Thrombopoietin Induces Tyrosine Phosphorylation and Activation of Mitogen-Activated Protein Kinases in a Human Thrombopoietin-Dependent Cell Line

Yamada, Minami; Komatsu, Norio; Okada, Koji; Kato, Takashi; Miyazaki, Hiroshi; Miura, Yasusada

doi:10.1006/bbrc.1995.2768

Cited by 54 publications

(23 citation statements)

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“…Several groups have reported that components of Janus kinase/signal transducer and activator of transcription (JAK/STAT) (Drachman et al, 1995;Wendling and Vainchenker, 1998) and Shc-Ras-MAPK pathways (Nagata and Todokoro, 1995;Yamada et al, 1995;Rouyez et al, 1997) are strongly activated in response to TPO. In particular, the importance of the mitogen-activated protein kinase (MAPK) pathway in megakaryocytopoiesis was corroborated by introduction of constitutively activated mutants of MAPK kinase (MEK) in several cell lines (Whalen et al, 1997;Racke et al, 1997;Melemed et al, 1997) and by treatment of cell lines and primary cells with pharmacological inhibitors of MEK such as PD98059 and UO126 (Rojnuckarin et al, 1999;Fichelson et al, 1999).…”

Section: Introductionmentioning

confidence: 99%

Inhibition of TPO-induced MEK or mTOR activity induces opposite effects on the ploidy of human differentiating megakaryocytes

Guerriero

Parolini

Testa

et al. 2006

Journal of Cell Science

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The megakaryocyte is a paradigm for mammalian polyploid cells. However, the mechanisms underlying megakaryocytic polyploidization have not been elucidated. In this study, we investigated the role of Shc-Ras-MAPK and PI3K-AKT-mTOR pathways in promoting megakaryocytic differentiation, maturation and polyploidization. CD34+ cells, purified from human peripheral blood, were induced in serum-free liquid suspension culture supplemented with thrombopoietin (TPO) to differentiate into a virtually pure megakaryocytic progeny (97-99% CD61+/CD41+ cells). The early and repeated addition to cell cultures of low concentrations of PD98059, an inhibitor of MEK1/2 activation, gave rise to a population of large megakaryocytes showing an increase in DNA content and polylobated nuclei (from 45% to 70% in control and treated cultures, respectively). Conversely, treatment with the mTOR inhibitor rapamycin strongly inhibited cell polyploidization, as compared with control cultures. Western blot analysis of PD98059-treated progenitor cells compared with the control showed a downmodulation of phospho-ERK 1 and phospho-ERK 2 and a minimal influence on p70S6K activation; by contrast, p70S6K activation was completely inhibited in rapamycin-treated cells. Interestingly, the cyclin D3 localization was nuclear in PD98059-induced polyploid megakaryocytes, whereas it was completely cytoplasmic in those treated with rapamycin. Altogether, our results are in line with a model in which binding of TPO to the TPO receptor (mpl) could activate the rapamycin-sensitive PI3K-AKT-mTOR-p70S6K pathway and its downstream targets in promoting megakaryocytic cell polyploidization.

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Section: Introductionmentioning

confidence: 99%

Inhibition of TPO-induced MEK or mTOR activity induces opposite effects on the ploidy of human differentiating megakaryocytes

Guerriero

Parolini

Testa

et al. 2006

Journal of Cell Science

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“…TCFs are important mediators of the MAP kinase pathway, 6,7 which plays a major role in MK differentiation. 57,58 Thus, a part of the effect of MAL overexpression might be related to an alteration in this pathway. However, Cheng et al reported also a decrease in ploidy level in bone marrow MKs of MKL1 Ϫ/Ϫ mice.…”

Section: Discussionmentioning

confidence: 99%

MAL/SRF complex is involved in platelet formation and megakaryocyte migration by regulating MYL9 (MLC2) and MMP9

