Tumor Suppressor Protein p53 Regulates Megakaryocytic Polyploidization and Apoptosis

Fuhrken, Peter G.; Apostolidis, Pani A.; Lindsey, Stephan; Miller, William M.; Papoutsakis, Eleftherios T.

doi:10.1074/jbc.m801923200

Cited by 40 publications

(58 citation statements)

References 43 publications

(44 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…For baseline ploidy analysis, cells were analyzed as described previously with minor modifications. 23 After BM isolation, 2 ϫ 10 6 cells were labeled with an FITC-conjugated CD41 antibody for 30 minutes at 4°C. The cells were then washed twice in 2mM EDTA in PBS and fixed in 0.5% formalin for 10 minutes at RT.…”

Section: Megakaryocyte Ploidy Analysismentioning

confidence: 99%

Calcium- and integrin-binding protein 1 regulates megakaryocyte ploidy, adhesion, and migration

Kostyak¹,

Naik

2012

Blood

View full text Add to dashboard Cite

Megakaryocytes are large, polyploid cells that produce platelets. We have previously reported that calcium-and integrin-binding protein 1 (CIB1) regulates endomitosis in Dami cells. To further characterize the role of CIB1 in megakaryopoiesis, we used a Cib1 ؊/؊ mouse model. Cib1 ؊/؊ mice have more platelets and BM megakaryocytes than wild-type (WT) controls (P < .05). Furthermore, subsequent analysis of megakaryocyte-CFU production revealed an increase with Cib1 deletion compared with WT (P < .05). In addition, BM from Cib1 ؊/؊ mice, cultured with thrombopoietin (TPO) for 24 hours, produced more highly polyploid megakaryocytes than WT BM (P < .05). Subsequent analysis of TPO signaling revealed enhanced Akt and ERK1/2 phosphorylation, whereas FAK Y925 phosphorylation was reduced in Cib1 ؊/؊ megakaryocytes treated with TPO. Conversely, platelet recovery in Cib1 ؊/؊ mice after platelet depletion was attenuated compared with WT (P < .05). This could be the result of impaired adhesion and migration, as adhesion to fibrinogen and fibronectin and migration toward an SDF-1␣ gradient were reduced in Cib1 ؊/؊ megakaryocytes compared with WT (P < .05). In addition, Cib1 ؊/؊ megakaryocytes formed fewer proplatelets compared with WT (P < .05), when plated on fibrinogen. These data suggest that CIB1 plays a dual role in megakaryopoiesis, initially by negatively regulating TPO signaling and later by augmenting proplatelet production. IntroductionMature megakaryocytes produce platelets by extending proplatelet projections into sinusoidal vessels in BM. 1 Before this process, the megakaryocyte must differentiate from progenitor cells by undergoing multiple rounds of endomitosis. Megakaryocyte endomitosis occurs primarily through cell signaling initiated by the cytokine thrombopoietin (TPO), which is produced constitutively in the liver. 2 TPO binding to its receptor, c-Mpl, results in the activation of Janus kinase 2 (Jak2). Jak2 phosphorylates tyrosine residues on c-Mpl, which enables the subsequent activation of other signaling pathways, such as PI3K/Akt and MAPK. [3][4][5][6][7][8] In addition, focal adhesion kinase (FAK) is activated and acts as a negative regulator of TPO-induced signaling by activating Lyn, an Src-family kinase. 9 Specifically, Western blot analysis revealed that impaired TPO-induced FAK phosphorylation is concomitant with heightened Akt and ERK1/2 phosphorylation. 9 Once mature, megakaryocytes must migrate from an "osteoblastic niche" to a "vascular niche" where they interact with BM endothelial cells. 10 Integrins play a key role in cell migration, and the most abundantly expressed integrin on the megakaryocyte is ␣ IIb ␤ 3 , which is the major receptor for the highly expressed BM extracellular matrix (ECM) protein fibronectin (Fn). 11,12 The exact mechanism underlying megakaryocyte migration toward the vasculature is unknown, but stromal cell-derived factor-1␣ (SDF-1␣) appears to provide the homing signal. 13 Calcium-and integrin-binding protein 1 (CIB1) was first identified as a binding partner of th...

show abstract

Section: Megakaryocyte Ploidy Analysismentioning

confidence: 99%

Calcium- and integrin-binding protein 1 regulates megakaryocyte ploidy, adhesion, and migration

Kostyak¹,

Naik

2012

Blood

View full text Add to dashboard Cite

show abstract

“…An interesting insight might be provided by the analysis of developmentally programmed tetraploids, as similar mechanisms might apply in both aberrant and scheduled tetraploidization. For example, it was recently shown that a deficiency in p53 increases the level of polyploidization in megakaryocytes, which become highly polyploid in the developmentally programmed process of thrombocyte formation (Fuhrken et al, 2008).…”

