Mechanics of proplatelet elaboration

Italiano, Joseph E.; Patel–Hett, Sunita; Hartwig, John H.

doi:10.1111/j.1538-7836.2007.02487.x

Cited by 97 publications

(82 citation statements)

References 31 publications

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“…32 Mature megakaryocytes form long processes within the BM, proplatelets, from which platelets (and sometimes larger fragments) are released. [33][34][35] Although the microtubule system is required for proplatelet elongation, actin-mediated forces are required for the bending and branching of the tubules leading to proplatelet amplification. However, little is known with regard to how actin contributes to this process and the cell signaling initiating this process.…”

Section: Discussionmentioning

confidence: 99%

Serum response factor is an essential transcription factor in megakaryocytic maturation

Halene

Gao²,

Hahn³

et al. 2010

Blood

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IntroductionSerum response factor (Srf), a nearly ubiquitous transcription factor that is expressed in all hematopoietic cell types, is a founding member of the MCM1-agamous-deficiens-Srf (MADS) domain-containing family of transcription factors, and binds to so called CArG sites (CC/AT-rich/GG, with CCTTATATGG emerging as a major consensus sequence). 1 More than 200 CArG boxes control expression of more than 150 Srf target genes, including genes of the cytoskeleton as well as several immediateearly genes, for example the proto-oncogene c-fos, 2 and also bcl2, 3 whereby Srf participates in apoptosis, cell growth, differentiation, and cell-cycle regulation. Srf is a downstream target of many pathways; for example, the mitogen-activated protein kinase pathway that acts through the ternary complex factors (TCF) as well as Rho signaling, 4 which promotes actin polymerization. Srf plays an important role in the regulation of smooth, skeletal, and cardiac muscle genes 5-8 during development and in adult life including aging. 9,10 Srf can be activated to promote transcription in response to extracellular signals that induce its association with specific cofactors. The 2 families of Srf cofactors are the TCF proteins (Elk1, SAP1, SAP2), 11 and the myocardin family of proteins that includes myocardin, megakaryoblastic leukemia 1 (Mkl1), and Mkl2. 1,12,13 In some cell types, TCF proteins and Mkl1 compete for binding to Srf to activate or inhibit distinct transcription targets. 14,15 In Drosophila, Srf interaction with Mkl (MAL-D) promotes cytoskeletal strength during cellular migration. 16,17 In murine embryonic stem cells, Srf is crucial for actin cytoskeletal organization and focal adhesion assembly. 18 Murine embryos lacking Srf fail to form mesoderm and thus die early in development. 19 Prior work in our laboratory has focused on acute megakaryoblastic leukemia with the t(1;22) translocation, involving fusion between an RNA binding motif protein 15 (RBM15) and Mkl1. Mkl1 is known to act as a cofactor for Srf-mediated gene activation in muscle differentiation. 12,20 We have demonstrated a physiologic role for Mkl1 in megakaryopoiesis. 21 Mkl1 expression increases with normal megakaryocyte (Mk) differentiation, and promotes Mk polyploidization. Mkl1 knockout (KO) mice have normal hematopoietic stem cells and megakaryocyte-erythroid progenitors (Pre-Meg-E). However, there is a dramatic increase in the number of CD41 ϩ megakaryocytes with most of these having low (2N) ploidy. There is a significant decrease in the number of polyploid megakaryocytes and thrombocytopenia. Mkl1 requires Srf to enhance polyploidization during megakaryocytic differentiation. Based on these findings, we have examined the effects of the Mkl1 cofactor Srf in megakaryocyte development.We show, that Srf deletion specifically in the megakaryocytic lineage leads to macrothrombocytopenia, whereas bone marrow (BM) and spleen show significant accumulation of abnormal megakaryocytes. Examination of candidate Srf target genes reveals that several actin cyt...

