NF-kappaB p50-p65 heterodimers. As expected, reduced IKK activity in Mpl ligand-treated cells was associated with a significant reduction in NF-kappaB DNA binding activity and in the activity of a NF-kappaB-dependent promoter. Our study is thus the first to identify a constitutive NF-kappaB activity in proliferating megakaryocytes as well as to describe a link between Mpl receptor signaling and IKK and NF-kappaB activities. Since a variety of proliferation-promoting genes and anti-apoptotic mechanisms are activated by NF-kappaB, retaining its low levels would be one potential mechanism by which inhibition of mitotic proliferation is maintained and apoptosis is promoted during late megakaryopoiesis.

Rapid recovery from panobinostat (LBH589)-induced thrombocytopenia in mice involves a rebound effect of bone marrow megakaryocytes

“…However, both methods indicate increased numbers of megakaryocytes, especially smaller immature forms, at day 6 in panobinostat-treated mice, followed by a return to normal megakaryocyte levels by day 19. This finding is also consistent with a model of thrombocytopenia involving temporary disruption of nuclear factor-kB signaling, as the NFkB transcription factor is important in megakaryocyte maturation, 8 and panobinostat is known to block nuclear translocation of NFkB. 6 A study by Matsuoka et al 12 also found that megakaryocyte numbers in rat spleen increase in response to the histone deacetylase inhibitor FR235225, and that inhibition of GATA-1 in megakaryocytes is associated with histone deacetylase inhibitor-induced thrombocytopenia.…”

“…The mechanisms of bortezomib-and panobinostat-induced thrombocytopenia are not well understood, but both may involve effects on nuclear factor-kB (NFkB) nuclear translocation and downstream gene targets. [6][7][8] Here, we evaluated the kinetics of thrombocytopenia and the fate of BM megakaryocytes in panobinostat-treated mice. We hypothesized that the rate of blood platelet reduction would reflect a panobinostat-induced cessation of platelet production, rather than megakaryocyte loss, and that platelet generation by megakaryocytes would resume on discontinuation of panobinostat treatment.…”

Mechanism of Aurora-B Degradation and Its Dependency on Intact KEN and A-Boxes: Identification of an Aneuploidy-Promoting Property

“…Protein was subjected to Western blot analysis (40 g/lane) as described previously (52). Immunoprecipitation experiments were pursued according to standard procedures (Santa Cruz Biotech, Santa Cruz, CA) and as described previously (53). Briefly, 500 g of soluble protein was first incubated with primary antibodies for 2 h at room temperature and further incubated overnight at 4°C after addition of 25 l of protein A/GSepharose beads (Santa Cruz Biotech, Santa Cruz, CA).…”

The A2B adenosine receptor protects against inflammation and excessive vascular adhesion

“…Fifty micrograms/sample of total protein was electrophoresed on 6% or 10% SDS-PAGE gels and transferred overnight at 4°C to Immobilon-P PVDF membranes (Millipore), and Western blotting was performed as described in ref. 58 …”