Gilles

Bluteau

Boukour

et al. 2009

Blood

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Megakaryoblastic leukemia 1 (MAL) is a transcriptional coactivator of serum response factor (SRF). In acute megakaryoblastic leukemia, the MAL gene is translocated and fused with the gene encoding one twenty-two (OTT). Herein, we show that MAL expression increases during the late differentiation steps of neonate and adult human megakaryopoiesis and localized into the nucleus after Rho GTPase activation by adhesion on collagen I or convulxin. MAL knockdown in megakaryocyte progenitors reduced the percentage of cells forming filopodia, lamellipodia, and stress fibers after adhesion on the same substrates, and reduced proplatelet formation. MAL repression led to dysmorphic megakaryocytes with disorganized demarcation membranes and ␣ granules heterogeneously scattered in the cytoplasm. Gene expression profiling revealed a marked decrease in metalloproteinase 9 (MMP-9) and MYL9 expression after MAL inhibition. Luciferase assays in HEK293T cells and chromatin immunoprecipitation in primary megakaryocytes showed that the MAL/SRF complex directly regulates MYL9 and MMP9 in vitro. Megakaryocyte migration in response to stromal cell-derived factor 1, through Matrigel was considerably decreased after MAL knockdown, implicating MMP9 in migration. Finally, the use of a shRNA to decrease MYL9 expression showed that MYL9 was involved in proplatelet formation. MAL/SRF complex is thus involved in platelet formation and megakaryocyte migration by regulating MYL9 and MMP9. IntroductionSerum response factor (SRF) is a widely expressed transcription factor required for the expression of immediate early, musclespecific, and cytoskeletal genes. 1-4 SRF contains a MADS domain that mediates homodimerization and DNA binding, and that allows recruitment of transcriptional cofactors. SRF binds to a CArG box present in promoter/enhancer regions of SRF-regulated genes. 5 Depending on cell lines, different extracellular stimuli activate SRF through 2 main signaling pathways: the MAP-kinase pathway through members of the ternary complex factor (TCF) 6,7 and the small GTPases pathway through the Rho family 8 members regulating the myocardin-related transcription factors (MRTFs). The Rho-actin signaling pathway 9-12 stimulates SRF by 2 ubiquitous MRTFs, megakaryoblastic leukemia 1 (MAL; MKL1, MRTF-A, BSAC) and MAL16 (MKL2, MRTF-B).MAL was initially identified in acute megakaryoblastic leukemia (AMKL, M7) as a chromosome 22 encoded protein fused in 3Ј with RNA-binding motif protein 15 (RBM15; OTT) located on chromosome 1. [13][14][15] The translocation t(1,22)(p13;q13) leads to the in-frame fusion of the quasi-totality of OTT/RBM15 to the MAL gene. The OTT-MAL fusion protein is restricted to AMKL occurring de novo in infancy, 16,17 in children older than 1 year or, occasionally, in Down syndrome patients. 18,19 The subcellular localization of MAL is regulated through its association with globular actin by its RPEL motifs in the Nterminal region. The modification of the actin treadmilling by the Rho pathway results in the nuclear accumulation of ...

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“…The MAPK pathway involves a protein kinase cascade, with two of the MAPKs, extracellular signal-related protein kinase 1 (ERK1) and ERK2, having been shown to be involved in TPO signaling. [86][87][88] Thrombopoietin has been shown to activate this pathway in human CD34 + cells, a population that includes HSCs, as well as in megakaryocytes and platelets. 89 MAPK signaling is required for TPO induced Hox activation.…”

Section: Signaling In Myeloproliferative Disordersmentioning

confidence: 99%

Thrombopoietin and hematopoietic stem cells

Graaf

Metcalf²

2011

Cell Cycle

129

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Thrombopoietin Induces Tyrosine Phosphorylation and Activation of Mitogen-Activated Protein Kinases in a Human Thrombopoietin-Dependent Cell Line

Cited by 54 publications

References 0 publications

Inhibition of TPO-induced MEK or mTOR activity induces opposite effects on the ploidy of human differentiating megakaryocytes

Inhibition of TPO-induced MEK or mTOR activity induces opposite effects on the ploidy of human differentiating megakaryocytes

MAL/SRF complex is involved in platelet formation and megakaryocyte migration by regulating MYL9 (MLC2) and MMP9

Thrombopoietin and hematopoietic stem cells

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