Integrins Regulate the Intracellular Distribution of Eukaryotic Initiation Factor 4E in Platelets

Lindemann, Stephan; Tolley, Neal D.; Eyre, Jennifer R.; Kraiss, Larry W.; Mahoney, Tracey M.; Weyrich, Andrew S.

doi:10.1074/jbc.m104281200

Cited by 103 publications

(168 citation statements)

References 24 publications

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“…Platelets retain numerous mRNAs from megakaryocytes, these mRNAs are functional and translate to protein by cellular activation [24]. There are some previous studies in which regular RT-PCR has been used to detect mRNA in platelets [19][20][21], but no-one has been able to set up a quantification method, probably due to the large amount of total RNA needed for previous RNA quantification methods.…”

Section: Discussionmentioning

confidence: 99%

Quantification of ADP and ATP receptor expression in human platelets

Wang

Östberg

Wihlborg

et al. 2003

Journal of Thrombosis and Haemostasis

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Summary. The mechanism of ADP-mediated platelet activation has been difficult to unravel due to the large number of receptors for extracellular nucleotides (P2 receptors). mRNA levels in circulating platelets are very low, but have been shown to be translationally active. By optimizing mRNA extraction and using real time (RT)-PCR we were able to establish a protocol for highly sensitive platelet mRNA quantification in human regular blood samples. In platelets from healthy volunteers, only P2X 1 , P2Y 1 and P2Y 12 were found in significant levels, with the following order of expression: P2Y 12 >> P2X 1 > P2Y 1 . Other P2 receptors (P2Y 2 , P2Y 4 , P2Y 6 , P2Y 11 , P2Y 13 , P2X 4 , P2X 7 ) had very low expression. As a control measurement to exclude contamination, P2 receptors in buffy coat were quantified but had a completely different profile. Incubation in vitro revealed a more rapid degradation rate for P2X 1 receptor mRNA than for P2Y 1 and P2Y 12 , indicating that the level of P2X 1 may be relatively higher in newly released platelets and in megacaryocytes. In conclusion, we have developed the first protocol for quantifying mRNA expression in human platelets limiting the P2 receptor drug development targets to P2Y 12 , P2Y 1 and P2X 1 . Furthermore, the method could be used to study platelet expression for any gene in human materials.Keywords: P2X receptor, P2Y receptor, platelets, real time-PCR, Western blot.Platelets play a crucial role in the maintenance of normal hemostasis and are involved in the development of pathological thrombus formation leading to vascular occlusion which is an important mechanism in myocardial infarction and stroke [1,2]. A large number of endogenous mediators can activate platelets, such as thromboxane A2, adenosine diphosphate (ADP), collagen, von Willebrand factor, thrombin, epinephrine and 5-hydroxitryptamine (5-HT). To get full activation of the platelets, these agonists are dependent on two positive feedback loops: the formation of thromboxane A2 by cyclooxygenase in the platelets and the release of ADP from dense platelet granules. Thromboxane A2 and ADP then activates specific receptors on the extracellular side of the platelet membrane.Therapeutic intervention aimed at the first positive feedback loop by inhibiting cyclooxygenase with aspirin is highly efficient in reducing death and cardiovascular events. However, ADP may be even more important as evidenced by the CAPRIE study, in which the ADP receptor inhibition was more beneficial than aspirin in reducing cardiovascular events [3,4].Recently, progress has been made in the understanding of the mechanisms of ADP mediated platelet aggregation, where at least three receptors are considered to be involved; the P2Y 12 , P2Y 1 , and P2X 1 receptors [5]. The importance of the P2Y 12 receptor is proven by the effects of clopidogrel, which after metabolization in the liver, acts as an irreversible antagonist at P2Y 12 receptors. Knockout of the P2Y 1 receptor in mice has demonstrated its importance for thrombus formation, blee...

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Section: Discussionmentioning

confidence: 99%

Quantification of ADP and ATP receptor expression in human platelets

Wang

Östberg

Wihlborg

et al. 2003

Journal of Thrombosis and Haemostasis

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“…Several other pseudopod-enriched proteins have also been reported to be associated with cancer including COTL1, YWHAE, ATP synthase, TPM4, SET, PTMA, and CNN2 (33)(34)(35)(36)(37)(38). Importantly, many of the identified proteins (Table 1), such as COTL1, YWHAE, TPM4, CNN2, septin 9, AHNAK, S100A11, and eIF4E, have well-defined associations with the actin cytoskeleton (39)(40)(41)(42)(43)(44)(45). In addition, we also identified five ribosomal proteins (RPL11, RPL23, RPL6A, RPL13, and RPL27) that, together with pseudopodial enrichment of eIF4E, elongation factor α (46), and various other proteins associated with RNA translocation and protein translation (19), further supports actin-rich pseudopodia as sites of active protein translation.…”