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Section: Discussionmentioning

confidence: 99%

Serum response factor is an essential transcription factor in megakaryocytic maturation

Halene

Gao²,

Hahn³

et al. 2010

Blood

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“…21,22 The final step in this process is typified by the extension of numerous proplatelets from the surface of mature megakaryocytes, in the form of long pseudopodia bearing organized platelet elements at their extremities. 23 This requires the recruitment of an internal pool of membranes 24 and dynamic reorganization of the microtubules, which drive the extension of pseudopodia and coil up to delimit disc-shaped platelet elements. 25 The recent live visualization of fluorescent-labeled megakaryocytes in transgenic mice has added support for this model, showing proplatelet elements protruding into bone marrow sinusoids.…”

Section: Introductionmentioning

confidence: 99%

Intrinsic impaired proplatelet formation and microtubule coil assembly of megakaryocytes in a mouse model of Bernard-Soulier syndrome

Strassel¹,

Eckly²,

Léon³

et al. 2009

Haematologica

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The online version of this article contains a supplementary appendix. BackgroundGiant platelets and thrombocytopenia are invariable defects in the Bernard-Soulier syndrome caused by deficiency of the GPIb-V-IX complex, a receptor for von Willebrand factor supporting platelet adhesion to the damaged arterial wall. Various properties of this receptor may be considered potential determinants of the macrothrombocytopenia. Design and MethodsTo explore the underlying mechanisms of the disease, megakaryopoiesis was studied in a mouse model deficient in GPIbβ. Megakaryocytes were initially characterized in situ in the bone marrow of adult mice, after which their capacity to differentiate into proplateletbearing cells was evaluated in cultured fetal liver cells. ResultsThe number of megakaryocyte progenitors, their differentiation and progressive maturation into distinct classes and their level of endoreplication were normal in GPIbβ -/-bone marrow. However, the more mature cells exhibited ultrastructural anomalies with a thicker peripheral zone and a less well developed demarcation membrane system. GPIbβ -/-megakaryocytes could be differentiated in culture from Lin -fetal liver cells in normal amounts but the proportion of cells able to extend proplatelets was decreased by 41%. Moreover, the GPIbβ -/-cells extending proplatelets displayed an abnormal morphology characterized by fewer pseudopodial extensions with thicker shaft sections and an increased diameter of the terminal coiled elements. GPIbβ -/-released platelets were larger but retained a typical discoid shape. Proplatelet formation was similarly affected in bone marrow explants from adult mice examined by videomicroscopy. The marginal microtubular ring contained twice as many tubulin fibers in GPIbβ -/-proplatelet buds in cultured and circulating platelets. ConclusionsAltogether, these findings point to a role of the GPIb-V-IX complex intrinsic to megakaryocytes at the stage of proplatelet formation and suggest a functional link with the underlying microtubular cytoskeleton in platelet biogenesis.Key words: megakaryocyte, platelet, Bernar-Soulier, microtubules, GPIb-V-IX.Citation: Strassel C, Eckly A, Léon C, Petitjean C, Freund M, Cazenave J-P, Gachet C, and Lanza F. Intrinsic impaired proplatelet formation and microtubule coil assembly of megakaryocytes in a mouse model of Bernard-Soulier syndrome. Haematologica 2009; 94:800-810. doi:10.3324/haematol.2008 This is an open-access paper.

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“…ProPLT-producing MKs in culture release intermediate barbell structures that yield individual PLTs 2,3 . ProPLTs have been identified extending and releasing into blood vessel sinusoids both in vitro and in vivo 4,5 , and are therefore believed to mature in the circulation 3,6 . We have recently identified a new intermediate stage called the prePLT in the maturation process.…”

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confidence: 99%

Microtubule and cortical forces determine platelet size during vascular platelet production

et al. 2012

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megakaryocytes release large preplatelet intermediates into the sinusoidal blood vessels. Preplatelets convert into barbell-shaped proplatelets in vitro to undergo repeated abscissions that yield circulating platelets. These observations predict the presence of circular-preplatelets and barbell-proplatelets in blood, and two fundamental questions in platelet biology are what are the forces that determine barbell-proplatelet formation, and how is the final platelet size established. Here we provide insights into the terminal mechanisms of platelet production. We quantify circular-preplatelets and barbell-proplatelets in human blood in high-resolution fluorescence images, using a laser scanning cytometry assay. We demonstrate that force constraints resulting from cortical microtubule band diameter and thickness determine barbellproplatelet formation. Finally, we provide a mathematical model for the preplatelet to barbell conversion. We conclude that platelet size is limited by microtubule bundling, elastic bending, and actin-myosin-spectrin cortex forces.

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Mechanics of proplatelet elaboration

Cited by 97 publications

References 31 publications

Serum response factor is an essential transcription factor in megakaryocytic maturation

Serum response factor is an essential transcription factor in megakaryocytic maturation

Intrinsic impaired proplatelet formation and microtubule coil assembly of megakaryocytes in a mouse model of Bernard-Soulier syndrome

Microtubule and cortical forces determine platelet size during vascular platelet production

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