Section: Signals That Trigger the Arrest Of Tetraploidsmentioning

confidence: 99%

The consequences of tetraploidy and aneuploidy

Storchová

Kuffer

2008

Journal of Cell Science

327

323

View full text Add to dashboard Cite

Polyploidy, an increased number of chromosome sets, is a surprisingly common phenomenon in nature, particularly in plants and fungi. In humans, polyploidy often occurs in specific tissues as part of terminal differentiation. Changes in ploidy can also result from pathophysiological events that are caused by viral-induced cell fusion or erroneous cell division. Tetraploidization can initiate chromosomal instability (CIN), probably owing to supernumerary centrosomes and the doubled chromosome mass. CIN, in turn, might persist or soon give way to a stably propagating but aneuploid karyotype. Both CIN and stable aneuploidy are commonly observed in cancers. Recently, it has been proposed that an increased number of chromosome sets can promote cell transformation and give rise to an aneuploid tumor. Here, we review how tetraploidy can occur and describe the cellular responses to increased ploidy. Furthermore, we discuss how the specific physiological changes that are triggered by polyploidization might be used as novel targets for cancer therapy. Tetraploid cells can also be created after an aberrant cell division. During mitosis, the chromosomes attach via proteinaceous structures called kinetochores to spindle microtubules that emanate from MTOCs (Box 1). This enables cells to segregate their chromosomes evenly into two daughter cells. Spindle-assembly checkpoint (SAC) activity holds back the onset of anaphase until all kinetochores are properly attached (Musacchio and Salmon, 2007). If there is a persistent error, the cell can escape SAC arrest (Brito and Rieder, 2006) and exit from mitosis without undergoing anaphase or cytokinesis, thereby producing a tetraploid cell with a single nucleus and two centrosomes (Azeddine et al., 1998;Lanni and Jacks, 1998). This so-called 'mitotic slippage' also occurs in cells that have an altered SAC, such as mouse embryonic fibroblasts (MEFs), which overexpress the SAC gene Mad2 (mitotic-arrest deficient 2) (Sotillo et al., 2007).In addition, cells that have entered anaphase might fail to finalize cell division. Cytokinesis might fail owing to a disturbance of cleavage-furrow formation, which occurs when bulk chromatin (Mullins and Biesele, 1977), or even a single lagging chromosome, is trapped in the cleavage furrow (Shi and King, 2005). The result is a single binucleated cell with two centrosomes. Abnormal spindle positioning and movements might also interfere with cytokinesis and lead to the accumulation of tetraploid cells, as has been observed, for example, in cells with deregulated integrin functions that inhibited spindle assembly (Reverte et al., 2006).The list of mechanisms that lead to tetraploidy is growing, and raises the issue of how frequently unscheduled tetraploidization occurs in normal tissues. Although difficult to estimate, tetraploid cells can be found with variable frequencies (0.5-20%) in nearly every human tissue (Biesterfeld et al., 1994), suggesting that tetraploidization is a more common process than was previously thought. In fact, spontaneous unsched...

show abstract

“…After that, the cells with the MPs were diluted in 600 mL IMDM medium supplemented with 5% BIT9500 and 50 ng/mL rhSCF, and cultured at 37°C and 20% O 2 until they were harvested at day 8 for CD41 and ploidy flow cytometry analysis. 25 At day 9, cells in coculture were examined using multiphoton confocal microscope (Zeiss 510 NLO) and Differential Interference Contrast images were collected. At day 10, cells from coculture were seeded onto human fibrinogen-coated coverslips and cultured overnight for staining for b1 tubulin (TUBB1), VWF, and serotonin (5-HT) at day 11.…”

Section: Isolation and Characterization Of Mkmpsmentioning

confidence: 99%

Shear enhances thrombopoiesis and formation of microparticles that induce megakaryocytic differentiation of stem cells

Jiang

Woulfe

Papoutsakis

2014

Blood

Self Cite

134

View full text Add to dashboard Cite

• Physiological shear stress promotes megakaryocytic maturation, DNA synthesis, phosphatidylserine exposure and caspase-3 activation.• Shear enhances the production and function of PLPs and Mk-derived microparticles possessing a novel function.In vivo visualization of thrombopoiesis suggests an important role for shear flow in platelet biogenesis. In vitro, shear stress was shown to accelerate proplatelet formation from mature megakaryocytes (Mks). Yet, the role of biomechanical forces on Mk biology and platelet biogenesis remains largely unexplored. In this study, we investigated the impact of shear stress on Mk maturation and formation of platelet-like particles (PLPs), pro/preplatelets (PPTs), and Mk microparticles (MkMPs), and furthermore, we explored a physiological role for MkMPs. We found that shear accelerated DNA synthesis of immature Mks in an exposure time-and shear stress level-dependent manner. Both phosphatidylserine exposure and caspase-3 activation were enhanced by shear stress. Exposure to physiological shear dramatically increased generation of PLPs/PPTs and MkMPs by up to 10.8 and 47-fold, respectively. Caspase-3 inhibition reduced shear-induced PLP/PPT and MkMP formation. PLPs generated under shear flow displayed improved functionality as assessed by CD62P exposure and fibrinogen binding. Significantly, coculture of MkMPs with hematopoietic stem and progenitor cells promoted hematopoietic stem and progenitor cell differentiation to mature Mks synthesizing a-and dense-granules, and forming PPTs without exogenous thrombopoietin, thus identifying a novel and unexplored potential physiological role for MkMPs. (Blood. 2014;124(13):2094-2103

show abstract

Tumor Suppressor Protein p53 Regulates Megakaryocytic Polyploidization and Apoptosis

Cited by 40 publications

References 43 publications

Calcium- and integrin-binding protein 1 regulates megakaryocyte ploidy, adhesion, and migration

Calcium- and integrin-binding protein 1 regulates megakaryocyte ploidy, adhesion, and migration

The consequences of tetraploidy and aneuploidy

Shear enhances thrombopoiesis and formation of microparticles that induce megakaryocytic differentiation of stem cells

Contact Info

Product

Resources

About