Section: Discussionmentioning

confidence: 99%

Pseudopodial Actin Dynamics Control Epithelial-Mesenchymal Transition in Metastatic Cancer Cells

et al. 2010

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A key cellular process associated with the invasive or metastatic program in many cancers is the transformation of epithelial cells toward a mesenchymal state, a process called epithelial to mesenchymal transition or EMT. Actin-dependent protrusion of cell pseudopodia is a critical element of mesenchymal cell migration and therefore of cancer metastasis. However, whether EMT occurs in human cancers and, in particular, whether it is a prerequisite for tumor cell invasion and metastasis, remains a subject of debate. Microarray and proteomic analysis of actin-rich pseudopodia from six metastatic human tumor cell lines identified 384 mRNAs and 64 proteins common to the pseudopodia of six metastatic human tumor cell lines of various cancer origins leading to the characterization of 19 common pseudopod-specific proteins. Four of these (AHNAK, septin-9, eIF4E, and S100A11) are shown to be essential for pseudopod protrusion and tumor cell migration and invasion. Knockdown of each of these proteins in metastatic cells resulted in reduced actin cytoskeleton dynamics and induction of mesenchymal-epithelial transition (MET) that could be prevented by the stabilization of the actin cytoskeleton. Actin-dependent pseudopodial protrusion and tumor cell migration are therefore determinants of EMT. Protein regulators of pseudopodial actin dynamics may represent unique molecular targets to induce MET and thereby inhibit the metastatic potential of tumor cells.Cancer Res; 70(9); 3780-90. ©2010 AACR.

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“…The platelet "secretome" is derived from intracellular storage granules, eicosanoid and phospholipid synthesis (47), and, as recently recognized, synthesis of proteins from constitutive mRNAs (48)(49)(50). It is now clear that platelets synthesize biologically relevant proteins in response to physiological stimuli that are regulated via gene expression programs at the translational level (51).…”

Section: Platelets Synthesize Biologically Active Proteinsmentioning

confidence: 99%

Platelets in inflammation and atherogenesis

Gawaz

Langer²,

May³

2005

Journal of Clinical Investigation

1,227

984

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Platelets represent an important linkage between inflammation, thrombosis, and atherogenesis. Inflammation is characterized by interactions among platelets, leukocytes, and ECs. These interactions trigger autocrine and paracrine activation processes that lead to leukocyte recruitment into the vascular wall. Platelet-induced chronic inflammatory processes at the vascular wall result in development of atherosclerotic lesions and atherothrombosis. This Review highlights the molecular machinery and inflammatory pathways used by platelets to initiate and accelerate atherothrombosis. Platelet interaction with endotheliumAt the site of vascular lesions, ECM proteins such as vWF and collagen are exposed to the blood. Platelet adhesion to the exposed matrix is considered to be the initial step in thrombus formation. Platelets adhere to vWF via the membrane adhesion receptor glycoprotein Ib/IX/V (GPIb/IX/V) (1) and to collagen via GPVI (2-4). This results in platelet activation and transformation of the integrin receptors α IIb β 3 (GPIIb/IIIa, fibrinogen receptor) (5, 6) and α 2 β 1 (collagen receptor) (4, 7), which firmly bind to the respective ECM components. Subsequently, platelets spread and form a surface for the recruitment of additional platelets via fibrinogen bridges between 2 α IIb β 3 receptors.In vitro studies with human ECs. In recent years, however, it has become increasingly evident that endothelial denudation is not an absolute prerequisite to allow platelet attachment to the arterial wall. The intact, nonactivated endothelium normally prevents platelet adhesion to the ECM. The adhesion receptors involved in platelet attachment to the subendothelial matrix, e.g., following rupture of an atherosclerotic plaque, have been well defined during the past decade; however, the molecular determinants that promote the interaction between platelets and endothelium are incompletely understood. Whereas endothelium normally controls platelet reactivity through inhibitory and modulating mechanisms involving COX-2, PGI 2 , or prostanoid synthetic systems, inflamed ECs develop properties that render them adhesive for platelets. In vitro studies showed that platelets adhere to the intact but activated human EC monolayer (8-10). Platelet adhesion to human umbilical vein ECs (HUVECs) is mediated by a GPIIb/IIIa-dependent bridging mechanism involving platelet-bound fibrinogen, fibronectin, and vWF (10). In HUVECs infected with herpes virus or stimulated with IL-1, platelet adhesion was effectively inhibited by antibodies to vWF or α IIb β 3 integrin, respectively (9,11,12). Furthermore, the involvement of the EC receptors ICAM-1, α v β 3 integrin, and GPIb in the binding of activated platelets to HUVECs has been described in vitro (10).In vivo studies of mouse models. Most in vitro studies have evaluated platelet-endothelium adhesion to human ECs under static conditions with limited attention to the dynamic situation in vivo. Studies using intravital microscopy, however, confirmed that platelet-endothelium adhesion occurs eve...

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Integrins Regulate the Intracellular Distribution of Eukaryotic Initiation Factor 4E in Platelets

Cited by 103 publications

References 24 publications

Quantification of ADP and ATP receptor expression in human platelets

Quantification of ADP and ATP receptor expression in human platelets

Pseudopodial Actin Dynamics Control Epithelial-Mesenchymal Transition in Metastatic Cancer Cells

Platelets in inflammation and atherogenesis

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